Molecular mimicry of human tRNA^Lys anti-codon domain by HIV-1 RNA genome facilitates tRNA primer annealing

Abstract

The primer for initiating reverse transcription in human immunodeficiency virus type 1 (HIV-1) is tRNA Lys3. Host cell tRNA Lys is selectively packaged into HIV-1 through a specific interaction between the major tRNA Lys -binding protein, human lysyl-tRNA synthetase (hLysRS), and the viral proteins Gag and GagPol. Annealing of the tRNA primer onto the complementary primerbinding site (PBS) in viral RNA is mediated by the nucleocapsid domain of Gag. The mechanism by which tRNA Lys3 is targeted to the PBS and released from hLysRS prior to annealing is unknown. Here, we show that hLysRS specifically binds to a tRNA anti-codon-like element (TLE) in the HIV-1 genome, which mimics the anti-codon loop of tRNA Lys and is located proximal to the PBS. Mutation of the U-rich sequence within the TLE attenuates binding of hLysRS in vitro and reduces the amount of annealed tRNA Lys3 in virions. Thus, LysRS binds specifically to the TLE, which is part of a larger LysRS binding domain in the viral RNA that includes elements of the Psi packaging signal. Our results suggest that HIV-1 uses molecular mimicry of the anti-codon of tRNA Lys to increase the efficiency of tRNA Lys3 annealing to viral RNA.

Anticodon-like binding of the HIV-1 tRNA-like element to human lysyl-tRNA synthetase

“…Interestingly, the tRNA mimicry of the tRNA-like element (TLE) extends to the global fold of the PBS/TLE region of the HIV-1 genome, which resembles the L-shaped tRNA fold (Jones et al 2014). Taken together, our previous data are consistent with a model wherein tRNA Lys3 is at least partially released from hLysRS through competition for hLysRS binding with the viral TLE (Jones et al 2013).…”

“…In particular, we reported that a U-rich tRNA Lys anticodon-like sequence in the gRNA proximal to the PBS interacts with hLysRS with relatively high affinity (Jones et al 2013). Importantly, mutation of the U-rich sequence had a significant impact on tRNA primer placement and HIV-1 infectivity (Jones et al 2013).…”

“…In particular, we reported that a U-rich tRNA Lys anticodon-like sequence in the gRNA proximal to the PBS interacts with hLysRS with relatively high affinity (Jones et al 2013). Importantly, mutation of the U-rich sequence had a significant impact on tRNA primer placement and HIV-1 infectivity (Jones et al 2013). Interestingly, the tRNA mimicry of the tRNA-like element (TLE) extends to the global fold of the PBS/TLE region of the HIV-1 genome, which resembles the L-shaped tRNA fold (Jones et al 2014).…”

“…We hypothesize that the ACB interacts with the U-rich TLE in the gRNA in a manner that mimics its interaction with the tRNA Lys anticodon domain. The high specificity of this interaction along with additional interactions between hLysRS and gRNA outside of the ACB interaction (Jones et al 2013) contribute to primer release from LysRS and targeting to the PBS. As a first step toward testing this hypothesis, we used NMR to map the interaction between the hLysRS ACB and a 23-nt RNA stem-loop derived from the wild-type (WT) HIV-1 TLE (Fig.…”

Distinct binding interactions of HIV-1 Gag to Psi and non-Psi RNAs: Implications for viral genomic RNA packaging

“…RNAs were fluorescently labeled with fluorescein on their 3 ′ ends as previously described (Pagano et al 2007;Jones et al 2013). Concentrations and labeling efficiencies were determined using the following extinction coefficients-fluorescein, 8. .…”