**1.2. Autophagy pathway**

1963. Prior to this term, however, autophagy was first observed, or at least hinted in as early as 1955 [1]. Kleinfeld and his colleagues found that the process involves three continuous stages of maturation and is seen as being used for the reutilization of cellular materials and organelle disposal. In Duve's definition, autophagy is a part of lysosomal function and glucagon being the main inducer of hepatic cell degradation. Together with his student, Russell Deter, they were the first to demonstrate that lysosomes play a central role for intracellular autophagy [2, 3]. Using Duve's work as reference, independent scientist groups discovered autophagy-related genes in yeast. In that period, Ohsumi and Michael Thumm studied on nonselective autophagy induced by starvation [4, 5]. At the same time, Klionsky discovered a form of selective autophagy called the cytoplasm-to-vacuole targeting (CVT) pathway [6, 7]. Not longer after, they discovered that their independent work actually revolves around the same pathway. Through collaborating, they published a paper titled "Cytoplasm-to-vacuole targeting and autophagy employ the same machinery to deliver proteins to the yeast vacuole" [8]. In 2003, a unified nomenclature was advocated by scientists in the field to use ATG for autophagy-related genes [9]. More than a decade later, in 2016, Ohsumi was finally awarded a Nobel Prize in Physiology or Medicine for his contribution toward the field of autophagy. While it is undeniable that Ohsumi deserves the Nobel Prize, some individuals have pointed out that the prize should have been more inclusive of other researchers who made Ohsumi's work possible [10]. In the second millennium, there was an accelerated growth of research in autophagy thanks to the work and contribution by these scientists on ATG genes. With the fundamentals on autophagy set strong, scientists began to study its association with human health and diseases. The first breakthrough discovery associating autophagy with cancer

76 Cell Culture

**Figure 1.** Cellular autophagy processes. (1) Phagophore forms and elongates to package the cargoes comprising damaged organelles and proteins. (2) LC3B proteins are then recruited and together with phagophore, it forms autophagosome. (3) LC3B proteins are then dissociated, and the cytoplasmic cargo is sequestered to fuse with endosome-derived lysosome to form autolysosome. (4) Hydrolytic enzymes then degrade the cargoes and release metabolic products and

building blocks such as amino acids and fatty acids for nutrient recycling.

Autophagy is an evolutionary conserved mechanism involved in maintaining homeostasis and metabolism at the cellular level by degrading proteins with long half-life and clearing cytoplasmic organelles through lysosomes. Lipids, nucleotides, and glycogen are also subjected to lysosomal degradation via autophagy. Like other pathways involved in homeostasis, the cellular pathways are highly regulated. There are evidences that the recycling of proteins and other macromolecules may contribute to protective roles in normal development, senescence, cell death, and as a defense mechanism against intracellular infections. **Figure 1** shows the detailed processes involved in the autophagy pathway. The dysregulation of autophagy has been shown to cause cancer, inflammatory, metabolic, and neurodegenerative diseases [12, 13].
