Divergent GW182 functional domains in the regulation of translational silencing.

ABSTRACT: MicroRNA (miRNA)-mediated gene regulation has become a major focus in many biological processes. GW182 and its long isoform TNGW1 are marker proteins of GW/P bodies and bind to Argonaute proteins of the RNA induced silencing complex. The goal of this study is to further define and distinguish the repression domain(s) in human GW182/TNGW1. Two non-overlapping regions, ?12 (amino acids 896-1219) containing the Ago hook and ?5 (amino acids 1670-1962) containing the RRM, both induced comparable silencing in a tethering assay. Mapping data showed that the RRM and its flanking sequences in ?5, but not the Ago hook in ?12, were important for silencing. Repression mediated by ?5 or ?12 was not differentially affected when known endogenous repressors RCK/p54, GW182/TNGW1, TNRC6B were depleted. Transfected ?5, but not ?12, enhanced Ago2-mediated repression in a tethering assay. Transfected ?12, but not ?5, released endogenous miRNA reporter silencing without affecting siRNA function. Alanine substitution showed that GW/WG motifs in ?12 (?12a, amino acids 896-1045) were important for silencing activity. Although ?12 appeared to bind PABPC1 more efficiently than ?5, neither ?5 nor ?12 significantly enhanced reporter mRNA degradation. These different functional characteristics of ?5 and ?12 suggest that their roles are distinct, and possibly dynamic, in human GW182-mediated silencing.

SUBMITTER: Yao B 

PROVIDER: S-EPMC3074120 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature