*2.1.3 Transcription factors with nutrigenomic potential in dairy cattle*

The TFs are fundamental to the study of nutrigenomics; they can act as intermediaries between dietary nutrients and the final alteration in gene expression. TFs that respond to nutrient effects can be activated directly or indirectly by nutrients, and upon activation, they translocate from the cytoplasm to the nucleus where they alter the transcription of specific target genes. Transcription factors can bind specific short DNA sequences (i.e., 6–12 nucleotides) called response elements located in the enhancer regions upstream of the actual gene sequence [57]. The ability of TF to modulate gene expression upon activation by nutrients confers these proteins a central stage in the field of nutrigenomics. Therefore, accurate identification and characterization of TF responding to specific nutrients and to what extent these TF can be manipulated through dietary effects should be the focus of future research in nutrigenomics in dairy cattle.

Although between 2000 and 3000 TFs with sequence-specific DNA-binding domains in the human genome were estimated, only ~100 have been experimentally verified for their DNA-binding and regulatory functions [58]. The AnimalTFDB website had collected information for almost 1300 TF and almost 400 transcription cofactors for *Bos taurus* [59]. Rather than TF working independently, a regulatory network of TF has been suggested, which is essential to coordinate the response to external stimuli and translate this into changes in gene expression [60].

Among the known TF, the nuclear receptor superfamily of TF, with 48 members in the human genome, is the most important group of nutrient sensors [61]. From this superfamily, a short list of TFs has been identified as ligand-dependent nuclear receptors (LdNR), which can bind and be activated by macro- and micronutrients [55]. Recently, the main LdNR with a potential role in nutrigenomics with an emphasis in large [62] and small [24] ruminants has been reviewed. Among the LdNR associated with macronutrients such as fatty acids are the peroxisome proliferator-activated receptors (PPAR), liver X receptors (LXR), and hepatic

nuclear factor 4 (HNF4) [63]. The LdNR associated with micronutrients are primarily vitamin-specific and include retinoid X receptors (RXR) and retinoic acid receptors (RAR) activated by retinoic acids or metabolites of vitamin A [64], vitamin D receptor (VDR), and pregnane X receptor activated by vitamin E [65].

The PPAR belongs to the LdNR group of TF and requires to form heterodimers with RXR in order to be functional. The main characteristic of PPAR is to have a prominent role in controlling expression of genes involved in lipid metabolism and inflammation. The potential nutrigenomic role of PPAR in ruminants has been reviewed at length previously [10]. This review discussed the role of PPARα in controlling lipid metabolism and inflammation in liver, the potential role of PPARβ/δ in controlling glucose uptake in mammary tissue, and the potential role of PPARγ in controlling milk fat synthesis and mastitis [10]. An interesting feature of PPAR is their capacity for binding and be activated by LCFA in both monogastrics and ruminants [10, 66]. However, in the case of ruminants, data indicate that activation of the PPAR isotypes PPARα and PPARγ is more consistent with saturated LCFA, primarily palmitate and stearate, than unsaturated LCFA [10].

#### *2.1.4 Nutriepigenomics in dairy cattle*

Environment factors such as diet and ambient conditions can not only affect the short- and medium-term gene expression, but there is also a medium- to long-term regulation of genes. The latter is primarily carried out through changes in the availability of gene sequences to be transcribed into mRNA. This concept is referred to epigenetics, where "epi" is a Greek-derived term meaning "over" then, epigenetics is commonly referred as "on-top-of genetics." The implications of epigenetics indicate that there could be a set of inherited characteristics, phenotypes, and chemical entities that are superimposed on the DNA and do not follow basic Mendelian laws. Every individual will have a set of epigenetic marks throughout the genome, which is also known as the epigenome.

Epigenetic modifications are carried out through several biological processes including DNA methylation, histone modifications (e.g., methylation and acetylation), noncoding RNA (e.g., micro- and long-RNA). And, when these biological processes respond to nutrients and compounds in the diet, it is associated with nutriepigenomic effects. In ruminants, such effects could serve important physiological adaptations during the onset of lactation including increasing the availability of gene sequences through alterations in DNA methylation for the transcription of essential proteins such as caseins in the mammary gland of dairy cows [67]. This new spinoff of nutrigenomics (i.e., nutriepigenomics) will provide essential information to our understanding of how nutrients can affect the biology of ruminants at a molecular level. However, at the same time, nutriepigenomics will add another layer of complexity to our field, where such interactions have to be fully understood, and in time, manipulated through dietary interventions.

Methylation is a major route for epigenetic modifications, through DNA and histone methylation. Therefore, methyl donors (e.g., choline, methionine, folic acid, etc.) found in the diets of dairy cattle can have a nutriepigenomic effect. These dietary methyl donors will likely increase the synthesis of SAM, which is the major biological methyl donor in the body [68]. The essential role of SAM within the context of the transition cow relies on the multiple biological processes that require this methyl donor, including transsulfuration, polyamine biosynthesis, DNA methylation [69], and histone methylation [70]. Among these, the epigenetic modifications caused by DNA and histone methylation are particularly important in order to understand the potential nutriepigenomic alterations caused by dietary methyl donor (e.g., methionine) supplementation.

**129**

*Gene Regulation in Ruminants: A Nutritional Perspective*

DNA methylation occurs through specialized enzymes called DNA methyltransferases, which utilized the methyl group provided by SAM to methylate cytosines within a Cyt-phosphate-Gua (CpG) region ("island") in the DNA and eventually creating methylated CpG patterns in the mammalian genome [71]. In ruminants, supplementation of rumen-protected to dairy cows resulted in a prepartal upregulation of *DNMT3A*, a gene that encodes for a DNA methyltransferase in charge of the de novo methylation of the DNA [72]. And, more recently, the significance of these findings was confirmed by observing alterations due to methionine supplementation in the liver of transition dairy cows in terms of global DNA methylation and specific region methylation of an important TF, the peroxisome proliferatoractivated receptor alpha PPARα [8]. The characteristics and uniqueness of this TF within the context of the transition dairy cow were initially presented by Drackley [73], and since then, this nuclear receptor has become an exciting area of research in dairy cattle nutrigenomics (i.e., nutrient-gene interaction) [10]. Therefore, the connection between Met and PPARα upregulation through DNA methylation during the transition period is another suitable mechanism to explain the consistent improvements in performance (e.g., milk yield and DMI) observed in transition

In the cellular nuclei, the DNA is normally packed in condensed structures called chromatins, consisting primarily of histone proteins, which serve as spools where the DNA winds around. Then, the genetic information contained in the DNA exists in two states: unavailable or wind around histone proteins, and available or unwound. Chromatin remodeling is the main mechanism by which DNA is wind or unwound from histones and these dynamic modifications occur by enzymatic modifications including acetylation, phosphorylation, ubiquitination, and methylation [75]. The latter being a potential mechanism through which Met can alter gene expression in dairy cows. Currently, the limited amount of data on histone methylation in dairy cows has been conducted in immune cells [76] primarily related to subclinical mastitis [77]. This work has provided nuances on the interactions between mastitis-related pathogens and histone methylation; however, dietary

In order to understand how changes in a single gene could significantly alter the body structure and physiology of beef cattle, we need to focus in mutations that occur in specific areas in the bovine genome. For example, a 11-nucleotide deletion in the Myostatin gene (MSTN) determines double muscling in Belgian Blue cattle, and a single nucleotide change produces the same effect on Piedmontese cattle [78]. Furthermore, the leptin gene (LEP) presents several single nucleotide polymorphisms (SNP), like AJ236854:c.73 T > C, which induces a cysteine-to-arginine amino acid substitution that could affect protein functionality [79]. The c.73C allele of LEP is associated with higher average daily gain (ADG), lower dressing percentage, and higher marbling scores [79], which are desirable characteristics in a beef carcass. Finally, intramuscular fat deposition could also be affected by a mutation in the fatty acid synthase gene (FASN), contributing to the characteristic fatty acid composition of Japanese Black beef [80]. However, not all mutations have beneficial effects on the productivity of meat-producing animals. There are also mutations that are considered lethal, affecting, for example, the reproductive performance of females through early embryonic loss [81], or mutations that produce genetic

*DOI: http://dx.doi.org/10.5772/intechopen.82193*

dairy cows supplemented with Met [74].

**3. Gene regulation in beef cattle**

**3.1 Mutations**

effects on histone methylation have not been investigated.

*Gene Regulation in Ruminants: A Nutritional Perspective DOI: http://dx.doi.org/10.5772/intechopen.82193*

*Gene Expression and Control*

nuclear factor 4 (HNF4) [63]. The LdNR associated with micronutrients are primarily vitamin-specific and include retinoid X receptors (RXR) and retinoic acid receptors (RAR) activated by retinoic acids or metabolites of vitamin A [64], vitamin D receptor (VDR), and pregnane X receptor activated by vitamin E [65]. The PPAR belongs to the LdNR group of TF and requires to form heterodimers with RXR in order to be functional. The main characteristic of PPAR is to have a prominent role in controlling expression of genes involved in lipid metabolism and inflammation. The potential nutrigenomic role of PPAR in ruminants has been reviewed at length previously [10]. This review discussed the role of PPARα in controlling lipid metabolism and inflammation in liver, the potential role of PPARβ/δ in controlling glucose uptake in mammary tissue, and the potential role of PPARγ in controlling milk fat synthesis and mastitis [10]. An interesting feature of PPAR is their capacity for binding and be activated by LCFA in both monogastrics and ruminants [10, 66]. However, in the case of ruminants, data indicate that activation of the PPAR isotypes PPARα and PPARγ is more consistent with saturated LCFA,

primarily palmitate and stearate, than unsaturated LCFA [10].

stood, and in time, manipulated through dietary interventions.

methyl donor (e.g., methionine) supplementation.

Environment factors such as diet and ambient conditions can not only affect the short- and medium-term gene expression, but there is also a medium- to long-term regulation of genes. The latter is primarily carried out through changes in the availability of gene sequences to be transcribed into mRNA. This concept is referred to epigenetics, where "epi" is a Greek-derived term meaning "over" then, epigenetics is commonly referred as "on-top-of genetics." The implications of epigenetics indicate that there could be a set of inherited characteristics, phenotypes, and chemical entities that are superimposed on the DNA and do not follow basic Mendelian laws. Every individual will have a set of epigenetic marks throughout the genome, which

Epigenetic modifications are carried out through several biological processes including DNA methylation, histone modifications (e.g., methylation and acetylation), noncoding RNA (e.g., micro- and long-RNA). And, when these biological processes respond to nutrients and compounds in the diet, it is associated with nutriepigenomic effects. In ruminants, such effects could serve important physiological adaptations during the onset of lactation including increasing the availability of gene sequences through alterations in DNA methylation for the transcription of essential proteins such as caseins in the mammary gland of dairy cows [67]. This new spinoff of nutrigenomics (i.e., nutriepigenomics) will provide essential information to our understanding of how nutrients can affect the biology of ruminants at a molecular level. However, at the same time, nutriepigenomics will add another layer of complexity to our field, where such interactions have to be fully under-

Methylation is a major route for epigenetic modifications, through DNA and histone methylation. Therefore, methyl donors (e.g., choline, methionine, folic acid, etc.) found in the diets of dairy cattle can have a nutriepigenomic effect. These dietary methyl donors will likely increase the synthesis of SAM, which is the major biological methyl donor in the body [68]. The essential role of SAM within the context of the transition cow relies on the multiple biological processes that require this methyl donor, including transsulfuration, polyamine biosynthesis, DNA methylation [69], and histone methylation [70]. Among these, the epigenetic modifications caused by DNA and histone methylation are particularly important in order to understand the potential nutriepigenomic alterations caused by dietary

*2.1.4 Nutriepigenomics in dairy cattle*

is also known as the epigenome.

**128**

DNA methylation occurs through specialized enzymes called DNA methyltransferases, which utilized the methyl group provided by SAM to methylate cytosines within a Cyt-phosphate-Gua (CpG) region ("island") in the DNA and eventually creating methylated CpG patterns in the mammalian genome [71]. In ruminants, supplementation of rumen-protected to dairy cows resulted in a prepartal upregulation of *DNMT3A*, a gene that encodes for a DNA methyltransferase in charge of the de novo methylation of the DNA [72]. And, more recently, the significance of these findings was confirmed by observing alterations due to methionine supplementation in the liver of transition dairy cows in terms of global DNA methylation and specific region methylation of an important TF, the peroxisome proliferatoractivated receptor alpha PPARα [8]. The characteristics and uniqueness of this TF within the context of the transition dairy cow were initially presented by Drackley [73], and since then, this nuclear receptor has become an exciting area of research in dairy cattle nutrigenomics (i.e., nutrient-gene interaction) [10]. Therefore, the connection between Met and PPARα upregulation through DNA methylation during the transition period is another suitable mechanism to explain the consistent improvements in performance (e.g., milk yield and DMI) observed in transition dairy cows supplemented with Met [74].

In the cellular nuclei, the DNA is normally packed in condensed structures called chromatins, consisting primarily of histone proteins, which serve as spools where the DNA winds around. Then, the genetic information contained in the DNA exists in two states: unavailable or wind around histone proteins, and available or unwound. Chromatin remodeling is the main mechanism by which DNA is wind or unwound from histones and these dynamic modifications occur by enzymatic modifications including acetylation, phosphorylation, ubiquitination, and methylation [75]. The latter being a potential mechanism through which Met can alter gene expression in dairy cows. Currently, the limited amount of data on histone methylation in dairy cows has been conducted in immune cells [76] primarily related to subclinical mastitis [77]. This work has provided nuances on the interactions between mastitis-related pathogens and histone methylation; however, dietary effects on histone methylation have not been investigated.

### **3. Gene regulation in beef cattle**

#### **3.1 Mutations**

In order to understand how changes in a single gene could significantly alter the body structure and physiology of beef cattle, we need to focus in mutations that occur in specific areas in the bovine genome. For example, a 11-nucleotide deletion in the Myostatin gene (MSTN) determines double muscling in Belgian Blue cattle, and a single nucleotide change produces the same effect on Piedmontese cattle [78]. Furthermore, the leptin gene (LEP) presents several single nucleotide polymorphisms (SNP), like AJ236854:c.73 T > C, which induces a cysteine-to-arginine amino acid substitution that could affect protein functionality [79]. The c.73C allele of LEP is associated with higher average daily gain (ADG), lower dressing percentage, and higher marbling scores [79], which are desirable characteristics in a beef carcass. Finally, intramuscular fat deposition could also be affected by a mutation in the fatty acid synthase gene (FASN), contributing to the characteristic fatty acid composition of Japanese Black beef [80]. However, not all mutations have beneficial effects on the productivity of meat-producing animals. There are also mutations that are considered lethal, affecting, for example, the reproductive performance of females through early embryonic loss [81], or mutations that produce genetic

disorders in beef cattle [82]. The Arachnomelia syndrome in Simmental cattle [83] that produce malformations of the skeleton mainly affecting the legs, the spinal column, and the skull is an example of these genetic disorders. These types of inborn errors can be prevented nowadays with techniques like genome editing [83]. Although the implementation of this technique in the animal production industry might generate controversy, it will offer tremendous potential for breeding animals with desirable traits.

Undesired mutations present in the bovine genome are difficult to avoid when they occur. Through the implementation of selection plans and genotyping the herds in order to avoid the reproduction of carrying individuals is the most commonly utilized strategy. Similarly, selection of animals that carry biomarkers in their genomes that will make them improve their meat production and marbling efficiency is the general goal of researchers passionate about beef production. Currently, identification of relevant genetic and genomic markers is ongoing, especially for tenderness—a top priority quality attribute [84].

#### **3.2 Nutrigenomics in beef cattle**

In spite of the emergence of new alternatives to beef production (e.g., cultivated meat) in order to meet the growing world population's food demand, researchers in beef production are also focusing on techniques to regulate the bovine genome with a more natural approach (i.e., nutrigenomics). Nutrigenomics study the interactions between nutrients and genes [85], that is how the nutrients present in the diet can affect gene expression. In beef animals, nutrigenomics was widely studied [26]; following, a more detailed description of how specific nutrients regulate the expression of genes related to muscle growth and intramuscular fat deposition will be addressed.

The composition of adipose tissue produced in a meat-producing animal can be "manipulated" by diet, with some variability between breeds. For example, high silage-based diets produce less proportion of the fatty acid 18:2 n-6, with the consequent decrease in the amount of total polyunsaturated fatty acids (PUFA), as compared to low silage-based diets in subcutaneous adipose tissue [86]. This variability in fatty acids composition was attributed to greater activity of fatty acid binding protein 4 (FABP4), lipoprotein lipase (LPL), and stearoyl CoA desaturase (SCD) genes in subcutaneous adipose tissue of animals fed a low silage-based diet [86].

In the same way, we are able to regulate the fatty acid profile composition of a specific fat depot by dietary changes, researchers are trying to prioritize the growth and development of the intramuscular fat that will lead to greater marbling. Intramuscular fat starts to accumulate in the late stages of growth, as compared to other adipose tissue depots that normally develop first (i.e., visceral, intermuscular, and subcutaneous fat). The ultimate goal is to improve marbling scores that will lead to premium prices for the carcasses that classify as prime or choice according to the USDA carcass grading scale [87].

When comparing dairy and beef cattle breeds, subcutaneous fatty acid profile presents several differences in terms of fatty acids profile, probably due to the difference in the degree of fatness, which is always lower for dairy cattle [88]. The important variability in fat composition between breeds could be explained by the variability in relative SCD1 expression in subcutaneous fat [88]. SCD1 seems to have a role in a depot-specific fashion.

#### **3.3 How management decisions can affect gene expression**

The combination of early weaning and high dietary starch leads to a strong programming effect in skeletal muscle tissue, with PPARγ and CCAAT

**131**

*Gene Regulation in Ruminants: A Nutritional Perspective*

intramuscular fat deposition in the LM following castration.

enhancer-binding protein alpha (CEBPA) as the central coordinators of this response. The implementation of early weaning (e.g., 2 months of age) in beef calves provides a different type of diet as compared to a calf weaned normally at 6–7 months of age. Therefore, the early administration of starch in a beef producing animal produces a precocious and sustained activation of the PPARG and its target genes, leading to greater intramuscular fat deposition and consequently more

Castration increases intramuscular fat (IMF) deposition, improving beef quality in cattle. In a transcriptome analysis performed in longissimus muscle (LM) samples, when comparing bulls and steers, castration showed to upregulate transcription of genes for lipogenesis, fatty acid oxidation, and also genes coding for enzymes associated with the tricarboxylic acid cycle and oxidative phosphorylation [90]. Therefore, castration shifts the transcriptome of lipid/energy metabolism to favor

Another beef producer decision that can affect the expression of genes is the selected calving season (i.e., different temperature/humidity index = THI). For example, two groups of pregnant cows that calved during thermoneutral temperature conditions (THI = 67.3) and cows that calved in summer season (THI = 79.9) were bleed during their transition period (i.e., cows between 3 weeks before and 3 weeks after calving) for RNA extraction of white blood cells [91]. Results showed that expression of CASP-3, BCL-2, BAK, P53, and ratio of BAX/BCL-2 in white blood cells increased during summer as compared to thermoneutral conditions, suggesting the susceptibility of these cells to apoptosis or cell death [91]. Consequently, cows calving in two different calving seasons will present differences in their inflammatory response, affecting the maternal recognition of the fetus during early pregnancy [92] or also will have a negative impact in the cow's milking

The beef industry has two main ways to produce beef: pasture-base, and grain-

based or feedlot. Consuming energy above requirements helps to increase the intramuscular fat deposition in beef cattle. Feeding high concentration of cereal grains is the way to reach surplus energy that can be utilized in the rumen and the small intestine to produce volatile fatty acids for glucose and energy production. The starch present in the grain is fermented by microbes in the rumen, producing propionate (a glucose precursor), which will be the signal received by the membrane receptors present in the cell activating a cascade of events that will end up with the activation of genes related to the process of adipogenesis. This type of diet is more commonly administered during the so-called finishing or fattening phase. When the diet is mainly based on the forage available, the rumen population consists of microbes that produce a greater proportion of acetic acid which increases the activation of 5′-prime-AMP-activated protein kinase alpha (AMPK) phosphorylation, reducing the transcriptional activity of the sterol regulatory element-binding protein 1c (SREBP1C) and the carbohydrate responsive element-binding protein (MLXIPL), which decreased the expression of lipogenic genes [94]. In beef cattle finished under a forage-based diet, the fatty acid profile varies considerably as compared to animals finished under feedlot diets. Beef finished under forage-based diets presents greater concentration of polyunsaturated fatty acids (PUFAs), especially the fatty acids with nutraceutical value (20:5 or EPA and 22:6 or DHA). These types of beef products are in the eye of consumers who care about eating healthy foods. These PUFAs mentioned above are upstream regulators of genes related to fatty acid synthesis and transport. FABP4, FASN, and PPARG are particularly activated by these additional PUFAs

*DOI: http://dx.doi.org/10.5772/intechopen.82193*

carcasses grading as "Choice" [89].

ability postcalving [93].

**3.4 Feedlot versus pasture**

#### *Gene Regulation in Ruminants: A Nutritional Perspective DOI: http://dx.doi.org/10.5772/intechopen.82193*

*Gene Expression and Control*

with desirable traits.

**3.2 Nutrigenomics in beef cattle**

the USDA carcass grading scale [87].

seems to have a role in a depot-specific fashion.

**3.3 How management decisions can affect gene expression**

The combination of early weaning and high dietary starch leads to a strong programming effect in skeletal muscle tissue, with PPARγ and CCAAT

disorders in beef cattle [82]. The Arachnomelia syndrome in Simmental cattle [83] that produce malformations of the skeleton mainly affecting the legs, the spinal column, and the skull is an example of these genetic disorders. These types of inborn errors can be prevented nowadays with techniques like genome editing [83]. Although the implementation of this technique in the animal production industry might generate controversy, it will offer tremendous potential for breeding animals

Undesired mutations present in the bovine genome are difficult to avoid when they occur. Through the implementation of selection plans and genotyping the herds in order to avoid the reproduction of carrying individuals is the most commonly utilized strategy. Similarly, selection of animals that carry biomarkers in their genomes that will make them improve their meat production and marbling efficiency is the general goal of researchers passionate about beef production. Currently, identification of relevant genetic and genomic markers is ongoing,

In spite of the emergence of new alternatives to beef production (e.g., cultivated meat) in order to meet the growing world population's food demand, researchers in beef production are also focusing on techniques to regulate the bovine genome with a more natural approach (i.e., nutrigenomics). Nutrigenomics study the interactions between nutrients and genes [85], that is how the nutrients present in the diet can affect gene expression. In beef animals, nutrigenomics was widely studied [26]; following, a more detailed description of how specific nutrients regulate the expression of genes related to muscle growth and intramuscular fat deposition will be addressed. The composition of adipose tissue produced in a meat-producing animal can be "manipulated" by diet, with some variability between breeds. For example, high silage-based diets produce less proportion of the fatty acid 18:2 n-6, with the consequent decrease in the amount of total polyunsaturated fatty acids (PUFA), as compared to low silage-based diets in subcutaneous adipose tissue [86]. This variability in fatty acids composition was attributed to greater activity of fatty acid binding protein 4 (FABP4), lipoprotein lipase (LPL), and stearoyl CoA desaturase (SCD) genes in subcutaneous adipose tissue of animals fed a low silage-based diet [86]. In the same way, we are able to regulate the fatty acid profile composition of a specific fat depot by dietary changes, researchers are trying to prioritize the growth and development of the intramuscular fat that will lead to greater marbling. Intramuscular fat starts to accumulate in the late stages of growth, as compared to other adipose tissue depots that normally develop first (i.e., visceral, intermuscular, and subcutaneous fat). The ultimate goal is to improve marbling scores that will lead to premium prices for the carcasses that classify as prime or choice according to

When comparing dairy and beef cattle breeds, subcutaneous fatty acid profile presents several differences in terms of fatty acids profile, probably due to the difference in the degree of fatness, which is always lower for dairy cattle [88]. The important variability in fat composition between breeds could be explained by the variability in relative SCD1 expression in subcutaneous fat [88]. SCD1

especially for tenderness—a top priority quality attribute [84].

**130**

enhancer-binding protein alpha (CEBPA) as the central coordinators of this response. The implementation of early weaning (e.g., 2 months of age) in beef calves provides a different type of diet as compared to a calf weaned normally at 6–7 months of age. Therefore, the early administration of starch in a beef producing animal produces a precocious and sustained activation of the PPARG and its target genes, leading to greater intramuscular fat deposition and consequently more carcasses grading as "Choice" [89].

Castration increases intramuscular fat (IMF) deposition, improving beef quality in cattle. In a transcriptome analysis performed in longissimus muscle (LM) samples, when comparing bulls and steers, castration showed to upregulate transcription of genes for lipogenesis, fatty acid oxidation, and also genes coding for enzymes associated with the tricarboxylic acid cycle and oxidative phosphorylation [90]. Therefore, castration shifts the transcriptome of lipid/energy metabolism to favor intramuscular fat deposition in the LM following castration.

Another beef producer decision that can affect the expression of genes is the selected calving season (i.e., different temperature/humidity index = THI). For example, two groups of pregnant cows that calved during thermoneutral temperature conditions (THI = 67.3) and cows that calved in summer season (THI = 79.9) were bleed during their transition period (i.e., cows between 3 weeks before and 3 weeks after calving) for RNA extraction of white blood cells [91]. Results showed that expression of CASP-3, BCL-2, BAK, P53, and ratio of BAX/BCL-2 in white blood cells increased during summer as compared to thermoneutral conditions, suggesting the susceptibility of these cells to apoptosis or cell death [91]. Consequently, cows calving in two different calving seasons will present differences in their inflammatory response, affecting the maternal recognition of the fetus during early pregnancy [92] or also will have a negative impact in the cow's milking ability postcalving [93].

#### **3.4 Feedlot versus pasture**

The beef industry has two main ways to produce beef: pasture-base, and grainbased or feedlot. Consuming energy above requirements helps to increase the intramuscular fat deposition in beef cattle. Feeding high concentration of cereal grains is the way to reach surplus energy that can be utilized in the rumen and the small intestine to produce volatile fatty acids for glucose and energy production. The starch present in the grain is fermented by microbes in the rumen, producing propionate (a glucose precursor), which will be the signal received by the membrane receptors present in the cell activating a cascade of events that will end up with the activation of genes related to the process of adipogenesis. This type of diet is more commonly administered during the so-called finishing or fattening phase.

When the diet is mainly based on the forage available, the rumen population consists of microbes that produce a greater proportion of acetic acid which increases the activation of 5′-prime-AMP-activated protein kinase alpha (AMPK) phosphorylation, reducing the transcriptional activity of the sterol regulatory element-binding protein 1c (SREBP1C) and the carbohydrate responsive element-binding protein (MLXIPL), which decreased the expression of lipogenic genes [94]. In beef cattle finished under a forage-based diet, the fatty acid profile varies considerably as compared to animals finished under feedlot diets. Beef finished under forage-based diets presents greater concentration of polyunsaturated fatty acids (PUFAs), especially the fatty acids with nutraceutical value (20:5 or EPA and 22:6 or DHA). These types of beef products are in the eye of consumers who care about eating healthy foods. These PUFAs mentioned above are upstream regulators of genes related to fatty acid synthesis and transport. FABP4, FASN, and PPARG are particularly activated by these additional PUFAs

generated due to the administration of forage-based diets [95]. Even though we could expect a greater proportion of IMF due to the mentioned additional PUFAs, there is an overall lower fat content in a grass-fed beef product. Furthermore, grass-fed beef is known to have a different flavor and aroma as compared to grain-fed beef when cook on the grill [96].

#### **3.5 Fetal programming in beef cattle**

The fetal programming concept is related to the important physiological changes that can occur due to environmental/nutritional events during prenatal development. For example, nutrient restriction of 85% as compared to 140% of total metabolizable energy requirements during the second half of gestation can alter in fetal muscle the expression of both, myogenic and adipogenic genes, without apparent differences in fetal phenotype [97].

The canonical Wnt pathway, a β-catenin-dependent signaling pathway called the Wnt/β-catenin signaling pathway is key in establishing the fate of the undifferentiated stem cells; hence, β-catenin plays an essential role in the regulation of embryonic, postnatal, and oncogenic growth of many tissues. If the β-catenin pathway is blocked, the total number of myocytes will be reduced, and the differentiation of mesenchymal stem cells into mature adipocytes will be potentiated [98]. In the same way, adipogenesis is initiated around mid-gestation in ruminant animals; therefore, a strategic maternal nutritional plan in order to enhance the number of mesenchymal cells committed to adipogenesis will increase the number of intramuscular adipocytes in a process known as hyperplasia; therefore, this outcome will be translated as more marbling in the offspring postnatally. PPARG alone can stimulate adipocyte differentiation [99], although the continuation of this process is regulated by many PPARG target genes [100].

Bioinformatics analysis revealed a pseudoinflammatory process in early-weaned beef calves during their growing phase [101], which it is associated with the activation of the innate immune system presumably due to macrophage infiltration of intramuscular fat [101], which is a typical obesity symptom. These results could be considered as a biological sign of a precocious beginning of the adipogenic metabolic machinery in young beef calves.

#### **3.6 Epigenetics regulations in beef cattle**

Changes caused by chromatin (the complex of DNA and histone proteins that forms chromosomes within the nucleus) modification due to environmental factors is called epigenetics [102]. Another epigenetics regulation approach is through microRNAs, which are endogenous noncoding small RNA molecules (20–24 base pairs) that prevent the production of a particular protein by binding to and destroying the messenger RNA that would have produced the protein [103].

Epigenetics regulation is based on chromatin remodeling rather than alteration of the DNA code. With the aim to identify methylated genes affecting bovine growth, an elegant study provides a genome-wide landscape of DNA methylomes and their relationship with mRNA and miRNA for fetal and adult muscle of Chinese Qinchuan beef cattle [104]. Presence of SNPs in epigenetic-related genes was analyzed in different beef breeds. Interestingly, three DNA (cytosine-5)-methyltransferases (DNMTs), DNMT1, DNMT3a, and DNMT3b were found significantly associated with beef quality parameters on the carcass. In particular, DNMT3b presented five SNPs related to carcass traits, becoming a potential candidate gene for beef quality improvement [105].

In the bovine genome, no microRNAs were identified on chromosome Y, while microRNA related to adipose tissue are expressed in chromosome X [106]. This

**133**

*Gene Regulation in Ruminants: A Nutritional Perspective*

expression in grass-fed as compared to grain-fed cattle.

could be a reason that explains the sexual differences in fat metabolism in mammals. Furthermore, there are highly expressed microRNAs for beef adipose tissue, miR-378 which was found upregulated in steers with high levels of subcutaneous fat [107] and miR-2478 [108] which potentially targets ELOVL fatty acid elongase 6 (ELOVL6) and stearoyl CoA desaturase (SCD), is bovine specific and had higher

A study in bovine skeletal muscle development used next-generation small RNA sequencing, a total of 512 miRNAs were identified [109]. Thirty-six miRNAs were differentially expressed between prenatal and postnatal stages of muscle development including several myomiRs (miR-1, miR-206, and let-7 families). Compared to miRNA expression between different muscle tissues, 14 miRNAs were upregulated and 22 miRNAs were downregulated in the muscle of postnatal stage [109]. Furthermore, a genome-wide landscape of DNA methylomes and their relationship with mRNA and miRNA for bovine fetal and adult muscle recently discovered will provide a solid basis for exploring the epigenetic mechanisms of muscle growth and

Muscling in cattle influenced by genetic background, ultimately affecting beef yield is of major interest to the beef industry. The best alternative to promote muscle development is through satellite cell proliferation [110]. In fact, myoblast or satellite cells are utilized for the proliferation and differentiation of cultured meat [111]. The transcription factor Sp1, an activator of myosin D (MyoD) and a suppressor of cyclin-dependent kinase inhibitor 1A (CDKN1A), plays an important role in bovine muscle cell proliferation and differentiation. This transcription factor is a target of miR-128 and, if this microRNA is overexpressed, it inhibits muscle satellite cell proliferation and differentiation [112]. Furthermore, miR-1 and miR-206 facilitate bovine skeletal muscle satellite cell myogenic differentiation by restricting the expression of their target genes, and that inhibition of miR-1 and miR-206 increased the paired box 7 (Pax7) and histone deacetylase 4 (HDAC4) protein levels

In Biceps femoris muscle of Japanese Shorthorn cattle, a grazing period up to 4 months increased the expression of miR-208b, which has a muscle fiber typeassociated role. Furthermore, a target for miR-208b, MyoD, a myogenic regulatory factor associated with the shifting of muscle property to the fast type, had lower expression in the grazed cattle after 4 months of grazing, as compared to feedlot cattle. During skeletal muscle adaptation to grazing, miR-206 expression remained higher as compared to housed animals in which it decreases [114]. MiR-206 is

Offspring's health depends on the maternal body condition at mid-gestation, which will make them be more predisposed to develop obesity at an early age, which in beef production is desirable. Fetal intramuscular adipogenesis was enhanced at mid-gestation due to alteration of microRNA expression; downregulation of let-7g was the main cause for this outcome [116]. This microRNA inhibits the mRNA expression of PPARG and CEBPA, both important regulators of adipogenesis [117]. Furthermore, cow plane of nutrition during the last third of gestation showed epigenetic effects on the offspring's skeletal muscle through the downregulation of miR-34a that has a role on the activation of cell cycle arrest by suppressing SIRT1

The importance of ruminants to the world food security is reflected in their contribution to the demand for animal protein around the world and particularly

*DOI: http://dx.doi.org/10.5772/intechopen.82193*

enhancing satellite cell proliferation [113].

known as the skeletal muscle-specific myomiRNA [115].

expression, which promotes adipocyte differentiation [118].

**4. Molecular nutrition in ruminants**

development [104].

*Gene Expression and Control*

cook on the grill [96].

**3.5 Fetal programming in beef cattle**

ent differences in fetal phenotype [97].

regulated by many PPARG target genes [100].

bolic machinery in young beef calves.

**3.6 Epigenetics regulations in beef cattle**

generated due to the administration of forage-based diets [95]. Even though we could expect a greater proportion of IMF due to the mentioned additional PUFAs, there is an overall lower fat content in a grass-fed beef product. Furthermore, grass-fed beef is known to have a different flavor and aroma as compared to grain-fed beef when

The fetal programming concept is related to the important physiological changes that can occur due to environmental/nutritional events during prenatal development. For example, nutrient restriction of 85% as compared to 140% of total metabolizable energy requirements during the second half of gestation can alter in fetal muscle the expression of both, myogenic and adipogenic genes, without appar-

The canonical Wnt pathway, a β-catenin-dependent signaling pathway called the Wnt/β-catenin signaling pathway is key in establishing the fate of the undifferentiated stem cells; hence, β-catenin plays an essential role in the regulation of embryonic, postnatal, and oncogenic growth of many tissues. If the β-catenin pathway is blocked, the total number of myocytes will be reduced, and the differentiation of mesenchymal stem cells into mature adipocytes will be potentiated [98]. In the same way, adipogenesis is initiated around mid-gestation in ruminant animals; therefore, a strategic maternal nutritional plan in order to enhance the number of mesenchymal cells committed to adipogenesis will increase the number of intramuscular adipocytes in a process known as hyperplasia; therefore, this outcome will be translated as more marbling in the offspring postnatally. PPARG alone can stimulate adipocyte differentiation [99], although the continuation of this process is

Bioinformatics analysis revealed a pseudoinflammatory process in early-weaned beef calves during their growing phase [101], which it is associated with the activation of the innate immune system presumably due to macrophage infiltration of intramuscular fat [101], which is a typical obesity symptom. These results could be considered as a biological sign of a precocious beginning of the adipogenic meta-

Changes caused by chromatin (the complex of DNA and histone proteins that forms chromosomes within the nucleus) modification due to environmental factors is called epigenetics [102]. Another epigenetics regulation approach is through microRNAs, which are endogenous noncoding small RNA molecules (20–24 base pairs) that prevent the production of a particular protein by binding to and destroy-

Epigenetics regulation is based on chromatin remodeling rather than alteration of the DNA code. With the aim to identify methylated genes affecting bovine growth, an elegant study provides a genome-wide landscape of DNA methylomes and their relationship with mRNA and miRNA for fetal and adult muscle of Chinese Qinchuan beef cattle [104]. Presence of SNPs in epigenetic-related genes was analyzed in different beef breeds. Interestingly, three DNA (cytosine-5)-methyltransferases (DNMTs), DNMT1, DNMT3a, and DNMT3b were found significantly associated with beef quality parameters on the carcass. In particular, DNMT3b presented five SNPs related to carcass

In the bovine genome, no microRNAs were identified on chromosome Y, while microRNA related to adipose tissue are expressed in chromosome X [106]. This

ing the messenger RNA that would have produced the protein [103].

traits, becoming a potential candidate gene for beef quality improvement [105].

**132**

could be a reason that explains the sexual differences in fat metabolism in mammals. Furthermore, there are highly expressed microRNAs for beef adipose tissue, miR-378 which was found upregulated in steers with high levels of subcutaneous fat [107] and miR-2478 [108] which potentially targets ELOVL fatty acid elongase 6 (ELOVL6) and stearoyl CoA desaturase (SCD), is bovine specific and had higher expression in grass-fed as compared to grain-fed cattle.

A study in bovine skeletal muscle development used next-generation small RNA sequencing, a total of 512 miRNAs were identified [109]. Thirty-six miRNAs were differentially expressed between prenatal and postnatal stages of muscle development including several myomiRs (miR-1, miR-206, and let-7 families). Compared to miRNA expression between different muscle tissues, 14 miRNAs were upregulated and 22 miRNAs were downregulated in the muscle of postnatal stage [109]. Furthermore, a genome-wide landscape of DNA methylomes and their relationship with mRNA and miRNA for bovine fetal and adult muscle recently discovered will provide a solid basis for exploring the epigenetic mechanisms of muscle growth and development [104].

Muscling in cattle influenced by genetic background, ultimately affecting beef yield is of major interest to the beef industry. The best alternative to promote muscle development is through satellite cell proliferation [110]. In fact, myoblast or satellite cells are utilized for the proliferation and differentiation of cultured meat [111]. The transcription factor Sp1, an activator of myosin D (MyoD) and a suppressor of cyclin-dependent kinase inhibitor 1A (CDKN1A), plays an important role in bovine muscle cell proliferation and differentiation. This transcription factor is a target of miR-128 and, if this microRNA is overexpressed, it inhibits muscle satellite cell proliferation and differentiation [112]. Furthermore, miR-1 and miR-206 facilitate bovine skeletal muscle satellite cell myogenic differentiation by restricting the expression of their target genes, and that inhibition of miR-1 and miR-206 increased the paired box 7 (Pax7) and histone deacetylase 4 (HDAC4) protein levels enhancing satellite cell proliferation [113].

In Biceps femoris muscle of Japanese Shorthorn cattle, a grazing period up to 4 months increased the expression of miR-208b, which has a muscle fiber typeassociated role. Furthermore, a target for miR-208b, MyoD, a myogenic regulatory factor associated with the shifting of muscle property to the fast type, had lower expression in the grazed cattle after 4 months of grazing, as compared to feedlot cattle. During skeletal muscle adaptation to grazing, miR-206 expression remained higher as compared to housed animals in which it decreases [114]. MiR-206 is known as the skeletal muscle-specific myomiRNA [115].

Offspring's health depends on the maternal body condition at mid-gestation, which will make them be more predisposed to develop obesity at an early age, which in beef production is desirable. Fetal intramuscular adipogenesis was enhanced at mid-gestation due to alteration of microRNA expression; downregulation of let-7g was the main cause for this outcome [116]. This microRNA inhibits the mRNA expression of PPARG and CEBPA, both important regulators of adipogenesis [117]. Furthermore, cow plane of nutrition during the last third of gestation showed epigenetic effects on the offspring's skeletal muscle through the downregulation of miR-34a that has a role on the activation of cell cycle arrest by suppressing SIRT1 expression, which promotes adipocyte differentiation [118].
