Two Dimerization Domains in the Trans-activation Response RNA-binding Protein (TRBP) Individually Reverse the Protein Kinase R Inhibition of HIV-1 Long Terminal Repeat Expression

Abstract

Trans-activation response (TAR) RNA-binding protein (TRBP) is a cellular protein that binds to the human immunodeficiency virus-1 (HIV-1) TAR element RNA. It has two double-stranded RNA binding domains (dsRBDs), but only one is functional for TAR binding. TRBP interacts with the interferon-induced protein kinase R (PKR) and inhibits its activity. We used the yeast two-hybrid assay to map the interaction sites between the two proteins. We show that TRBP and PKR-N (178 first amino acids of PKR) interact with PKR wild type and inhibit the PKR-induced yeast growth defect in this assay. We characterized two independent PKR-binding sites in TRBP. These sites are located in each dsRBD in TRBP, indicating that PKR-TRBP interaction does not require the RNA binding activity present only in dsRBD2. TRBP and its fragments that interact with PKR reverse the PKR-induced suppression of HIV-1 long terminal repeat expression. In addition, TRBP activates the HIV-1 long terminal repeat expression to a larger extent than the addition of each domain. These data suggest that TRBP activates gene expression in PKR-dependent and PKR-independent manners.

Merlin, a Tumor Suppressor, Interacts with Transactivation-responsive RNA-binding Protein and Inhibits Its Oncogenic Activity

“…The two isoforms of TRBP, TRBP1 (original TRBP) and TRBP2, are expressed by alternative transcriptional initiation, resulting in the TRBP2 protein that is longer than TRBP1 by 21 amino acids at its amino terminus (14 -16). TRBP was originally cloned as an HIV-1 transactivation-response RNA-binding protein and belongs to the family of double-stranded RNA (dsRNA)-binding proteins with two clearly defined dsRNAbinding domains (dsRBDs) and a carboxyl-terminal basic region (17)(18)(19)(20)(21). TRBP dsRBD2 binds transactivation-response with higher affinity than dsRBD1, because of the presence of a KR helix motif (16,17).…”

Emergence of a Complex Relationship between HIV-1 and the microRNA Pathway

“…TRBP is also known to inhibit the interferon (IFN)-induced dsRNA-regulated protein kinase R (PKR) (94), and to be involved in miRNAguided RNA silencing, more specifically, in assisting Dicer function within a pre-miRNA processing complex (24,25). Immunoprecipitation approaches identified TRBP as a Dicerinteracting protein (24,25).…”

RNA interference and HIV‐1 infection

“…Another interesting function of the TRBP protein is the translation-independent PKR enhancement that depends on a structured RNA [99] and on inhibition of PKR activity [100] suggesting a strong link between RNAi and IFN-PKR pathways. On the other hand, PKR could also phosphorylate Tat [101] which in turn can inhibit PKR by RNA-dependent and independent mechanisms [102].…”