MicroRNA antagonism of the picornaviral life cycle: alternative mechanisms of interference.

In addition to modulating the function and stability of cellular mRNAs, microRNAs can profoundly affect the life cycles of viruses bearing sequence complementary targets, a finding recently exploited to ameliorate toxicities of vaccines and oncolytic viruses. To elucidate the mechanisms underlying microRNA-mediated antiviral activity, we modified the 3' untranslated region (3'UTR) of Coxsackievirus A21 to incorporate targets with varying degrees of homology to endogenous microRNAs. We show that microRNAs can interrupt the picornavirus life-cycle at multiple levels, including catalytic degradation of the viral RNA genome, suppression of cap-independent mRNA translation, and interference with genome encapsidation. In addition, we have examined the extent to which endogenous microRNAs can suppress viral replication in vivo and how viruses can overcome this inhibition by microRNA saturation in mouse cancer models.

Desensitization, internalization, and signaling functions of beta-arrestins demonstrated by RNA interference.

Beta-arrestins bind to activated G protein-coupled receptor kinase-phosphorylated receptors, which leads to their desensitization with respect to G proteins, internalization via clathrin-coated pits, and signaling via a growing list of "scaffolded" pathways. To facilitate the discovery of novel adaptor and signaling roles of beta-arrestins, we have developed and validated a generally applicable interfering RNA approach for selectively suppressing beta-arrestins 1 or 2 expression by up to 95%. Beta-arrestin depletion in HEK293 cells leads to enhanced cAMP generation in response to beta(2)-adrenergic receptor stimulation, markedly reduced beta(2)-adrenergic receptor and angiotensin II receptor internalization and impaired activation of the MAP kinases ERK 1 and 2 by angiotensin II. This approach should allow discovery of novel signaling and regulatory roles for the beta-arrestins in many seven-membrane-spanning receptor systems.