Abstract
HIV-1 encodes an envelope glycoprotein (Env) that contains a long cytoplasmic tail (CT) harboring trafficking motifs implicated in Env incorporation into virus particles and viral transmission. In most physiologically relevant cell types, the gp41 CT is required for HIV-1 replication, but in the MT-4 T-cell line the gp41 CT is not required for a spreading infection. To help elucidate the role of the gp41 CT in HIV-1 transmission, in this study we investigated the viral and cellular factors that contribute to the permissivity of MT-4 to gp41 CT truncation. We found that the kinetics of HIV-1 production are faster in MT-4 than in the other T-cell lines tested, but MT-4 express equivalent amounts of HIV-1 proteins on a per-cell basis relative to cells not permissive to CT truncation. MT-4 express higher levels of plasma-membrane-associated Env than non-permissive cells and Env internalization from the plasma membrane is slower compared to another T-cell line, SupT1. Paradoxically, despite the high levels of Env on the surface of MT-4, two-fold less Env is incorporated into virus particles in MT-4 compared to SupT1. Cell-to-cell transmission between co-cultured 293T and MT-4 is higher than in co-cultures of 293T with most other T-cell lines tested, indicating that MT-4 are highly susceptible to this mode of infection. These data help to clarify the long-standing question of how MT-4 cells overcome the requirement for the HIV-1 gp41 CT and support a role for gp41 CT-dependent trafficking in Env incorporation and cell-to-cell transmission in physiologically relevant cell lines.
Importance The HIV-1 Env cytoplasmic tail (CT) is required for efficient Env incorporation into nascent particles and viral transmission in primary CD4+ T cells. The MT-4 T-cell line has been reported to support multiple rounds of infection of HIV-1 encoding a gp41 CT truncation. Uncovering the underlying mechanism of MT-4 T-cell line permissivity to gp41 CT truncation would provide key insights into the role of the gp41 CT in HIV-1 transmission. This study reveals that multiple factors contribute to the unique ability of a gp41 CT truncation mutant to spread in cultures of MT-4 cells. The lack of a requirement for the gp41 CT in MT-4 is associated with the combined effects of rapid HIV-1 protein production, high levels of cell-surface Env expression, and increased susceptibility to cell-to-cell transmission compared to non-permissive cells.
List of abbreviations
- CT
- cytoplasmic tail
- VS
- virological synapse
- C-C
- cell-to-cell
- Env
- envelope glycoprotein
- STR
- short tandem repeat
- MFI
- median fluorescence intensity
- PM
- plasma membrane
- Radio-IP
- radio-immunoprecipitation
- hPBMC
- human peripheral blood mononuclear cells
- RT
- reverse transcriptase
- VSV-G
- Vesicular stomatitis virus glycoprotein
- PBS
- phosphate buffered saline
- FBS
- fetal bovine serum
- WT
- wild type
- HRP
- horseradish peroxidase
- ER
- endoplasmic reticulum
- MA
- HIV-1 Matrix protein
- EBV
- Epstein-Barr virus
- HTLV
- Human T-cell leukemia virus
- ALL
- Acute lymphocytic leukemia
- ATL
- Adult T-cell lymphoma
- ATCC
- American Type Culture Collection
- NIH ARP
- NIH AIDS Reagent Program
- n.s.
- not statistically significant
- IN
- HIV-1 Integrase protein
- CA
- HIV-1 Capsid protein
- RIN
- RNA Integrity Number