Yan polymerization contributes to active repression and thus contributes to precise gene expression

Abstract

Downstream of the receptor tyrosine kinase (RTK) pathway in Drosophila, a conserved network mediates the robust regulation of genes involved in developmentally important processes such as cell proliferation and differentiation. This regulation is accomplished by a bistable switch, termed the Yan-Pointed (Pnt) switch, in which Yan functions as a transcriptional repressor while Pnt activates the transcription of target genes. Both the mechanism of Yan’s repression as well as any importance of its ability to self-associate, via an N-terminal protein-protein interaction sterile alpha motif (SAM) domain, remain unknown. As Yan exists as an oligomer in vivo and in vitro it is hypothesized that higher order structures, resulting from SAM mediated polymerization, form and favor stable the formation of active complexes contributing to active repression. In the present study and to test this hypothesis, a genetic approach was taken in tandem with an in vivo expression analysis. Polymeric, dimeric, and monomeric constructs of Yan were examined in lethality assays and in target gene expression analyses. Both the dimeric and monomeric constructs revealed decreased rescue ability as well as increased expression of target genes in comparison to the polymer. These studies strongly suggest a higher order structure beyond a dimer is required for robust transcriptional repression by Yan and thereby provide a direct relationship between the SAM domain and the mechanism of the Yan repression.