Two cellular proteins that interact with a stem loop in the simian hemorrhagic fever virus 3′(+)NCR RNA

Maines, Taronna R.; Young, Mary; Dinh, Nikita Nhu-Nguyen; Brinton, Margo A.

Both full-length and subgenomic negative-strand RNAs are initiated at the 3′ terminus of the positive-strand genomic RNA of the arterivirus, simian hemorrhagic fever virus (SHFV). The SHFV 3′(+) non-coding region (NCR) is 76 nts in length and forms a stem loop (SL) structure that was confirmed by ribonuclease structure probing. Two cell proteins, p56 and p42, bound specifically to a probe consisting of the SHFV 3′(+)NCR RNA. The 3′(+)NCR RNAs of two additional members of the arterivirus genus specifically interacted with two cell proteins of the same size. p56 was identified as polypyrimidine tract-binding protein (PTB) and p42 was identified as fructose bisphosphate aldolase A. PTB binding sites were mapped to a terminal loop and to a bulged region of the SHFV 3′SL structure. Deletion of either of the PTB binding sites in the viral RNA significantly reduced PTB binding activity, suggesting that both sites are required for efficient binding of this protein. Changes in the top portion of the SHFV 3′SL structure eliminated aldolase binding, suggesting that the binding site for this protein is located near the top of the SL. These cell proteins may play roles in regulating the functions of the genomic 3′ NCR.