Regulation of the Drosophila melanogaster protein, enhancer of rudimentary, by casein kinase II

The Drosophila melanogaster gene enhancer of rudimentary, e(r), encodes a conserved protein, ER. Most ER homologs share two casein kinase II (CKII) target sites. In D. melanogaster, these sites are T18 and S24. A third CKII site, T63, has been seen only in drosophilids. The conservation of these CKII sites, particularly T18 and S24, suggests a role for these residues in the function of the protein. To test this hypothesis, these positions were mutated either to alanine as a nonphosphorylated mimic or to glutamic acid as a phosphorylated mimic. The mutations were tested individually or in double or triple combinations for their ability to rescue either a wing truncation characteristic of the genotype e(r)(p1) r(hd1-12) or the synthetic lethal interaction between e(r)(p2) and the Notch allele N(nd-p). All of the substitutions as single mutations rescued both mutant phenotypes, arguing that individually the phosphorylation of the three residues does not affect ER activity. The double mutants T18A-S24A and T18E-S24E and the triple mutants T18A-S24A-T63A and T18E-S24E-T63E failed to rescue. Together the data support the following model for the regulation of ER by CKII. ER that is unphosphorylated at both T18A and S24 is inactive. CKII activates ER by phosphorylating either T18 or S24. Further phosphorylation to produce the doubly phosphorylated protein inactivates ER.     

Drosophila Sex-lethal protein mediates polyadenylation switching in the female germline

The Drosophila master sex-switch protein Sex-lethal (SXL) regulates the splicing and/or translation of three known targets to mediate somatic sexual differentiation. Genetic studies suggest that additional target(s) of SXL exist, particularly in the female germline. Surprisingly, our detailed molecular characterization of a new potential target of SXL, enhancer of rudimentary (e(r)), reveals that SXL regulates e(r) by a novel mechanism--polyadenylation switching--specifically in the female germline. SXL binds to multiple SXL-binding sites, which include the GU-rich poly(A) enhancer, and competes for the binding of CstF64 in vitro. The SXL-binding sites are able to confer sex-specific poly(A) switching onto an otherwise nonresponsive polyadenylation signal in vivo. The sex-specific poly(A) switching of e(r) provides a means for translational regulation in germ cells. We present a model for the SXL-dependent poly(A) site choice in the female germline.