Human Immunodeficiency Virus Type 1 (HIV-1) Induces the Cytoplasmic Retention of Heterogeneous Nuclear Ribonucleoprotein A1 by Disrupting Nuclear Import

Abstract

Human immunodeficiency virus type 1 (HIV-1) co-opts host proteins and cellular machineries to its advantage at every step of the replication cycle. Here we show that HIV-1 enhances heterogeneous nuclear ribonucleoprotein (hnRNP) A1 expression and promotes the relocalization of hnRNP A1 to the cytoplasm. The latter was dependent on the nuclear export of the unspliced viral genomic RNA (vRNA) and to alterations in the abundance and localization of the FG-repeat nuclear pore glycoprotein p62. hnRNP A1 and vRNA remain colocalized in the cytoplasm supporting a post-nuclear function during the late stages of HIV-1 replication. Consistently, we show that hnRNP A1 acts as an internal ribosomal entry site trans-acting factor up-regulating internal ribosome entry site-mediated translation initiation of the HIV-1 vRNA. The up-regulation and cytoplasmic retention of hnRNP A1 by HIV-1 would ensure abundant expression of viral structural proteins in cells infected with HIV-1.During the late stages of the HIV-1 3 replication cycle, the full-length HIV-1 viral RNA (vRNA) must be exported from the nucleus and both translated and packaged into new viral particles (1). The orchestration of these events is directed by a diversity of viral and host proteins that interact with each other and with the vRNA to form HIV-1 ribonucleoprotein (RNP) complexes that originate in the nucleus and persist in the cytoplasm (2). Investigations into the composition and functions of the HIV-1 RNP will reveal new information about innate immunity as well as identify new potential therapeutic targets (3-6).The heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) is a predominantly nuclear protein engaged in a number of cellular and viral RNPs for RNA-processing activities, including splicing regulation, nuclear export, microRNA processing, mRNA stability, telomere maintenance, and IRES-mediated translation initiation (7-12). What enables hnRNP A1 to have such broad functions in RNA metabolism is its ability to bind both nuclear and cytoplasmic RNAs (10). In addition to its well characterized role in nuclear RNA processing, hnRNP A1 binds to purine-rich sequences of mRNAs for mRNA turnover and translation (13,14). Close examination of the HIV-1 vRNA reveals many AG-and AU-rich sequences closely resembling hnRNP A1 binding motifs (15). In fact, hnRNP A1 binds to a number of sequences on the HIV-1 vRNA such as the cis-acting repressive sequences, instability elements, and exonic splicing silencer elements, and indeed, hnRNP A1 is implicated in the fate of HIV-1 RNA, including splicing regulation, nucleocytoplasmic export, and cytoplasmic stability (16 -21).Our previous work demonstrated that hnRNP A1 efficiently immunoprecipitated with the HIV-1 vRNA (22) and that siRNA-mediated knockdown of hnRNP A1 caused a dramatic decrease in HIV-1 structural protein, pr55Gag (herein termed Gag) expression and virus production with little effect on steady-state levels of the three HIV-1 RNA species (i.e. 9-, 4-, and 2-kb RNAs) (23). In this work, we show that HIV-1 ...

Translation initiation of the HIV-1 mRNA

“…10,126 Therefore, it is plausible that both viral and cellular proteins such as Rev and hRNP A1 are initially loaded on the viral mRNA in the nucleus before being used for cytoplasmic RNA expression. 118,127 In line with this possibility, a recent report confirms the cotranscriptional formation of a stable export complex that includes both Rev and CRM1. 128 It is noteworthy that Rev and associated proteins not only determine the export of the full-length mRNA, they also play a role in its translation.…”

“…The HIV-1 5 0 UTR IRES demonstrated activity in various biological systems such as HeLa based translation extracts, 86 in cells such as HeLa, U20S, HEK293-T, Jurkat T cells, 71,86,88,90,118,119 and in Xenopus laevis oocytes (see Fig. 2).…”

“…71 Using a similar strategy, Monette et al identified hnRNP A1 as a cellular factor that enhances HIV-1 5 0 UTR IRES function. 118 They further presented evidence that HIV-1 mRNA is loaded with hnRNP A1 in the cell nucleus and that the protein shuttles to the cytoplasm associated to the viral mRNA where it can exert its function at the level of IRES-mediated translation initiation. 118 As for nascent cellular mRNAs, the HIV-1 mRNA is expected to first encounter RNA-binding proteins in the nucleus.…”

“…118 They further presented evidence that HIV-1 mRNA is loaded with hnRNP A1 in the cell nucleus and that the protein shuttles to the cytoplasm associated to the viral mRNA where it can exert its function at the level of IRES-mediated translation initiation. 118 As for nascent cellular mRNAs, the HIV-1 mRNA is expected to first encounter RNA-binding proteins in the nucleus. 82 In fact, the full-length viral mRNA reaches the cytoplasm as part of a distinct ribonucleoprotein (RNP) complex with nuclear RNA-binding factors.…”

“…41 The impacts of some well known ITAFs have been assayed on translation initiation of the HIV-1 5 0 UTR IRES. 71,118,120 As such, Liu et al showed that eIF5A, the human Rev-interacting protein (hRIP) and DDX3 were all capable of stimulating translational activity from the HIV-1 5 0 UTR IRES when located in a bicistronic context in HEK293T cells. Interestingly, the same report discarded Sam68, a member of the signal transduction and activation of RNA (STAR) protein family, as a possible ITAF for the HIV-1 5 0 UTR IRES.…”