Phospho-ATG13 Ser355 ELISA Kit

Sample

cell lysates

Species Reactivity

Human

Intended Use

The Phospho-ATG13 Ser355 ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Atg13 protein when phosphorylated at Ser355.

Sensitivity

The Phospho-ATG13 (Ser355) ELISA Kit detects endogenous levels of Atg13 protein phosphorylated at Ser355. The kit sensitivity is shown in Figure 1. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.


Figure 1. Atg13 is phosphorylated at Ser355 in 293T cells that were co-transfected with constructs expressing human Atg13 and mouse ULK1, but is not phosphorylated in cells transfected with only an Atg13 expression construct. The relationship between lysate protein concentration from Atg13/ULK1 co-transfected and Atg13-only transfected cells and the absorbance at 450 nm using the Phospho-ATG13 Ser355 ELISA Kit is shown in the upper figure. The corresponding western blots using phospho-Atg13 (Ser355) antibody (left panel) and Atg13 antibody (right panel) are shown in the lower figure. 293T cells were either co-transfected with constructs expressing Myc/DDK-tagged full-length human Atg13 (hAtg13-Myc/DDK) and mouse ULK1 (mULK1) or just transfected with a construct expressing Myc/DDK-tagged full-length human Atg13 (hAtg13-Myc/DDK), and then lysed.

General Description

Autophagy is a catabolic process for the autophagosomic-lysosomal degradation of bulk cytoplasmic contents (1,2). Autophagy is generally activated by conditions of nutrient deprivation but has also been associated with a number of physiological processes including development, differentiation, neurodegeneration, infection, and cancer (3). The molecular machinery of autophagy was largely discovered in yeast and referred to as autophagy-related (Atg) genes.
Atg13/Apg13 was originally identified in yeast as a constitutively expressed protein that was genetically linked to Atg1/Apg1, a protein kinase required for autophagy (4). Overexpression of Atg1 suppresses the defects in autophagy observed in Atg13 mutants (4). Autophagy requires a direct association between Atg1 and Atg13, and is inhibited by TOR-dependent phosphorylation of Atg13 under high-nutrient conditions (5). Similarly, mammalian Atg13 forms a complex with the Atg1 homologues ULK1/2, along with FIP200, which localizes to autophagic isolation membranes and regulates autophagosome biogenesis (6-8). mTOR phosphorylates both Atg13 and ULK1, suppressing ULK1 kinase activity and autophagy (7-9). ULK1 can directly phosphorylate Atg13 at a yet unidentified site, presumably to promote autophagy (7,8). Additional studies suggest that Atg13 and FIP200 can function independently of ULK1 and ULK2 to induce autophagy through an unknown mechanism (10).
ULK1-dependent phosphorylation of Atg13 at Ser355, which corresponds to Ser318 of isoform 2 of Atg13, leads to the recruitment of Atg13 to damaged mitochondria, enabling efficient mitophagy (11).