pLVX-IRES-Puro

Human myxovirus resistance protein B (hMXB) is a restriction factor for HIV-1 and also inhibits a variety of retroviruses. However, hMXB is not resistant to equine infectious anemia virus (EIAV). Studies here show that equine MX2 (eMX2) effectively restricts EIAV in vitro. Additionally, eMX2 inhibits HIV-1 and other lentiviruses, including murine leukemia virus. Previously, it was reported that hMXB repression was reduced in hMXB Δ1-25 but not in the GTP-binding mutant K131A and the GTP-hydrolyzing mutant T151A. In contrast to this phenomenon, this study shows that eMX2 restriction is not reduced in eMX2 Δ1-25 but is reduced in eMX2 K127A and T147A, which correspond to hMXB K131A and T151A, respectively. Therefore, eMX2 may inhibit retroviral replication through a novel mechanism distinct from hMXB.

In this study, wild-type human and equine MX2 as well as mutants were expressed from the lentiviral vector pLVX-IRES-Puro. Stable MX protein-expressing cell lines were generated by transduction with lentiviral vectors by transfection of pLVX-IRES-Puro (empty vector) or pLVX-IRES-Puro encoding wild-type hMXB, eMX2, or mutants, together with psPAX2, pRSV-Rev and pMD.G. Puromycin-resistant cell pools were tested for MX protein expression by immunoblotting.

Figure 1. Stable expression of hMXB, hMXB Δ1–25, or eMX2 in HeLa cells transduced with lentiviral MX expression vector or pLVX-IRES-Puro (empty vector). MX expression was detected by immunoblotting with anti-FLAG antibody. (Meier, K., et al. 2018)