ATG18D Antibody

Target Overview: ATG18/WIPI Proteins

ATG18 is a conserved autophagy-related protein with homologs across species:

• Human homologs: WIPI1 (ATG18A) and WIPI2 (ATG18B) [1].

• Function: Regulates autophagosome formation by binding phosphatidylinositol 3-phosphate (PtdIns(3)P) and interacting with ATG2 to mediate membrane expansion [3,4].

• Isoforms:

  • WIPI1 (48.7 kDa, 446 residues) has two isoforms [1].

  • WIPI2 has six isoforms (42–48 kDa) [7].

3.1. Molecular Interactions

• ATG18 binds PtdIns(3)P via a FRRG motif, facilitating recruitment to autophagic membranes [3].

• The ATG18-ATG2 complex is essential for autophagosome formation; mutations in the FRRG motif disrupt this interaction and impair autophagy [4].

3.2. Functional Assays

• Liposome Binding: Purified ATG18 binds liposomes in a PtdIns(3)P-dependent manner, confirmed by surface plasmon resonance [4].

• Co-immunoprecipitation: ATG18 interacts with ATG2 even when PtdIns(3)P binding is disrupted, indicating a secondary binding mechanism [3].

4.1. Western Blot (WB)

• Primary Antibody Dilution: 1:300–1:5,000 [6].

• Sample Preparation: Use RIPA buffer for lysates; detect bands at 42–48 kDa [7].

4.2. Immunoprecipitation

• Cross-linking: DSP (dithiobis-succinimidyl-propionate) stabilizes ATG18-ATG2 complexes for co-IP [5].

Limitations and Considerations

• Isoform Specificity: Many antibodies cross-react with WIPI1/WIPI2 due to sequence homology.

• Phosphorylation Effects: ATG18 activity is modulated by phosphorylation, which may affect antibody binding [4].