Figure 7. EPG-7 is not required for starvation-induced autophagic degradation of protein substrates. (A and B) C17E4.2::GFP is weakly expressed and diffusely localized in a wild-type embryo. (A) DIC image of the embryo shown in B. (C and D) C17E4.2::GFP is expressed at dramatically elevated levels and accumulates into many aggregates in
lgg-1 (C) and
epg-7 (D) mutant embryos. (E and F) W07G4.5::GFP is expressed in the intestine and a few aggregates are formed in a wild-type embryo at the fourfold stage. (E) DIC image of the embryo shown in F. (G and H) W07G4.5::GFP is expressed at dramatically elevated levels and accumulates into a large number of aggregates in
atg-3 (G) and
epg-7 (H) mutants. Images in B-D or F-H were taken using the same exposure time. (I) Levels of C33D9.6::GFP and W07G4.5::GFP are dramatically elevated in
atg-3 and
epg-7 mutant embryos. (J and K) C17E4.2 aggregates in
atg-3 mutants overlap with EPG-7 aggregates (J), but are separable from SQST-1 aggregates (K). Insets show a magnified view. (L) Colocalization frequency of C17E4.2, C33D9.6, and W07G4.5 aggregates with EPG-7 and SQST-1 aggregates in
atg-3 mutants. (M) SQST-1::GFP accumulates in hypodermal cells in
epg-7 mutants at the larval stage. (N) SQST-1::GFP aggregates disappear in
rheb-1(RNAi);
epg-7 mutant larvae. (O) Dramatic accumulation of SQST-1::GFP aggregates in hypodermal cells in
atg-3 mutant larvae. (P) SQST-1::GFP aggregates persist in
rheb-1(RNAi);
atg-3 mutant larvae. (Q-T) At larval stages, W07G4.5::GFP forms aggregates in intestinal cells (Q and R). These aggregates disappear upon RNAi knockdown of
let-363 (S and T). (U-X) W07G4.5::GFP aggregates accumulate in intestinal cells in
epg-7 mutant larvae (U and V). These aggregates disappear when
let-363 is simultaneously depleted (W and X). (Y and Z) In
atg-3 mutants, W07G4.5::GFP aggregates accumulate in intestinal cells (Y) and persist after RNAi knockdown of
let-363 (Z). (A2) Levels of W07G4.5::GFP in wild type,
epg-7 mutants, and
atg-3 L1 larvae before and after 6 h starvation treatment. (B2) Model for the role of
epg-7 in degradation of SQST-1 aggregates. SQST-1 directly interacts with EPG-7 and is recruited into EPG-7 aggregates. EPG-7 also associates with multiple Atg proteins, which may be recruited to the SQST-1-EPG-7 complex in a hierarchical order. IM, isolation membrane. (C2) Hierarchical relationship of the scaffold proteins EPG-2 and EPG-7 and other essential ATG and EPG proteins in the aggrephagy pathway. Bars: (A-H, J, and K) 10 um; (J and K, insets) 5 um; (M-Z) 10 um.