Preface

The force of natural selection tightly checks the adaptability of any living population to environmental conditions. On the other hand, the evolutionary success of a species in the phylogenetic path is measured by its flexibility against changing conditions. Like other intracellular components, peroxisomes also play an important role in the adaptation of the organism to its surrounding conditions. In eukaryotic organisms, the peroxisome, enclosed by a single membrane, is indispensable for lipid metabolism and free radical detoxification. The importance of peroxisome can be better grasped when the absolute harmful effects of free radicals as well as their newly discovered roles in intracellular signaling are evaluated holistically.

Despite extensive and rigorous scientific research, satisfactory information about the evolutionary origin of peroxisomes has yet to be elucidated. According to a theorybased argument, the ancestor of the peroxisome, which has an endosymbiotic origin, is a hydrogen-producing anaerobic prokaryotic organism that lost its DNA long ago or migrated to the host cell and dedicated itself to the endosymbiotic partnership with the eukaryotic cell earlier than the mitochondria and chloroplast. As evidence of the endosymbiotic origin of the peroxisome, the phylum Apicomplexa can be cited as a good example because this parasitic unicellular Sporozoa group contains mitochondria but not peroxisomes. However, the molecular composition and structural architecture of the peroxisomal membrane show homology to eukaryotic membranes. In addition, the molecular mobilization processes observed in the peroxisomal matrix protein import system and endoplasmic reticulum-associated protein degradation are interestingly similar.

Many diseases are related to peroxisome-induced free radical damage and intracellular signaling defect. As such, this book discusses prevention and sustainable treatment approaches for these diseases. Some of the highlights of the book are summarized below.

Disturbances in the reactive oxygen species (ROS) removal cascade of peroxisomes have important consequences for cell activities. The environment in which the cell interacts can either increase or decrease ROS damage. Today, the direct and indirect effects of natural and herbal supplements on the health of the organism are examined in a wide range. Communication disorders between the peroxisome and mitochondria can cause the oxidative balance to deteriorate in favor of reactive species. Peroxisomal dysfunction severely affects the morphological and molecular stability of the cell, resulting in apoptosis. Increased apoptosis is associated with many metabolic diseases that are often very difficult or impossible to treat. Molecules called peroxisome proliferator-activated receptors (PPARs) are nuclear receptor proteins that regulate the expression of genes that have important roles in many cellular processes, including tumorigenesis, as transcription factors. There are three subtypes of PPARs (PPARalpha, PPAR-beta/delta, and PPAR-gamma) found to be effective in the control of cancer metabolism steps. Disproportionate expression of any of these PPARs in the

organism may lead to cell proliferation and abnormal survival (immortality) in various types of cancer. These molecules thus provide a potential alternative therapeutic possibility as both an agonist and antagonist target for cancer therapy. In this context, some natural supplements such as multifunctional PPAR modulators are likely to provide important instruments in the therapy of various types of cancer. Several recent studies have demonstrated that various phytochemicals, including phenolics, are associated with PPARs-mediated anticarcinogenicity. Considering the proposed mechanisms, it is envisaged that plant phenolics have both palliative and therapeutic opportunities in the fight against cancer. The ghrelin molecule is a hunger-stimulating peptide hormone produced by the stomach and pancreatic cells, and its levels rise before meals and then fall. This hormone has been shown to reduce food intake and body weight in rodents. Preproghrelin, the immature (unprocessed) form of ghrelin, is likely cleaved by a protease to synthesize a regulatory peptide, obestatin. One of the N-terminal fragments (Nt8U) of three different obestatins synthesized in the laboratory was found to be as active as obestatin in the experiments performed on Swiss albino mice. Obestatin and its fragment analog Nt8U are reported to upregulate glycerolipid metabolism and PPAR-gamma signaling and reduce fat accumulation in Swiss albino mice. Soon, computer-aided computational methods and advances in data processing analysis will be more widely used to answer the remaining questions about peroxisome regulation, mechanism, function, and its biogenesis in the context of healthy and pathological phenotypes. It is thought that advanced computational methods for the analysis of imaging data, protein structural modeling, proteomics, and genomics will come forward thanks to artificial intelligence (AI) such as machine learning and deep learning.

This book highlights the role of peroxisomes in the antioxidant mechanism as well as in the synthesis, maturation, and homeostatic regulation of some vital molecules. It also discusses diseases in humans caused by disruptions in peroxisome function and the modulatory effects of natural exogenous supplements on peroxisome activity.

> **Dr. Hasan Basri İla** Professor, Faculty of Science and Letter, Department of Biology, Cukurova University, Adana, Turkey

Section 1 Introduction

#### **Chapter 1**

## Introductory Chapter: Peroxisome-Virus Interaction from SARS-CoV-2 Perspective

*Hasan Basri İla*

#### **1. Introduction**

Shannon Butalla and Melissa Gamble, parents of children affected by the peroxisomal disorder, took concrete steps in 2010 to establish the "Global Peroxisomal Disorders Foundation (GFPD)." The foundation has become an effective and complementary center providing research and family support for patients and their relatives struggling with rare peroxisomal diseases in the community. Peroxisomes, which are active in almost all our cells under normal conditions, are organelles that take an active role in lipid metabolism and render specific biochemical toxins harmless. Sabotaging the peroxisomes, which have supercritical initiatives for cell homeostasis, has irreparable consequences. Peroxisomal disorders include single-enzyme defects that affect a specific biochemical pathway and/or biogenesis defects that affect the entire peroxisome and therefore the organism. Many patients with peroxisome biogenesis defect (PBD) simultaneously lack the multiple functions of the peroxisome. These patients show a clinical gradient termed peroxisome biogenesis disorders-Zellweger Spectrum Disorders (PBD-ZSD). PBD-ZSD symptoms manifest in a wide range of patients. The disease symptoms can vary from mild as progressive hearing and vision problems after childhood to moderate-/high-level diseases including feeding problems, low muscle tone, and brain and liver disorders since infancy. Many patients with severely affected PBD-ZSD present with multiple medical problems, persistent seizures, developmental abnormalities, and even death in the neonatal period [1].

In patients with PBD-ZSD, autosomal recessive mutations have been found in a group of peroxisomal genes called peroxin (PEX), which plays a role in peroxisome biogenesis. In a typical PBD-ZSD family, each of the parents may be a carrier of a mutation in a gene such as PEX1. For example, the mutant PEX1 gene in both carrier parents does not affect the health of the carrier and does not produce severe disease for them. This is because there are two copies (diploidy) of each gene in each individual's genome, including PEX1. A healthy second copy can protect a carrier from the disease, but two carrier parents for the same gene have a 25% chance of transmitting the mutation to their children in each pregnancy. Because of their importance for peroxisome, the functions of PEX genes and their products, of which 37 have been discovered so far, are being studied intensively [2].

#### **2. Peroxisome virus interaction**

#### **2.1 Pandemic declaration and SARS-CoV-2**

On March 11, 2020, it was recognized by the World Health Organization (WHO) that the epidemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (later named COVID-19) has become a pandemic. As of July 10, 2022, around 553 million confirmed cases and more than 6.3 million deaths have been reported worldwide due to COVID-19 [3, 4].

SARS-CoV-2 is a new strain of the coronavirus family that triggers and deepens health, social, and economic crises worldwide. Considering the effects of this virus on the organism, its interaction with the peroxisome is critical. SARS-CoV-2 infection in some human cell lines (Huh-7 and SK-N-SH) causes striking changes in peroxisome morphology. Since the peroxisomal membrane integrity is affected by the virus, peroxisomal matrix proteins leak into the cytosol and a marked reduction in the number of mature peroxisomes in the infected cells. Some proteins encoded by ORF14 of SARS-CoV-2 interact physically with human PEX14, which act as a membrane protein during matrix protein import and peroxisome biogenesis. Given the role of peroxisomes in innate immunity, the argument is getting stronger that SARS-CoV-2 can either directly target peroxisomes or is dangerous because it targets them. Therefore, a viral attack can cause the loss of function in the structural integrity, matrix protein content, and antiviral signaling of the peroxisome [5].

#### **2.2 Interactions of some viruses with peroxisomes**

A newly discovered mechanism in human immunodeficiency virus (HIV) patients (AIDS) has offered hope as a different approach for treating COVID-19. More recently, the steps of HIV attack on peroxisomes, which regulate many cellular mechanisms, including the immune system, have been demonstrated. It has been demonstrated that some viruses, including West Nile virus (WNV) and ZIKA virus (ZIKV), have different instruments to inhibit the host-cell interferon response, such as reducing peroxisomes. In fact, all viruses have ways of blocking the host-interferon response. This situation is consistent with the concept that peroxisomes are an important target for viral infections. At this point, researchers evaluated that SARS-CoV-2 could also attack peroxisomes and started testing drugs that increase peroxisome activity against the virus in cell cultures. Recently, a group of scientists reported the mechanistic basis for why many HIV patients suffer from premature aging, lipodystrophy (change in the body fat metabolism cascade), and several diseases. The team identified four microRNAs with increased expression in the brains of HIV patients with dementia. All four microRNAs increased in the brains of these patients appeared to target the peroxisome biogenesis pathway. These increased expression molecules downregulate protein expression required to form peroxisomes. On the other hand, it has been shown that increased peroxisomes by manipulating appropriate genes can inhibit ZIKV replication. Tests of drugs that increase peroxisomes are ongoing to evaluate the success against SARS-CoV-2 and HIV. Many of the agents that researchers have tried have been approved as cancer drugs. During these investigations, it was discovered that a pathway that inhibits peroxisome formation was targeted. Despite possible indications for drug use, it is noted that some peroxisome boosters can be taken orally and have low side effects [6].

#### **3. The plasmalogen importance for viral defense**

The cellular functions of plasmalogens, which are abundant in many mammalian tissues, have not yet been fully elucidated. However, it is evaluated that they can protect cells against the harmful effects of endogenous reactive oxygen species. It is also thought to act as signaling molecules and modulators in the membrane [7–9]. Plasmalogen biosynthesis requires functional peroxisomes that are oxidative organelles in the cell. Peroxisomes are important host organelles where certain viral replication can take place [10]. Plasmalogen deficiency observed in cardiometabolic and multiple neurodegenerative diseases may predispose humans to SARS-CoV-2 and other similar viral infections. On the other hand, increased plasmalogen levels were detected in humans infected with some viruses (ZIKV, HBV, and HIV) and in the virion lipidome of cytomegalovirus [11–14]. Increased plasmalogen levels revealed a strong association between the ZIKV lifecycle and host peroxisomes. This finding scientifically explains the upregulation of resident plasmalogens and peroxisomemediated lipidome changes in the serum of patients infected with ZIKV [14, 15]. Meanwhile, it has been reported that the plasmalogen phosphatidylcholine molecule plays an important role in influenza virus infection [16].

The infection response of the host cell can be used as the basic determinant in the pathogenesis of any infectious disease, including the COVID-19 pandemic. The lipid composition of the host cell plasma membrane has a decisive role in the life cycle of a virus, because the first step of the infection process is the entry of the virus into the cell by crossing the membrane. In viral infections in a cell, peroxisomes act as vital immune signaling foci and assist the host by regulating antiviral signaling [10]. Peroxisomes are critical organelles that act as a double-edged sword in the viral infection process. Since this cellular organelle, which both harbors and kills the pathogen, has a primary role in host antiviral defense, it can transform into a useful apparatus that serves viral replication through critical manipulations [17]. This interaction in favor of the virus brings to mind the famous aphorism of nineteenth-century German philosopher Friedrich Nietzsche "What doesn't kill me, makes me stronger."

With the discovery of mitochondrial antiviral signaling (MAVS), an innate host immune response, mitochondria were recognized as an important subcellular signaling center. Initially, MAVS, which produced a rapid antiviral reaction, was thought to be specific to mitochondria only. However, considering the known roles of peroxisomes in detoxification, the identification of peroxisomal MAVS (or PAVS) has revealed the importance of peroxisome in host defense as an antiviral signaling organelle. This finding was supported by the discovery of increased host peroxisome biogenesis during human cytomegalovirus (HCMV) and herpes simplex type 1 (HSV-1) infections [18]. The synthesis of some vital molecules, which are essential for viral penetration and the success of the virus life cycle, only in the peroxisome is the most important link in the chain. These molecules are plasmalogens and some cellular lipids such as docosahexaenoic acid, a very long chain omega-3 fatty acid. The units that synthesize plasmalogen and some lipids are peroxisomes, which are indispensable for the construction of viral envelopes, modulation of host cholesterol homeostasis, and maintenance of virus-host balance during infection. It is therefore not surprising that peroxisomes are an attractive candidate for cellular remodeling during some viral infections. Consistent with this proposition, it has been suggested that increased plasmalogen in HCMV virions [12] and alterations in peroxisomal lipid metabolism may be a general characteristic of enveloped virus infections. Plasmalogen and some lipids are essential components of many enveloped viruses, including HCMV and

influenza. Therefore, it is considered that this argument may also be valid for SARS-CoV-2. However, further lipidomic analysis is required in COVID-19 samples for a more detailed projection [19].

Macrophages, professional phagocytes of the host immune system, can detect and clear invading pathogens such as viruses and damaged cells. Plasmalogen deficiency in macrophages is associated with a reduced ability to phagocytosis. This situation is significantly reversed when cells are exposed to lysophosphatidylethanolamine plasmalogen [20]. Similarly, restoration of plasmalogen levels leads to increases in the number and size of lipid microdomains in the membranes of macrophages. Therefore, exogenous plasmalogen administration is likely to be adopted as an innovative strategy for optimizing macrophage function [19]. According to the results of a comprehensive bioinformatics study on macrophage differentiation, the plasmalogen phosphatidylethanolamine (PE) molecule is a biomarker of immune system activation. As an interesting finding, a significant decrease in plasmalogen levels was observed in obese subjects [21]. In light of these findings, the potential link between host plasmalogen dysregulation and the high morbidity and mortality levels observed in COVID-19 patients is considered significant. Again, a strong correlation is observed between decreased plasmalogen levels and a number of pathological conditions, including neurodegenerative and cardiometabolic disorders, as well as severe COVID due to coronavirus infection. In coronavirus-induced lipidome patterns, irregular plasmalogen levels in the infected patients are of interest. This finding indicates that the plasmalogen molecule is among the key lipids in potentially modulating viral infection [19].

#### **4. The host cell reprogramming by SARS-CoV-2**

SARS-CoV-2 replication alters the morphology, number, and function of many cellular structures. Peroxisome accumulation was observed in regions containing double-membrane vesicle (DMV) clusters formed by the endoplasmic reticulum (ER). This finding was confirmed by detecting a marked increase in peroxisomeassociated protein, PMP70, in infected cells by confocal microscopy and Western blot analysis. The convergence between the peroxisome and the viral RNA replication site serves to protect viral RNA from oxidative damage [10] or to establish a signaling platform that generates a cytokine response [22].

It has been discovered that many cellular components other than the ER and peroxisomes are also remodeled in cells infected with SARS-CoV-2. It is noteworthy that this transformation is accompanied by decreased mitochondrial ATP synthase, swollen cristae, and matrix condensation. The related observation is consistent with reduced oxidative phosphorylation in cells infected with SARS-CoV-2 and transcriptional changes pointing to virus-induced metabolic reprogramming [23]. The virus in the infected cell manages dynamic changes in the biosynthesis and utilization of macromolecules such as glucose, nucleotides, fatty acids, and amino acids to expand the maneuvering space. These changes are directly related to the success of viral infection [24].

#### **5. Inference**

As can be seen from the results of the studies discussed above, there is a multilayered, dynamic, and complex interaction between peroxisome health and the virus life

#### *Introductory Chapter: Peroxisome-Virus Interaction from SARS-CoV-2 Perspective DOI: http://dx.doi.org/10.5772/intechopen.106925*

cycle. The coincidental and rapid evolution of viruses in the direction of manipulating the peroxisomal pathway at any stage during the infection process may lead to dead ends or irreparable damage to the host cell defense system. In general, the possibility that extraordinary viruses such as SARS-CoV-2 may favor people with fragile peroxisomes for genetic or environmental reasons should be seriously considered. In this context, handling new insights such as peroxisome health in the control of viral pandemics will provide an important achievement in the fight against the disease.

### **Author details**

Hasan Basri İla Faculty of Science and Letter, Department of Biology, Çukurova University, Adana, Turkey

\*Address all correspondence to: ila@cu.edu.tr; hasanbasriila@gmail.com

© 2022 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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[12] Liu STH, Sharon-Friling R, Ivanova P, Milne SB, Myers DS, Rabinowitz JD, et al. Synaptic vesicle-like lipidome of human cytomegalovirus virions reveals a role for SNARE machinery in virion egress. Proceedings of the National Academy of Sciences of the United States of America. 2011;**108**(31):12869-12874

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## **Chapter 2** The Mystery of Peroxisomes

*Hasan Basri İla*

#### **Abstract**

According to the evolutionary perspective, an organism must manage and optimize organized complexity effectively to achieve a strong adaptation. Within the scope of sustainable homeostasis, the subcellular components of the organism must strictly comply with the principle of minimum error and maximum efficiency in coordination. Advanced defense systems are evolution's greatest gift to the cell. One of the most important components of cellular defense systems is the antioxidant defense. When it comes to antioxidant defense, the first thing that comes to mind is the peroxisome organelle, because the peroxisome is a cytoplasmic organelle surrounded by a single membrane in which the very important enzyme, catalase, is localized. Furthermore, the role of this organelle in vital processes, such as lipid metabolism, antimicrobial defense, and intracellular signaling, is undeniable. In this chapter, attention has been tried on the mysteries related to peroxisome by performing a wide literature review. The chapter covers topics such as peroxisome production, targeted protein transport, roles in the oxidative mechanism, relationship with diseases, and mitochondria interaction. This chapter, which highlights the polygenic formation and pleiotropic features of peroxisome, will provide an important future projection for curious researchers and medical doctors seeking innovative treatment strategies.

**Keywords:** peroxisome, oxidative stress, mitochondria, protein import, cell differentiation, cancer

#### **1. Introduction**

The major oxidation event worldwide occurred about 2.45 billion (B) years ago. However, there is heated debate that an earlier and failed oxygenation event may have occurred about 3.2 B years ago. However, the absence of sedimentary rock deposits stained red with iron oxide in stratigraphic units older than 2.5–2.0 B years is considered to be the most convincing evidence of non-oxygenation [1]. The study of sulfur assets on earth shows that between 2.45 and 2.09 B years ago a change occurred in the sulfur cycle. Atmospheric reactions at that time and the partial pressure of atmospheric oxygen (O2) also played a role in determining the oxidation state of sulfur in the earth's crust. The findings indicate that the atmospheric oxygen partial pressures were low at that time and their roles in oxidative decomposition, microbial oxidation, and sulfur reduction were minimal [2].

According to the consensus accepted by the majority of scientists, the axis of the evolution of living things changed markedly as atmospheric oxygen started to increase from 2.5 to 2 B years ago. Since the first atmospheric oxygen appeared, there have been drastic fluctuations in oxygen concentration. After a significant increase 375–275 million years ago, it decreased slightly and reached today's level (21%) [3]. Mitochondria allowed the development of eukaryotes, thanks to the exploitation of this dangerous and highly reactive element, O2 (oxygen respiration). Reactive oxygen species (ROS), such as superoxide, hydrogen peroxide, and hydroxyl (O2·−, H2O2, and •OH), some of which are quite dangerous, have widely entered cellular life due to the oxidation and reduction (redox) reactions that take place in aerobic respiration. In addition to being a reactive species, H2O2, one of these ROS, also functions as a signal molecule in the intracellular metabolic pathways [4].

The primary source of H2O2 production in the cell is mitochondria [5]. Complex I, one of the components of the electron transport chain in mitochondria, firstly causes the production of superoxide (O2·−), which is then converted to H2O2 by being dismutated by manganese superoxide dismutase (Mn-SOD) induced by NF-κB activation [4]. Peroxisomes, which play a central role in lipid metabolism, owe their name to activities that produce and scavenge hydrogen peroxide and are eukaryotic organs that also play key roles in the conversion of reactive oxygen species. Peroxisomes are responsible for the catabolism of very-long-chain fatty acids, branched-chain fatty acids, bile acid intermediates, D-amino acids, and polyamines, and the reduction of reactive oxygen species, especially hydrogen peroxide [6]. In addition, peroxisomes play a role in the biosynthesis of ether phospholipid plasmalogens, which are critical for the normal function of organs such as the mammalian brain, heart, and lungs. In humans, plasmalogens constitute approximately 18% of the total phospholipid mass and show a cell- and tissue-specific distribution [7]. Plasmalogens (1–0-alk-1′-enyl-2-acyl glycerophospholipids) constitute a special class of phospholipids characterized by the presence of a vinyl-ether bond at the sn-1 position [8].

In addition to the presence in the cytosol of two enzymes in the pentose phosphate pathway (glucose-6-phosphate dehydrogenase and 6-Phosphogluconate dehydrogenase), which are important for energy metabolism (NADPH), the same enzymes are also found in intact peroxisomes secretly and a part of the total activity is associated with these enzymes located peroxisomal [9]. This finding is an indication of the tight relationship and strong interaction between peroxisome and other subcellular compartments.

#### **2. Peroxisome**

Peroxisomes were identified in 1960 as part of the pioneering work of Christian René de Duve, who developed cell lysis techniques. De Duve's method separates organelles according to their precipitation and density characteristics, and it has been determined that peroxisomes are more intensive in the cell than other organelles [10]. Peroxisomes, which are versatile (pleiotropic) organelles, constantly adapt (plasticity) to current environmental conditions. The number of this organelle is rapidly adjusted in response to cyclical changes or various stimuli to maintain a steady state in the cell. Although the mystery of peroxisomes is still not fully resolved, it can be generalized that it is an organelle involved in (anti)oxidation and some synthesis pathways. Although morphologically similar to lysosomes, the structure is matured by the transport of proteins synthesized in free ribosomes and then functionalized to

peroxisomes. At this point, the debate over whether peroxisomes proliferate by division or by *de novo* synthesis draws attention. Peroxisomes do not have their genome but replicate similar to mitochondria and chloroplasts. Peroxisomes contain at least 50 different enzymes involved in various biochemical pathways in different cell types. Initially, they were described as organelles that carry out oxidation reactions that lead to the production of hydrogen peroxide. On the other hand, since hydrogen peroxide is harmful to the cell, it is vital to render it harmless. For this purpose, the peroxisome either breaks down hydrogen peroxide directly into water and oxygen via its catalase enzyme.

$$2\text{H}\_2\text{O}\_2 \Rightarrow 2\text{H}\_2\text{O} + \text{O}\_2 \tag{1}$$

Various substrates, including uric acid, amino acids, and fatty acids, are broken down by oxidative reactions that take place in peroxisomes. The oxidation of fatty acids is an important example as it provides a great source of metabolic energy. Fatty acid oxidation in peroxisomes is accompanied by the production of hydrogen peroxide (H2O2). Although fatty acids are oxidized in both mitochondria and peroxisomes in animal cells, fatty acid oxidation in yeast and plants is limited to peroxisomes. In animal cells, cholesterol and dolichol are synthesized in peroxisomes as well as in the ER. Peroxisomes in the liver are also involved in the synthesis of bile acids derived from cholesterol. In addition, peroxisomes regulate sterol regulatory element-binding proteins (SREBPs) and SREBP cleavage-activating protein (SCAP) traffic that activates this protein. This regulation is of functional importance for maintaining cholesterol homeostasis and efficient cholesterol synthesis. In addition, peroxisomes contain enzymes necessary for the synthesis of plasmalogens, a member of the glycerophospholipid family. In the chemical structure of plasmalogen, one of the hydrocarbon chains is attached to glycerol with an ether bond instead of an ester bond. Although not widely observed in other tissues, plasmalogens are important membrane components in some tissue cells, especially in the heart and brain [11, 12].

#### **2.1 Glyoxysome**

Special peroxisomes found especially in seed plants as well as filamentous fungi are called glyoxysomes. Peroxisomes play two particularly important roles in plants. First, peroxisomes are responsible for metabolizing fatty acids stored in seeds into critical carbohydrates to provide growth energy and raw materials to the germinating plant. This process, called the glyoxylate cycle, is a variation of the citric acid cycle that occurs through a series of reactions. Second, peroxisomes in plant leaves are involved in the metabolism of the by-product formed in the photorespiration process [12].

#### **2.2 Reproduction and growth of peroxisomes**

As noted earlier, the peroxisome is similar in form to mitochondria and chloroplasts in terms of some fundamental metabolism. Structural and enzymatic proteins of peroxisomes are synthesized by cytosolic free ribosomes (sometimes ER polysome) and shipped specifically to the peroxisome. Similarly, phospholipids are transferred from major synthesis sites in the ER to peroxisomes via phospholipid transfer proteins (PLTP). With the import of protein and phospholipids, peroxisomes grow and then divide to form new peroxisomes.

Thirty-seven peroxisome biogenesis factor proteins (known as Peroxin or PEX) that are involved in peroxisome biogenesis and proliferation identified so far are encoded by PEX genes. However, not all eukaryotes contain peroxisomes, so there are several PEX proteins involved in possible alternative processes in organisms lacking peroxisomes. Initial data revealed that several protist species (*Cryptosporidium parvum, Theileria annulata, Babesia bovis, Monosiga brevicollis, Plasmodium falciparum, Blastocystis hominis,* and *Entamoeba histolytica*) lack most PEX proteins [13]. Some eukaryotes, such as anaerobic protists, plasmodium, and parasitic platyhelminths, lost their peroxisomes in the evolutionary process. Surprisingly, *Oikopleura dioica*, a free-living pelagic tunic in the oxygen-containing niches of marine waters, contains no peroxisomal gene in its genome. It was also found that the putative peroxisomal enzyme set was considerably reduced and none contained a predicted peroxisomal targeting signal (PTS). It has been shown that several metazoan lineages, such as *O. dioica*, independently lost peroxisomes showing that peroxisome loss is not only related to adaptation to anaerobic habitats and/or parasitic lifestyle [14].

PEX genes are required for peroxisome growth and division. Overexpression of the PEX11 gene causes peroxisome proliferation, while its deletion leads to the enlargement and reduction of peroxisomes. While Pex11 causes membrane tubulation *in vitro*, the cytoskeleton also contributes to this situation likely [15].

#### **2.3 Protein transport into the peroxisome**

According to the peroxisome database, a complete peroxisomal proteome is encoded by 61 genes in *Saccharomyces cerevisiae* and 85 genes in *Homo sapiens* [16]. Targeting of proteins into the peroxisome is achieved by at least two conserved pathways from yeasts to humans. Most proteins are targeted to peroxisomes by the simple amino acid sequence (Ser-Lys-Leu tripeptide) at their carboxy terminus (peroxisome targeting signal 1 or PTS1) [12]. However, there is a different source suggesting that amino acids in this signal (PTS1) show polymorphism as (Cys/Ala/Ser)-(Lys/Arg/His)- (Leu/Ile) [17]. Other peroxisomal proteins are targeted to the organelle by another signal peptide (PTS2) located at the amino terminus. Although there are multiple variants of PTS2, it consists of the nine amino acid consensus nonapeptide [(-Arg/Lys) (Leu/Val/Ile) XXXXX (His/Gln) (Leu/Ala)] sequence (X in this sequence represents any amino acid) showing amino acid polymorphism. However, some proteins may be targeted by alternative signals that are not yet well-defined [18]. PTS1 and PTS2 are recognized by different receptors and then transferred to a translocation complex in the peroxisome membrane. Cytosolic heat shock protein 70 (Hsp70) has been associated with protein import into peroxisomes, but the possible role of molecular chaperones in peroxisomes is unclear [12]. According to new data, Hsp70s mediates the delivery of components such as the ubiquitin E3 ligase Stub1 to the damaged peroxisome. In this way, peroxisome may promote autophagic degradation (pexophagy) of itself due to oxidative stress. In summary, Hsp70s are involved in the negative regulation of peroxisomes [19].

Three receptors, namely Pex5p, Pex7p, and Pex19p have been identified so far that recognize various types of PTS ligands. Pex5p functions as a cyclic transport receptor for proteins carrying newly synthesized PTS1 or not carrying any targeting signals. Pex5p recognizes and binds proteins to be transported in the cytosol and transports them to the peroxisomal matrix. This process is based on complex and transient protein-protein interactions involving cargo recognition, cargo-loaded receptor docking to the peroxisomal membrane, cargo translocation, and release. To fulfill

these functions, all Pex5p orthologs contain multiple flexible segments. It contains two clusters of three tetratricopeptide repeat (TPR) motifs in the C-terminal part of the molecule, connected by a flexible joint region. This construct creates a single binding site for PTS1. The N-terminal part of the molecule has multiple diaromatic pentapeptide motifs (usually expressed as TrpXXXPhe/Tyr). The number of these peptide motifs varies between species, with two in *S. cerevisiae* and nine in *Arabidopsis thaliana*. Duly, both the C- and N-terminal domains of Pex5p undergo significant conformational transitions after binding to their ligands.

The Pex7p molecule is a soluble protein that serves as a targeting signal recognition factor for newly synthesized PTS2 proteins. It exhibits a cytosolic and intraperoxisomal distribution pattern and can be repeatedly translocated inside and outside of the organelle. All Pex7p orthologs are characterized by the presence of six WD401 repeat motifs, which together with a distinct N-terminal region are predicted to form a seven-bladed β-propeller-like structure. Mutations that affect the conformation of this structure almost always abolish activity. PTS2's receptor, Pex7p, recruits cofactors for its function in peroxisomal protein uptake. These co-factors termed "PTS2 coreceptors," are species-specific and include an insertion form similar to Pex5pL, the long isoform of Pex5p in mammals. All these proteins are largely cytosolic and show a low overall similarity between their primer sequences.

The Pex19p molecule is a predominantly cytosolic, partially peroxisome localized multifunctional protein and plays a central role in the early steps of peroxisomal membrane synthesis. It has been determined that this Peroxin can (i) bind newly synthesized distinct peroxisome membrane proteins (PMP) in the cytosol, (ii) keep these PMPs in a competent conformation for attachment to the membrane, (iii) transport them to the peroxisomal membrane, and (iv) return to the cytosol with a shuttle-like movement. Because of this feature, the suggestion that Pex19p functions as a chaperone and soluble transport receptor for "class I" PMPs is strengthened. Members of this PMP class contain common Pex19p binding motifs that are an integral part of targeting signals (PTSs). Interestingly, many PMPs contain multiple Pex19p binding motifs, but not all of these regions can directly bind to PTS [20].

Some peroxisome membrane proteins are similarly synthesized by cytosolic ribosomes and targeted to the peroxisomal membrane by different internal signals. However, the expression of some peroxisomal membrane proteins in membrane-bound polysomes of the endoplasmic reticulum and their transport to peroxisomes point to the important role of the endoplasmic reticulum for peroxisome integrity [12].

Specific protein complexes exist that act as docking sites on the peroxisomal membrane for cargo-loaded PTS receptors. Currently, two complexes have been identified that perform this task: one recognizes the incoming Pex5p- and Pex7pcargo complexes, and the other recognizes the binding of the Pex19p-cargo complex. Here, it was determined that Pex5p Peroxin showed a higher affinity for Pex14p and the amount of peroxisome-associated Pex5p was proportional to the amount of Pex14p. In addition, it has long been known that in many species deficient in Pex3p, Pex16p, or Pex19p, the peroxisomal membrane structures of cells lack integrity. This observation led to the hypothesis that these Peroxins are essential for peroxisome membrane biogenesis. It has been shown that Pex8p from *Pichia pastoris*, a

<sup>1</sup> It is a short structural motif of about 40 amino acids, usually ending with a tryptophan-aspartic acid dipeptide. Successive copies of these repeats fold together to form a type of circular solenoid protein called the WD40 domain.

methylotrophic yeast species, requires only Pex5p and Pex14p for PTS1-dependent transfer to peroxisomes. It has been reported that the diameter of the ion-gated channel formed in studies with proteoliposomes may expand when it encounters cytosolic Pex5p-cargo complexes. The Pex5p/Pex14p proteins herein are regulatory elements of the matrix protein transition pore in the peroxisome membrane. In summary, there is currently substantial evidence that some PMPs migrate through the ER to peroxisomes, while others separate directly from the cytosol into these organelles. There is also no consensus on whether these processes require an energy source. While the interactions between PTS receptors Pex5p, Pex7p, and Pex19p and their cargo proteins are relatively well-characterized, little information is available about how the cargo, carrier, and receptor complexes are separated during the delivery of cargo to its destination. It was initially suggested that Pex8p, an intraperoxisomal protein containing both PTS1 and PTS2 signaling, may act as a PTS1 receptor-cargo release factor. However, since Pex8p is only found in fungi (such a mechanism is not active in higher eukaryotes), it has been suggested that dissociation of the Pex5p-PTS1 complex may be mediated by a change in pH. According to this hypothesis, Pex5p exists in different oligomeric conformations in the cell, and these structures change with pH. Pex5p is in the monomeric form at pH 6.0 and evolves into a tetrameric form at pH 7.2. While PTS1 peptides bind to tetrameric Pex5p predominantly under slightly alkaline conditions, due to the slight acidity of the peroxisomal matrix, the structure changes to monomeric conformation, resulting in the release of cargo. However, factors other than pH may also trigger the cargo release step at a higher level. The addition of single or multiple ubiquitin to conserved cysteine and lysine residues at the N-terminus of Pex5p, Pex18p, Pex20p, or Pex21p molecules can be expressed as a dissociation factor other than pH. Conjugation of ubiquitin-like molecules to a protein requires the concerted action of an ATP-requiring ubiquitinactivating enzyme (E1), a ubiquitin-conjugating enzyme (E2), and a ubiquitin-ligase (E3). It should be noted that mutations in any of these proteins will result in a defect in peroxisomal matrix protein import. For sustainable protein uptake (import) cycle, (ubiquitinated) PTS (co)receptors must be returned to the cytosol with (by) shuttle-like movement. Selective elimination of unnecessary and dysfunctional peroxisomes is required to regulate peroxisome function properly and limit damage during cellular aging. Biochemical and genetic studies in different organisms have shown that the degradation used to eliminate peroxisome can occur through at least three different mechanisms. These are macropexophagy, micropexophagy, and 15-lipoxygenase-mediated autolysis. More than 35 autophagy-related (ATG) genes have been identified to date. The proteins encoded by these genes, collectively called Atg protein genes, are essential for selective and nonselective autophagy pathways. Interestingly, all these pathways have been stated to require a core molecular machinery conserved from yeast to humans [20].

#### **3. Effect on oxidative stress**

Peroxisomes, which emerged as the product of the selective strategy of evolution seeking a way out, break down various substrates including amino acids, fatty acids, and uric acid in the cell, as is commonly known, through oxidative reactions, while reactive byproducts are also produced. Peroxisomes both render harmless the reactive products formed by redox reactions and are important actors in the biosynthesis of a phospholipid, dolichol, plasmalogen, cholesterol, and bile acid [12].

#### *The Mystery of Peroxisomes DOI: http://dx.doi.org/10.5772/intechopen.105063*

The radical scavenging function of peroxisome has been emphasized in the historical process from its discovery to the present. However, it produces radicals for various reasons in metabolic pathways, and then it renders them harmless in the cell. Reactive oxygen species (ROS) take an active role in the natural biosynthetic cascade, both as an intracellular signaling molecule and for the breaking or rebuilding of existing molecular bonds, due to their effect on electrical charge distribution [21, 22]. Interestingly, highly dangerous reactive molecules take an active mission in cell molecular architecture. On the other hand, the fact that peroxisomes contain antioxidant enzymes to eliminate the damage of radical species is related to the dual activity of the peroxisome. This reality brings to mind the struggle of Ahura Mazda - Ahriman in the Zoroastrian belief. Peroxisomes interact and cooperate closely with other cell organelles (mitochondria and chloroplasts) for the optimization of cellular homeostasis. In addition, due to the excellent plasticity of the peroxisomes, their number, shape, and protein contents are dynamically regulated in response to changing cellular and environmental conditions. Although the peroxisomal β-oxidation pathway is similar to the mitochondrial pathway, the related enzyme sets in the reaction are different. Unlike mitochondria, ATP is not released in the fatty acid breakdown in peroxisomes. The fatty acid is activated by the fatty acyl CoA synthetase in the first step in the degradation process. First, a double bond is added to the beta position of the fatty acyl-CoA ester. This step, catalyzed by FAD-containing acyl-CoA oxidase, generates hydrogen peroxide by transferring hydrogen atoms to molecular oxygen. Constitutive activation of peroxisome proliferator-activated receptor alpha (PPARα) occurs when fatty acids accumulate due to defective degradation. PPAR activation upregulates the levels of peroxisomal and mitochondrial β-oxidation enzymes that can lead to oxidative stress. In addition, complex forms of transition elements such as iron and copper are abundant in the peroxisome. Therefore, some toxic xenobiotics (pyrimidines, dialuric acid, divicine, isouramil, etc.) cause the iron release from iron-binding proteins such as ferritin. In the Fenton reaction, it has been shown that in the presence of ascorbate, histidine, or ADP, Fe2+ induces lipid peroxidation by breaking the H2O2 molecule into more dangerous hydroxyl (OH) radicals. On the other hand, peroxisome contains a number of antioxidant enzymes to prevent oxidative damage of hydrogen peroxide itself and free radicals derived from it. Hydrogen peroxide is cleared by catalase and glutathione peroxidase. Again, superoxide anions MnSOD and CuSOD are rendered harmless by converting them to hydrogen peroxide by Zn-SOD. Along with the last two enzymes, catalase and glutathione peroxidase are mainly found in mitochondria and cytosol, but these enzymes have also been reported to be present in peroxisomes. However, questions about how peroxisomal response signals are created have only recently begun to be answered. Located in the peroxisome membrane and a central component of the protein import process, Pex14 is first phosphorylated in mammalian cells in response to oxidative stresses such as H2O2. In this process, H2O2-induced phosphorylation of the Ser232 residue of Pex14 suppresses the import of catalase *in vivo*, and this phosphorylation selectively impairs the *in vitro* interaction of catalase with the Pex14-Pex5 complex. It has been found that this has an effect on cytosolic and peroxisomal catalase levels [23–25].

#### **4. Peroxisome-related illness or genetic disorders**

Enzymatic antioxidants in the peroxisome like catalase play important roles. Some *de facto* conditions in the enzymes and other auxiliary factors that take part in the

conversion of superoxide to the reduction stages to more stable molecules and the breakdown of H2O2 into water and oxygen will cause the creation of the •OH radical. The •OH, the hazardous premature product of H2O2 is a free radical that carries a serious risk for all organic molecules. It attacks all forms of molecules, including nucleic acids (DNA and RNA), and changes their molecular conformation. H2O2, which is also a known DNA disrupter, has taken its place in test protocols as a known mutagen in some genetic tests (e.g., Comet assay). There is no doubt the costs of the weaknesses that may arise in antioxidant enzymes will be heavy for the cell. However, a serious paradox stands here. Accordingly, do increased genetic defects (mutations) lead to oxidation, or do increased oxidations lead to mutations? This situation is similar to a popular Ancient Greek paradox, "*Which came first, the chicken or the egg*?" The undeniable fact is that increased ROS causes mutations, while increased mutations produce more ROS. It is like the Ouroboros2 phenomenon. Acatalasemia with inherited catalase activity deficiency was initially considered an asymptomatic disorder. Decreases in the extracellular hydrogen peroxide (H2O2) removal capacity of catalasedeficient tissues have been determined in animal models. In patients with catalase deficiency, H2O2 can cause methemoglobinemia3 . The high (18.5%) prevalence of diabetes and the onset of the disease 10 years earlier in individuals with hereditary catalase deficiency may be attributed to oxidative damage to oxidant-sensitive, insulin-producing pancreatic beta cells. Oxidative stress and aging-related diseases were diagnosed in 97 of 114 acatalasemias. Oxidative stress due to catalase deficiency alone may contribute to the onset of diabetes and may also be one of the causative factors of other diseases. Reactive species produced in the cell during normal metabolism enter into chemical reactions with cellular biomolecules such as nucleic acids, proteins, and lipids, causing their harmful oxidative modifications. It is assumed that catalase deficiency or its defective function, which occurs for any reason, is closely related to the pathogenesis of many age-related degenerative diseases such as cancer, anemia, hypertension, Diabetes mellitus, vitiligo, Parkinson's disease, Alzheimer's disease, schizophrenia and bipolar disorder [26, 27]. Many disease phenotypes can arise from the direct or indirect degradation of the genome, transcriptome, and proteome of tissues and organs by H2O2. But, further research is needed to confirm this argument.

#### **5. Peroxisomal effects on cell proliferation/differentiation, vitality, and apoptosis**

Regulatory molecules involved in cell proliferation/differentiation, vitality, and apoptosis are also vulnerable to damage from oxidative attacks. Damage to molecular regulators has important consequences such as loss of proliferation control, defective cell differentiation/transformation, and apoptosis. This title was designed and shaped considering the antioxidative roles of peroxisomes against oxidative stress.

In a study with a malignant cell line, varying degrees of oxidative stress inhibited the proliferation of hepatoma cells. The number of apoptotic cells increased with the increase of oxidative stress. On the other hand, it is stated that low oxidative stress

<sup>2</sup> It is interpreted as a snake eating its own tail. It is an iconography symbolized for the eternal cyclical renewal or cycle of life, death, and rebirth, as well as the spirit of the world.

<sup>3</sup> Methemoglobin is a type of hemoglobin in which normal hemoglobin has ferric cation (Fe3+) instead of ferrous cation (Fe2+) in the heme group. Methemoglobinemia (MetHb) is a blood disorder in which abnormal amounts of methemoglobin are produced.

#### *The Mystery of Peroxisomes DOI: http://dx.doi.org/10.5772/intechopen.105063*

levels in all of the 4 different indices determined in hepatoma cells (such as accumulation of Con-A on the cell surface, alpha-fetoprotein, gamma-glutamyl transpeptidase, and tyrosine-alpha-ketoglutarate transaminase) result in a tendency for cell differentiation by losing some malignant features. Researchers in that study suggested the possibility that hepatoma cell growth could be inhibited, differentiation and apoptosis could be promoted, and thus reverse transformation could be initiated by tight regulation of oxidative stress [28]. In a study investigating the effect of advanced oxidation protein products (AOPP) in rat osteoblasts, it was determined by gene expression markers that proliferation and differentiation were inhibited in osteoblasts treated with AOPP [29]. It is emphasized that physiological ROS levels, which are accepted as secondary messengers, mediate numerous cellular functions in stem cells, and the need for better quantification of ROS for the correct control of stem cell fate [30]. It has been shown that miR-424, one of the miRNAs identified as key regulators of proliferation and differentiation of mesenchymal stem cells, plays a role in the regulation of bone formation *in vitro*. In this study, it was determined that the down-regulation of miR-424 under oxidative stress induced by H2O2 mediates bone formation [31]. It has long been known that the presence of high levels of ROS leads to impaired cell function and apoptosis. However, within the physiological range, ROS shows a wide range of variable effects. Within these effects, there are consequences ranging from preserving the strength and qualities of pure stem cells to differentiation for a particular cell group. In addition, the effects of ROS may vary in different directions according to stem cell lineage and differentiation stage. For example, the presence of ROS may be associated with decreased embryonic stem cell potency while increasing proliferation in mesenchymal stem cells and an increased likelihood of genomic instability in induced pluripotent stem cells. Again, ROS inhibits osteogenesis while increasing the differentiation of stem cells into cardiomyocytes, adipocytes, endothelial cells, keratinocytes, and neurons, and increases hypertrophic differentiation of cartilage associated with chondrocyte death [32]. Paraquat-induced oxidative stress, a known herbicide, suppressed the expression of stem cell markers including NANOG, OCT4, and TDGF1, while it increased the spontaneous expression of neuronal differentiation markers such as PAX6, NEUROD1, HOXA1, NCAM, GFRA1, and TUJ1. In addition, it has been stated that an increase in the level of intracellular ROS may trigger the exit from the stem cell state and promote neuronal differentiation of human embryonic stem cells (hESC) [33].

#### **6. Peroxisome and immune system modulation**

A healthy immune system in the individual makes effective use of ROS molecules to render many pathogens harmless. Since it is closely related to peroxisome, the known important interactions of ROS at the physiological level in the immune system should be considered.

It is known that peroxisomes, which are single membrane-enclosed organelles in the ROS turnover center, are required to inactivate bacteria by engulfing them with macrophages. Decreased peroxisome function, therefore, disrupts the rapid, temporary, large amounts of ROS production (oxidative burst) turnover needed to fight infection. This impaired response in bacterial control negatively affects the survival of the host cell and organism. Considering phagocytosis and innate immunity, a previously unknown peroxisome requirement has emerged. Dysfunction of peroxisomes in intestinal epithelial cells triggers chain events that affect cell health. This state,

which increases cell death and epithelial instability, activates Tor kinase-dependent autophagy, which alters the gut microbiota, compromises immune pathways in the gut in response to infection, and affects organism survival. In addition, peroxisomes function effectively as centers that coordinate responses from metabolic, immune, and stress signaling pathways to maintain the balance in the functionality of the gut-microbe interface [34, 35]. After host cell invasion, the expression of NADPH oxidase 2 (NOX2) is induced by mycobacteria to form superoxide radicals (O−2). These superoxide anions are first converted to the more toxic hydrogen peroxide (H2O2) via superoxide dismutase (SOD), and catalase reduces H2O2 to water. Contrary to what has long been assumed, accumulating evidence has shown that peroxisomes play a vital role in maintaining cellular redox balance in eukaryotic cells. Deletions in peroxisome-associated Peroxin genes impair detoxification of reactive oxygen species and post-infection peroxisome transformation, leading to the altered synthesis of transcription factors and thus various cell signaling cascades in favor of bacilli [36]. The ability of peroxisomes to modulate fatty acids explains their role as metabolic regulators in various immune functions. It has also been reported that polyamines catabolized by peroxisomes are important for T-cell clonal expansion, alternative macrophage activation, and dendritic cell modulation [37]. Peroxisomes are known to perform important roles in lipid metabolism, but recent studies focus on their essential role in modulating the immune response and inflammation [38]. At least partial peroxisome function is required for the c-Fos and NF-kB-mediated pathways in a study investigating the effects of dysfunctional peroxisomes on the infectionfighting ability of the organism. Without peroxisomes, Drosophila melanogaster has been shown to have a significantly reduced chance of survival after infection. Interestingly, it has been reported that high fatty acid concentration has an inhibitory effect on the cell's immune response [39]. The complex and dynamic interactions that exist between virus and host cells include manipulating peroxisome dynamics in the context of viral infection. Different viruses take advantage of specific peroxisome properties to counteract the host's antiviral response/promote virus particle formation and spread; to this end, viruses modulate peroxisome biogenesis and metabolism. Different studies with the same virus report opposite results, as in SARS-CoV-2 infection. For example, one study reported an increase in the number of peroxisomes, while another study pointed to peroxisome depletion. These differences are likely due to the different stages of infection analyzed. As with human cytomegalovirus (HCMV), probably many viruses interact differently with peroxisomes at different stages of their infection cycle. Viruses can promote peroxisome number depletion early in infection to inhibit host antiviral signaling, but then stimulate peroxisome metabolism and biogenesis to increase lipid metabolism and promote the formation of new virus particles [40].

#### **7. Peroxisome and mitochondria**

There are strong interactions between mitochondria and peroxisomes in terms of the production and removal of reactive molecules. Given that H2O2 can rapidly cross the peroxisomal and mitochondrial membrane, it is reasonable to expect that changes in peroxisomal or mitochondrial H2O2 metabolism will also affect other organelle functions. Whether peroxisomal H2O2 acts directly or indirectly on mitochondria remains to be determined. In this context, inhibition of peroxisomal catalase not only rapidly increases mitochondrial oxidative damage but also decreases the expression of

#### *The Mystery of Peroxisomes DOI: http://dx.doi.org/10.5772/intechopen.105063*

PPAR-γ co-activator (PGC) 1α, a key regulator of mitochondrial biogenesis and function. Moderate levels of mitochondrial ROS are upregulated by multiple antioxidant enzymes (nuclear respiratory factor (NRF) 2 and Forkhead box O (FOXO)) including catalase. Reliable evidence has been found that this stimulation can promote the expression of stress-sensitive transcription factors that mediate stress tolerance. Peroxisome biogenesis is dramatically induced by activation of retrograde signaling pathways in respiratory failure *S. cerevisiae* without inhibition of mitochondrial ATP synthesis. Considering that peroxisomes and mitochondria play a central role in cellular lipid metabolism and lipids have multiple roles related to bioenergetics, cellular signaling, membrane structure and function, changes in lipid metabolism of either peroxisome or mitochondria affect the function of the other organelle. A time- and inhibitory concentration-dependent pattern of pharmacological inhibition of mitochondrial β-oxidation and upregulation of peroxisomal β-oxidation in human and rat liver has been demonstrated [41]. It has also been shown that impaired mitochondrial fatty acid oxidation in skeletal muscle leads to a compensatory increase in peroxisomal fatty acid oxidation [42]. Complex peroxisomal and mitochondrial processes include common substrates (e.g., FAD, NAD+ , O2 ve α-ketoglutarate) and metabolites (e.g., acetyl-CoA, succinate) that have the potential to directly or indirectly modulate the metabolic activities of other subcellular compartments. Many of these common substrates and metabolites can also serve as substrates or inhibitors of DNA methyltransferases, histone (de)methyltransferase/ (de)acetylases. Thus, changes in peroxisomal or mitochondrial activity are likely to affect other organelle activity through epigenetic remodeling. It has long been reported that mitochondria can communicate with the cell through the release of cytochrome c, which has a central role in apoptotic signaling. In addition, it should be well known that other death-promoting factors in the mitochondrial inner membrane space could be released into the cytosol upon induction of apoptosis. Recently, it has been shown that peroxisomes can release matrix proteins into the cytosol. Surprisingly, this release appears to be dependent on voltage-dependent anion-selective channel 2 (VDAC2), a redox-sensitive outer mitochondrial membrane protein. Loss of VDAC2 in Chinese hamster ovary cells shifts the localization of BCL2-antagonist/killer (BAK) 1, a B-cell lymphoma family member central to the mitochondrial pathway of apoptosis, from mitochondria to peroxisomes. This localization shift increases peroxisomal membrane permeability in a manner similar to that in mitochondria, resulting in the release of peroxisomal matrix proteins, including catalase, into the cytosol. It has become increasingly clear that defects in peroxisome biogenesis, peroxisomal fatty acid metabolism, or peroxisomal antioxidant capacity have a negative impact on mitochondrial function. Such mitochondrial defects can be generalized as abnormal cristae, decreased membrane potential and respiratory rates, increased ROS production, decreased fatty acid oxidation, and DNA depletion. In addition, it has been reported to induce ultrastructural and/or functional mitochondrial changes such as mass gain in various organs (e.g., brain, liver, and kidney) and cell types (e.g., skeletal and smooth muscle cells). It shows that changes in peroxisome turnover rates can affect human pathophysiology. Increased peroxisome disruption leads to worsening mitochondrial health. Here, the importance of functional peroxisomes for the maintenance of mitochondrial health is prominently emphasized. Recently, it has become increasingly clear that peroxisomes and mitochondria cooperate to fight viral infections through activation of the RIG-I-like receptors-mitochondrial antiviral signaling (RLR-MAVS) pathway. There is strong evidence to suggest that both peroxisomal and mitochondrial dysfunction may contribute to aging and age-related diseases of the organism. It is recognized

that peroxisomes serve as guardians of mitochondrial health during cellular aging and age-related disease development in patients suffering from congenital peroxisomal disorders [43]. It shows that in the fungus *Podospora anserina*, the activities of peroxisomes and mitochondria in the required process for different stages of sexual development are interrelated. Peroxisomes and mitochondria share proteins that mediate the division of this fungus [44]. The family of phosphatidylinositol 5-phosphate 4-kinases (PI5P4Ks), a class of phosphoinositide kinases, phosphorylates PI-5-P to PI-4,5-P2. This phosphorylated molecule regulates peroxisomal fatty acid oxidation by mediating the transport of lipid droplets to peroxisomes, which is essential for maintaining mitochondrial metabolism [45].

#### **8. Peroxisome and cancer**

Loss of cell division control due to disruptions in the mechanism regulating cell proliferation may cause cancer, which is a devastating disease of this century. Studies focused on the role of the peroxisome in the loss of this control have pointed to peroxisome-proliferator-activated receptor (PPAR) functions. PPARs are nuclear hormone receptors that mediate the effects of fatty acids and their derivatives at the transcriptional level. Through these pathways, PPARs can regulate cell proliferation, differentiation, and survival, thereby controlling carcinogenesis in various tissues [46]. PPARs are also ligand-activated transcription factors involved in the regulation of glucose and lipid homeostasis, inflammation, proliferation, and differentiation. All of these functions draw attention to the influence of PPARs in carcinogenesis [47]. Studies on peroxisomes, which are cellular organelles that affect cancer cell growth and survival, have investigated whether the expression of peroxisome-related genes changes in more than one tumor type. The results of research gave rise to the idea that peroxisomal proteins and their metabolites may support pro-tumorigenic functions. Overexpression of several peroxisomal-associated mRNAs has been reported across a wide range of cancer types (melanoma, breast, non-small cell lung, ovarian, pancreatic, and prostate cancers). In more than 10% of all tumor types included in the analysis, progesterone-induced decidual protein (DEPP) expression was at least twofold higher than the mean expression level. Due to its ability to indirectly induce ROSmediated DNA damage and meet a certain threshold of pro-tumorigenic genomic instability, DEPP may be relatively abundant in multiple tumor types. Moreover, elevated DEPP can activate autophagy, an aberrant process in tumorigenesis, which is particularly upregulated in treatment-resistant cancers. Similarly, PEX16 protein appears to be relatively elevated in tumors compared to normal tissue [48–50]. In different studies investigating the role of peroxisomes in spontaneously formed tumor tissue, it was observed that peroxisomal function is decreased in neoplastic tissue such as colon carcinoma when compared to unaffected tissue. Publications are reporting the amount of peroxisomal protein (catalase, ABCD3, ACOX1, PXMP2) or enzyme (catalase, D-amino acid oxidase, polyamine oxidase, peroxisomal β-oxidation) activities are decreased in breast and hepatocellular carcinomas similarly in colon tumor tissue. A high α-methylacyl-CoA racemase (AMACR) level has been suggested as a reliable prostate cancer tumor marker, as the expression of peroxisomal AMACR was found to be quite high in tissue from prostate carcinoma compared to benign prostate tissue. Not only prostate, but high AMACR expression has also been reported from stomach, colon, breast, kidney, and hepatocellular carcinoma. In the same review, the peroxisomal membrane protein PMP24/PXMP4 has been associated with prostate

#### *The Mystery of Peroxisomes DOI: http://dx.doi.org/10.5772/intechopen.105063*

cancer development. Here, PMP24 is a member of the TIM17 family of membrane proteins. Peroxisomes in glioblastomas have been investigated for tumor-grade progression. Increased immunocytochemistry staining was observed for peroxisome increase associated with progressive tumor grade. This finding was confirmed by the detection of Pex14, PMP70, ACOX1, and 3-ketothiolase proteins, which indicate increased organelle number, namely peroxisome proliferation, using immunoblotting [51]. It was observed that peroxisomal Lon peptidase (LonP2) expression increased in cervical cancer tissue. Basically, LonP2 functions as a combined chaperone/protease by refolding or cleaving degraded peroxisomal proteins. Downregulation of LonP2 in HeLa and SiHA tumor cell lines reduced oxidative stress and inhibited cervical cancer cell proliferation and migration [52, 53]. An unexpected function of peroxisomes in the control of cell division that may be associated with tumor progression is discussed. It has been demonstrated that the correct positioning of peroxisomes during mitosis is required for asymmetric cell division in skin epithelial cells. RNAi-mediated knockdown of Pex11β and Pex14 induced mitotic delay in targeted cells and resulted in an imbalance in growth and differentiation in basal and supra-basal skin cells, together with a reduction in terminal differentiation markers in tissue. The detected mitotic disorder was not associated with an impairment in peroxisomal functions but was found to result from the misplacement of peroxisomes during spindle formation [54].

A study reporting that peroxisomes are indispensable for the survival of liver cancer cells reported that tumor growth was significantly reduced by RNAi silencing of Pex2 in hepatocellular carcinoma xenografts [55]. Consistent with this data, decreased catalase activities were also found in kidney tumors [56]. Peroxisomal protein levels or enzyme activities are greatly reduced in some types of cancer (such as renal, breast, hepatocellular, and colon cancer). Here, hypoxia-inducible transcription factor (HIF-2α) has been shown to promote peroxisome degradation. A decrease in peroxisome abundance was observed in renal carcinoma cells with high HIF-2α levels. These studies report a reduction in peroxisomal activity in some tumor types, while other reports suggest that the metabolic activity of the peroxisome promotes tumor growth. The tumor-promoting or tumor-suppressing function of the peroxisome probably depends on the tumor type in the particular microenvironment. However, peroxisomal genes may be the target of a potential anticancer therapeutic strategy due to their functional role in tumorigenesis. One of the most prominent candidates as a target is alkylglycerone phosphate synthase (AGPS), which is involved in peroxisomal ether lipid biosynthesis. Inactivation of AGPS reduced the levels of ether lipids, including plasmalogens, in breast cancer and melanoma cells, inhibiting their tumorigenicity *in vitro* and *in vivo*. In addition, loss of AGPS decreased the invasive capacity by down-regulating ether lipid expression in glioma and liver cancer cells. Therefore, peroxisomes, which are the center of the antioxidant system, have the potential to be an anticancer target in combination with chemotherapy to generate oxidative stress. Considering many data, it is considered that cancer cells are prone to ROS-induced apoptosis, and factors that impair peroxisome integrity/function may increase the success of cancer therapy. For example, PEX3 degradation predisposes lymphoma cells to ROS-induced apoptosis. Similarly, the loss of proteins such as PEX2 and PEX5 involved in peroxisome protein import stimulates apoptosis in hepatocellular carcinoma (HCC) [57]. In a new study to determine the prognostic value of the peroxisomal pathway in colorectal cancer (CRC), the combined evaluation of T-cell immunoglobulin and mucin domain 3 (TIM3) expression and genes involved in the peroxisome pathway or Fatty acid alpha oxidation (FAAO). Data from here can be used for diagnosis and can be helpful for personalized treatment [58].

In summary, the roles of peroxisomes with pleiotropic effects should be evaluated separately for healthy and cancer cell types. ROS molecules, which are dangerous for healthy cells in the survival process, are also dangerous for cancer cells and their apoptotic effect is dominant. ROS, which is an enemy for healthy cells, is also an important enemy for cancer cells and thus is the enemy of our enemy. This situation, which offers an important perspective for the homeostasis of the organism, reminds us of the proverb "Amicus meus, inimicus inimici mei" (Enemy of my enemy is my friend) that was first mentioned in the work on state administration called "Arthashastra" written in Sanskrit in the 4th century BC.

#### **9. Stimulation or inhibition of peroxisomes' abundance**

Peroxisome abundance is regulated dynamically according to the general conditions of the cell. This regulation is sometimes stimulation and sometimes inhibition. Cell requirements and complex interactions with other cell components determine the direction of this regulation.

Consensus has been reached that the main mode of peroxisome proliferation in yeast is fission. Also, in mammals, fission is most likely the main mode of peroxisome proliferation, although *de novo* synthesis can also occur. *De novo* peroxisome synthesis from the ER was investigated by the reintroduction of the PEX3 gene into PEX3 mutant or peroxisome-deficient yeasts [59].

After treatment of fibroblasts with the peroxisome biogenesis disorder (PBDs) phenotype with 4-phenylbutyrate, an approximately two-fold increase in peroxisome number, again an increase in transcription of the adrenoleukodystrophyrelated gene and PEX11alpha was found. In addition, there was an increase in very-long-chain fatty acid beta-oxidation and plasmalogen concentrations and hence a decrease in very-long-chain fatty acid concentrations in fibroblasts of patients with PBD.

In the process of pexophagy, a type of macroautophagy that selectively degrades peroxisomes, double-membraned autophagosomes surround the peroxisomes, fusing them with lysosomes for degradation. Yeasts (*S. cerevisiae, P. pastoris,* and *Hansenula polymorpha*) can use different carbon sources to obtain energy, so they rapidly increase biogenesis-related peroxisome abundance and the formation of giant peroxisome clusters when grown in environments with oleic acid, methanol, oleate, or amines based on peroxisome metabolism. Conversely, the shift of yeast from peroxisome-dependent carbon sources, such as lipids, to peroxisome-independent carbon sources, such as glucose, triggers their abrupt degradation by pexophagy. Similarly, treating rodents with peroxisome-enhancing stimuli (phthalate esters and hypolipidemic drugs) rapidly increases peroxisome abundance, again triggering large-scale pexophagy upon removal of the stimulus. Pexophagy plays a critical role in peroxisome quality control, as peroxisomes have a half-life of 1.5–2 days in mammals. Failure to import matrix proteins (Pex1p, Pex6p, and Pex15p) identifies peroxisomes for quality control pexophagy. By the accumulation of ubiquitinated Pex5p on the cytosolic surface of the peroxisomal membrane, the pexophagy receptor appears to induce autophagy. However, it has been suggested that other signals, such as phosphorylation of some of the autophagy-related (Atg) proteins in yeast, could be the pexophagy receptor. Overexpression of C-terminal EGFP-tagged PEX5 in transformed mouse embryonic fibroblasts (MEFs) stimulates accumulation and monoubiquitination of PEX5-EGFP in the peroxisome, resulting in pexophagy. During

#### *The Mystery of Peroxisomes DOI: http://dx.doi.org/10.5772/intechopen.105063*

oxidative stress, PEX5 is phosphorylated via mutant ataxia-telangiectasia (ATM) proteins, making it possible to add ubiquitin and subsequently target it by autophagosomes. Yeast pexophagy is regulated via glucose-sensitive and mitogen-activated protein kinase (MAPK) cascades, but the precise mechanisms of these cascades are not fully understood. Although pexophagy is generally viewed as a similar pathway in yeast and mammals, there are distinct differences in the way the pexophagy pathway is governed. Mammalian pexophagy receptors NBR1 and p62 selectively target ubiquitinated peroxisomes for pexophagy, while Atg30p or Atg36p on yeast phosphoactivated pexophagy receptors interact with Pex3p and Atg37p on peroxisomes to mediate pexophagy. Mammals allow differentiation between peroxisomes based on the state of ubiquitin accumulation, while it is not clear whether this type of mechanism exists in yeast [60].

#### **10. Conclusion and projection**

The peroxisome, which is at the center of the lipid metabolism and antioxidant defense system in the cell, is a versatile organelle surrounded by a single membrane. Numerous components work in concert for a flawless and sustainable peroxisome function. Most of these components are Peroxin (PEX) group molecules, and they have signal, transport, and receptor functions in protein import. Some components have structural functions in the peroxisome, and some have enzymatic functions. Syndromes due to peroxisome defects usually represent a heterogeneous group of congenital diseases. Interestingly, mutations in some components of the peroxisome import pathways are not only associated with yeast but also with serious human diseases including peroxisome-related disorders. In some such diseases, only one peroxisomal enzyme is deficient. However, in some other diseases that result from defects in peroxisome function, multiple peroxisomal enzymes cannot be transferred to peroxisomes or are caused by deficiencies in the PTS1 or PTS2 pathways responsible for peroxisome protein transport.

Despite the large body of data available, we are still far from the goal of a full understanding of the peroxisome nature, which is surprisingly plastic and pleiotropic. Each new experimental study illuminates the peroxisome mystery and opens new horizons in terms of interaction between organelles and their effects on cell homeostasis. With the expansion of the peroxisome knowledge pool, we will have a better understanding of the organelle's function and subcellular interaction dynamics, and we will have advanced instruments for new revolutionary treatment strategies.

#### **Conflict of interest**

The author declared no financial or commercial conflict of interest.

*The Metabolic Role of Peroxisome in Health and Disease*

### **Author details**

Hasan Basri İla Faculty of Science and Letter, Department of Biology, Çukurova University, Adana, Turkey

\*Address all correspondence to: ila@cu.edu.tr; hasanbasriila@gmail.com

© 2022 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Section 2
