Fig. 4. UNC-43 CaMKII activation in the proximal gonad. (A) The rat sequence used to generate CaMKII-pT is highly conserved in C. elegans UNC-43, including the phosphorylated threonine (^*, T284 in UNC-43 and T286 in rat CaMKII). (B-D) CaMKII-pT stains the oocytes of wild-type gonads. In mid-focal plane images, staining co-localizes with F-actin at the oocyte cortex. (E-G) CaMKII-pT staining is not observed in unc-43(n1186)-null gonads. (H-J) At the sheath and oocyte interface of wild-type gonads, CaMKII-pT staining is observed in the same focal plane as sheath cell F-actin. (K) In contrast to CaMKII-pT, MAPK-YT staining is uniformly distributed throughout the cytoplasm, and sometimes in the nucleus. (L) CaMKII-pT stains sheath cells surrounding the sperm in fem-3(q20) gonads. The genetic requirements of UNC-43 T284 phosphorylation are shown in M-U. (M) No CaMKII-pT staining is observed in unmated fog-3(q443) females, indicating that sperm are required for UNC-43 T284 phosphorylation. (N,O) CaMKII-pT stains unmated fog-2(q71) female gonads microinjected with 100 nM MSP (N), but not buffer-injected control gonads (O). (P) CaMKII-pT staining is not observed in unmated vab-1 RNAi ceh-18(mg57) females, which undergo constitutive oocyte maturation independent of sperm presence. CaMKII-pT staining is not observed in vab-1(dx31) (Q) and nmr-1(ak4) gonads (S), but it is observed in vab-1(e2) kinase dead (R), itr-1(sa73) (T) and itr-1(sy290gf) (U) gonads. All images in B-M were processed with deconvolution software and include DAPI to visualize DNA. All gonads are oriented as shown in Fig. 1A. Scale bars: 10 µm.