PMC:4502371 / 24054-29102 JSONTXT

{"project":"2_test","denotations":[{"id":"25917920-16452136-43299639","span":{"begin":4515,"end":4519},"obj":"16452136"},{"id":"25917920-15579684-43299640","span":{"begin":4760,"end":4764},"obj":"15579684"},{"id":"25917920-17015622-43299641","span":{"begin":4904,"end":4908},"obj":"17015622"}],"text":"Mutations in dpy-17 suppress mua-3(uy19) lethality\nTo identify genes that interact with mua-3(uy19), we performed two independent unbiased genetic screens using ethyl methyl sulfonate (EMS) mutagenesis to identify suppressors. We used the temperature-dependent lethality of mua-3(uy19) to sort the resulting mutants of the genetic screens.\nAmong 20 isolates, seven suppressors were picked for further study after we removed weak suppressors, sterile mutants, and escapers (animals that produced nonviable progeny at nonpermissive temperatures) (Table 1). We confirmed their rescue by measuring survival at 25° (Figure 6A). Because the screens were performed with F2 progeny from mixed F1s, it is possible that some suppressors share the same mother and contain mutations on the same alleles; yet, several suppressors have distinct phenotypes. For example, S4, one of the strong suppressors (Figure 6A), does not show a Dpy phenotype but shows Eat (pale body color) and Unc (uncoordinated movement). Interestingly, several suppressors of mua-3(uy19) from two independent screens are smaller (Sma) or shorter (Dpy) in body length compared to wild-type. Because one of the major pathways that determines body size in C. elegans is a TGFβ pathway, our results suggest the intriguing possibility that MUA-3 may be involved in TGFβ regulation in C. elegans as in Marfan syndrome pathology in humans.\nTable 1 Characterization of appearance of isolated suppressors of the mua-3(uy19) lethality\nSuppressors Phenotypes\nS1 Small body size, pale body color, uncoordinated movement (Unc)\nS3 Pale body color, small body size (Sma)\nS4 Pale body color, uncoordinated movement (Unc)\nS5 Uncoordinated movement (Unc)\nS6 Small body size, pale body color\nS7 Small body size, pale body color, uncoordinated movement (Unc)\nS9 Short body length (Dpy), similar to dpy-17 mutants\nS10 Small body size\nS11 Short body length (Dpy), similar to dpy-17 mutants\nS16 Slow growth, slow movement, small body size (Sma), pale body color\nS18 Short body length (Dpy), similar to dpy-17 mutants\nS19 Short body length (Dpy), similar to dpy-17 mutants\nS20 Thin body width\nS21 Long body length (Lon)\nS22 Slow growth, slow movement, small body size (Sma), pale body color\nS23 Short body length (Dpy), similar to dpy-17 mutants\nS24 Small body size (Sma)\nS25 No obvious phenotype\nS26 Dark body color\nS27 Dark body color\nS1–S11 are suppressors isolated from the first screen. S16–S27 are suppressors isolated from the second screen. The suppressors tested in Figure 3A are in bold.\nFigure 6 Mutations in dpy-17 rescue mua-3(uy19) lethality. (A) Several suppressors were shown to suppress the mua-3 lethality. The P values were calculated by comparing the percent survival of each suppressor with that of the mua-3 mutants using the Student t test. (B) RNAi of dpy-17 rescued the mua-3 lethality, validating dpy-17 suppression of mua-3. (C) A model to suggest potential interactions among mua-3, dpy-17, and dpy-31 to regulate TGFβ signaling. Fibrillin-1, a human homolog of mua-3, sequesters TGFβ (DBL-1) to reduce the signal, whereas BMP-1/Tolloid metalloprotease (DPY-31) increases the signal. Based on the known interaction between DPY-17 and DPY-31 and its implication based on a function of DPY-31 homolog in mammals, it may be possible that the excess TGFβ (DBL-1) signal in the absence of MUA-3 could be ameliorated by reducing DPY-31 signal when DPY-17 was missing. Four suppressors (S11, S18, S19, S23) looked very similar to each other and shorter in body length than wild-type, exhibiting the Dpy phenotype. They failed to complement dpy-17(e164), indicating that they all carry mutations in dpy-17. We isolated dpy-17 mutations from two independently performed screens (S11 is from the first screen; S18, S19, and S23 are from the second screen), strongly suggesting that dpy-17 mutations suppress mua-3 lethality. S11 is close to sterile, probably due to other mutations; we excluded it from further study. To confirm dpy-17 suppression of mua-3 lethality, we treated mua-3 mutants with dpy-17 RNAi. dpy-17 RNAi rescued mua-3 lethality (Figure 6B), validating dpy-17 as a suppressor. The maximum rescue rate we saw was approximately 50% survival. Only approximately 50% of wild-type animals treated with dpy-17 RNAi showed a clear Dpy phenotype (data not shown), suggesting the low rescue percentage could be due to incomplete RNAi effect.\ndpy-17 encodes a cuticle collagen required for normal body morphology (Novelli et al. 2006). Genetic studies show that a gain-of-function mutation of dpy-17 suppresses phenotypes of dpy-31 mutations, indicating a genetic interaction between the two. dpy-31 encodes a homolog of the human bone morphogenic protein-1 (Novelli et al. 2004). The human bone morphogenic protein-1 is a tolloid-like gene whose proteolytic activity is required for TGFβ activation (Ge and Greenspan 2006). Our screen results suggest a potential link between MUA-3-associated and TGFβ-associated molecules such as DPY-31 via DPY-17 (Figure 6C)."}