TRBP, a regulator of cellular PKR and HIV‐1 virus expression, interacts with Dicer and functions in RNA silencing

Abstract

Dicer is a key enzyme involved in RNA interference (RNAi) and microRNA (miRNA) pathways. It is required for biogenesis of miRNAs and small interfering RNAs (siRNAs), and also has a role in the effector steps of RNA silencing. Apart from Argonautes, no proteins are known to associate with Dicer in mammalian cells. In this work, we describe the identification of TRBP (human immunodeficiency virus (HIV-1) transactivating response (TAR) RNA-binding protein) as a protein partner of human Dicer. We show that TRBP is required for optimal RNA silencing mediated by siRNAs and endogenous miRNAs, and that it facilitates cleavage of pre-miRNA in vitro. TRBP had previously been assigned several functions, including inhibition of the interferon-induced double-stranded RNA-regulated protein kinase PKR and modulation of HIV-1 gene expression by association with TAR. The TRBP-Dicer interaction shown raises interesting questions about the potential interplay between RNAi and interferon-PKR pathways.

Principles and effects of microRNA-mediated post-transcriptional gene regulation

“…In addition, orthologues of R2D2 are important for RNAi and miRNA processing in worms flies and mammals. The miRNA specific fly Dicer-1 requires Loquacious, for proper miRNA processing, Dicer in worms coimmunoprecipitates with RDE-4, a dsRNA-binding protein essential for RNAi initiated with long dsRNA, and human Dicer requires two R2D2 orthologues, TRBP and PACT, for miRNA-mediated gene silencing (Tabara et al, 2002;Forstemann et al, 2005;Haase et al, 2005;Jiang et al, 2005;Saito et al, 2005;Lee et al, 2006). Furthermore, Argonautes seem to interact physically with the miRNA maturation and loading machinery as immunoprecipitated hAgo2 complexes contain functional Dicer activity that can asymmetrically process exogenous pre-miRNAs, generating mature RISC that can cleave a perfect complementary target RNA Maniataki and Mourelatos, 2005;Meister et al, 2005).…”

Principles of micro-RNA production and maturation

“…Nevertheless, just like DGCR8 in the case of Drosha, proteins with double-stranded RNA-binding domains, such as TRBP and PACT in humans (Figure 3), bind to Dicer and contribute to Dicer function Forstemann et al, 2005;Gregory et al, 2005;Haase et al, 2005;Jiang et al, 2005;Maniataki and Mourelatos, 2005;Saito et al, 2005;Lee et al, 2006). Thus, they can enhance the affinity of Dicer for RNAs and participate in the selection of mature miRNA strands and/or the transfer of miRNAs to their final stop, the Argonaute proteins.…”

“…Dicer or its complex interacts with a family of conserved proteins called Argonautes (Bernstein et al, 2001;Hammond et al, 2001;Doi et al, 2003;Tahbaz et al, 2004;Chendrimada et al, 2005;Forstemann et al, 2005;Gregory et al, 2005;Haase et al, 2005;Jiang et al, 2005;Maniataki and Mourelatos, 2005;Saito et al, 2005;Lee et al, 2006). Mature miRNAs are eventually transferred to Argonaute proteins and serve as guides in RNA silencing (Caudy et al, 2002;Hutva´gner and Zamore, 2002;Ishizuka et al, 2002;Mourelatos et al, 2002;Liu et al, 2004;Meister et al, 2004).…”

Biochemical mechanisms of the RNA-induced silencing complex

“…Transactivating response RNA-binding protein (TRBP) and protein activator of protein kinase R (PACT) each harbor three annotated dsRNA-binding domains and have been shown to interact with Dicer [35][36][37]. siRNA knockdown of TRBP resulted in diminished miRNA production and loss of reportergene silencing in HeLa cells [35,36].…”