Figure 4. SEPA-1 Forms Cytoplasmic Aggregates and Is Degraded by Autophagy. (A and B) No obvious expression of
sepa-1 could be detected by anti-SEPA-1 antibody in embryos before the 16-cell stage. An 8-cell-stage embryo is shown. (A) DAPI image of the embryo shown in (B).(C and D) The number of SEPA-1 aggregates increases during early stages of embryogenesis (from 20-cell to 100-cell stages). (C) DAPI image of the embryo shown in (D).(E and F) SEPA-1 aggregates are almost undetectable in a comma-stage embryo. (E) Nomarski image of the embryo shown in (F).(G-J) SEPA-1 is not present in germline P granules (also see Figure S7). P granules are detected by anti-PGL-3 antibody. (G)-(J) are confocal images.(K and L) Expression of
sepa-1::gfp is first detected in 16-cell-stage embryos. SEPA-1 distribution is mainly diffuse, with a few aggregates. (K) Nomarski image of the embryo shown in (L).(M-P) The number of SEPA-1::GFP aggregates increases during early stages of embryogenesis (before 100-cell stage) and decreases as development proceeds. SEPA-1::GFP is still diffusely distributed in a comma-stage embryo (O and P) but is almost undetectable in a 4-fold-stage embryo (data not shown). (M and O) Nomarski images of the embryos shown in (N) and (P), respectively.(Q and R) SEPA-1::GFP is localized in the intestine at larval stages, especially in the anterior and posterior pairs of intestine cells. (R, upper panel) Nomarski image.(S-W) Expression of
sepa-1, detected by anti-SEPA-1 antibody (S and T) or
sepa-1::gfp (U-W), is dramatically increased in
lgg-1 animals. (S and T) Comma stage. (U and V) 4-fold stage. (W) L1 larva. (S and U) Nomarski images of the embryos shown in (T) and (V), respectively.(X) The mRNA level of
sepa-1, detected by RT-PCR, is the same in wild-type and
lgg-1 mutants. The mRNA level of actin serves as a control.