CONICET | Buscador de Institutos y Recursos Humanos

Smaug is an mRNA repressor that defines a novel family of RNA-binding proteins. Mammalian Smaug1 (mSmaug1) is restricted to mature neurons and mSmaug1 knockdown provokes the formation of immature synapses and impairs the response to synaptic stimulation (Baez et al., JCB, 2011; Pascual et al., CIB 2012). Both Drosophila Smaug and mSmaug 1 repress the translation of reporter mRNAs carrying specific motifs termed Smaug-Recognition-Element (SRE) and form mRNA silencing foci termed S-foci when expressed in cell lines (Baez and Boccaccio, JBC 2005). In mammalian neurons, the S-foci are located at the post-synapse. The S-foci are static under basal conditions and highly dynamic upon neuron stimulation, and they dissolve and release key mRNAs thus allowing their translation. Here we show that mSmaug 1 aggregation is independent of its binding to RNA and requires two conserved regions, a D domain located at the N-terminus and the C-terminal region, which includes a conserved PHAT (pseudo-HEAT repeat analogous topology) domain. Co-tranfection experiments suggest that both the D domain and the C-terminal regions interact homotypically thus facilitating mSmaug1 oligomerization. Aggregation of the Drosophila molecule requires the conserved D domain and is additionally mediated by protein regions absent from the mammalian molecule. In addition, we found that the aggregation domains are required for mRNA repression, suggesting that foci formation modulates Smaug-mediated silencing. Finally, we found that several cell components involved in miRNA-mediated silencing and deadenylation are recruited to the S-foci. Ago2, GW182/TNRC6, PABPC and the PAN2/PAN3 deadenylase complex co-localize or associate with S-foci when co-transfected with mSmaug1. Altogether, these observations suggest a role for the miRNA pathway in mSmaug1-mediated repression, which will be additionally regulated by conserved aggregation domains that respond to synaptic stimulation.