Protein disulfide bond formation or reduction are important modification steps for controlling protein function and assembly. Proteins expressed in the extracellular matrices are usually good target substrate for such modifications. One of the enzymes that facilitates this process is thioredoxin, which may be an important player in matrix organization and thus morphogenesis of C. elegans .
dpy-11 is one of many ram genes that affect sensory ray morphogenesis. Mutations of it give swollen sensory rays at the male tail.
dpy-11 has been cloned and is defined as the predicted thioredoxin-like-protein encoding gene, F46E10.9. Using a gfp reporter, we determine its expression pattern to be in the hypodermal cells in all larval and adult stages. To characterize the cellular defect of
dpy-11 mutants, different cell-type specific GFP markers were employed to show that all ray cells within a ray process have abnormal morphology. It implies that the
dpy-11 mutants have defects in protein modification and possibly cell differentiation facilitated by cell-cell communication. The protein structure of DPY-11 was predicted to have a putative signal peptide, thioredoxin-like domain, a spacer, transmembrane domain and a 46 amino acid carboxyl terminus. To understand the function of
dpy-11 , phenotypic characterization and mutation mapping of 16 existing alleles have been carried out. Mutations on the signal peptide and transmembrane domain affect the function of DPY-11 on body shape, but not the rays. The null mutants have most severe dumpiness and severe swollen rays while the animals with point mutations on thioredoxin-like domain have mild dumpiness and mild swollen rays. Heterozygous mutants of severe and mild alleles have been examined, and the results show that DPY-11 acts in a dosage dependent manner. For further characterization of each protein domain, deletion constructs are made for mutant rescue assay. The enzymatic activity of thioredoxin-like domain is expressed in bacteria and purified for reduction assay. Different versions of DPY-11 GFP fusion protein will also be employed to reveal the membrane-bound property and subcellular localization of this protein. These studies will provide us a clear picture when, where and how
dpy-11 takes part in the morphogenesis of male sensory rays.