X-I and X-II open reading frames of HTLV-I are not required for virus replication or for immortalization of primary T-cells in vitro

In contrast to other retroviruses of the oncovirinae subgroup, the primate and bovine leukemia viruses (HTLV, STLV, and BLV) encode genes in the X-region of the genome, between the env gene and the 3' long terminal repeat. In HTLV-I, two overlapping open reading frames (ORFs) in the distal half of the X-region encode tax and rex genes, while two ORFs (X-I and X-II) in the proximal half of this region potentially encode proteins designated p12(XI) (or roil and p30(XII) (or tof). The biological functions and mechanisms of tax and rex have been studied extensively whereas the roles of the other ORFs have not yet been established. To identify possible functions for ORFs X-I and X-II, an infectious molecular clone of HTLV-I and a mutant provirus lacking these ORFs were compared with respect to virus replication, gene expression, and ability to immortalize primary T-cells. When transiently transfected into 293 cells, both intact and deleted proviruses directed the synthesis of virus mRNAs and proteins that were quantitatively and qualitatively identical. These viruses were also indistinguishable in their abilities to infect and replicate in DBS-FRhL cells, which are permissive for HTLV-I propagation. Immortalized T-cell lines were established after cell-free or coculture methods for infection of activated, human peripheral blood or cord blood lymphocytes with each of the cloned viruses. The growth kinetics, cytokine dependence, and cell surface markers of the infected T-cell cultures were similar for each provirus clone. Thus, ORFs X-I and X-II are not essential for virus infectivity, replication, gene expression, or T-cell immortalization in vitro.