microRNA strand selection
Mature microRNAs are generated when Dicer enzyme cleaves the precursor microRNA to produce a microRNA duplex, consisting of the “guide” and “passenger,” or miR*, strands. The miR* strand is ejected and rapidly degraded, leaving a single microRNA bound to miRISC free to bind complementary target sites, resulting in translational repression and/or mRNA degradation. Because the sequence of the microRNA dictates the identity of the gene targets to be repressed, selection, or choice, of the correct strand from the precursor duplex is essential for microRNA-mediated gene silencing.The process of microRNA strand selection is highly asymmetric, with one strand chosen as the dominant guide. In addition, strand choice is highly regulated and under certain conditions miR* strands can accumulate and function. In this strand switching phenomenon, any duplex-intrinsic rules of strand selection must be overridden to preferentially incorporate or stabilize the miR* strand via unknown mechanisms.
While great in vitro work from multiple laboratories has shown that 5’ nucleotide identity and thermodynamic properties of the miR::miR* duplex impact strand selection, little is known about what contributes to strand choice in vivo. We have previously found that C. elegans Argonaute ALG-1 plays a role inmicroRNA strand selection. We are continuing to investigate the role of Argonautes in microRNA strand choice. In addition, we are employing a combination of approaches to identify other factors and features that influence microRNA strand selection in vivo using C. elegans as a model. We hope that this work gets us closer to understanding the molecular mechanisms that govern the extreme fidelity of microRNA strand selection in vivo and its regulation in developmental and disease contexts.
Modulation of microRNA function by miRISC co-factors
Very little is known about how microRNA production and activity are regulated by specific developmental and physiological signals. To identify factors that modulate microRNA biogenesis and/or function, we employed a proteomics approach. We have identified a number of Argonaute ALG-1 interactors. We are currently conducting functional studies to understand the roles of these proteins in modulating microRNA activity.
Phenotypic analyses of animals carrying mutations for Argonaute- and microRNA- interacting proteins allow us to determine the functional significance of these factors. We also seek to determine where and when these factors affect microRNA activity. Hence, we are further investigating the physical interactions of these factors with miRISC components.
Zinovyeva AY, Veksler-Lublinsky I, Vashisht AA, Wohlschlegel JA, Ambros VR. 2015 Caenorhabditis elegans ALG-1 antimorphic mutations uncover functions for Argonaute in microRNA guide strand selection and passenger strand disposal. Proc Natl Acad Sci U S A. Sep 8. pii: 201506576. PMID: 26351692
Zinovyeva AY, Bouasker S, Simard MJ, Hammell CM, Ambros V. 2014. Mutations in conserved residues of the C. elegans microRNA Argonaute ALG-1 identify separable functions in ALG-1 miRISC loading and target repression. PLoS Genet. 10(4):e1004286. doi: 10.1371/journal.pgen.1004286. eCollection 2014 Apr.
Zou Y, Chiu H, Zinovyeva A, Ambros V, Chuang CF, Chang C. 2013. Developmental decline in neuronal regeneration by the progressive change of two intrinsic timers. Science. Apr 19;340(6130):372-6. doi: 10.1126/science.1231321.
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