Point mutations in the C-terminus of HIV-1 gp160 reduce apoptosis and calmodulin binding without affecting viral replication

Abstract

One hallmark of AIDS progression is a decline in CD4+ T lymphocytes, though the mechanism is poorly defined. There is ample evidence that increased apoptosis is responsible for some, if not all, of the decline. Prior studies have shown that binding of cellular calmodulin to the envelope glycoprotein (Env) of HIV-1 increases sensitivity to fas-mediated apoptosis and that calmodulin antagonists can block this effect. We show that individual mutation of five residues in the C-terminal calmodulin-binding domain of Env is sufficient to significantly reduce fas-mediated apoptosis in transfected cells. The A835W mutation in the cytoplasmic domain of gp41 eliminated co-immunoprecipitation of Env with calmodulin in studies with stably transfected cells. Four point mutations (A835W, A838W, A838I, and I842R) and the corresponding region of HIV-1 HXB2 were cloned into the HIV-1 proviral vector pNL4-3 with no significant effect on viral production or envelope expression, although co-immunoprecipitation of calmodulin and Env was decreased in three of these mutant viruses. Only wild-type envelope-containing virus induced significantly elevated levels of spontaneous apoptosis by day 5 post-infection. Fas-mediated apoptosis levels positively correlated with the degree of calmodulin co-immunoprecipitation, with the lowest apoptosis levels occurring in cells infected with the A835W envelope mutation. While spontaneous apoptosis appears to be at least partially calmodulin-independent, the effects of HIV-1 Env on fas-mediated apoptosis are directly related to calmodulin binding.

Novel Recombinant Virus Assay for Measuring Susceptibility of Human Immunodeficiency Virus Type 1 Group M Subtypes To Clinically Approved Drugs

“…Standard deviations and coefficients of variation were within the ranges found for other recombinant virus assays, and changes were consistent with the scientific literature in terms of magnitude and specificity (15,17,31,34). The single-cycle format ensures very little sequence evolution and selection bias against less replication-competent variants and ensures that potential effects of mutations present in the recombinant virus on interaction with calmodulin or other cellular proteins involved in apoptosis will have no influence on the assay readout (26). An additional value lies in the potential to study recently described resistance mechanisms.…”

HIV and Chronic Obstructive Pulmonary Disease: Is It Worse and Why?

“…HIV-infected persons might have increased susceptibility to apoptosis because the HIV proteins Tat and Nef may induce endothelial cell apoptosis (49). It is also possible that HIV may cause apoptosis directly (50,51). It has been proposed that apoptosis, oxidative stress, and inflammation work in concert to promote COPD, and this scenario may be true in HIV-associated COPD as well (37).…”

Screening for Chronic Obstructive Pulmonary Disease (COPD) in an Urban HIV Clinic: A Pilot Study

“…[2][3][4] This increase in COPD is thought to be due to an increased prevalence of smoking in patients with HIV/ AIDS [5][6][7][8][9][10][11][12] in combination with a detrimental amplified inflammatory response in the lungs of those exposed to toxins in cigarette smoke. [13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29] Patients with HIV/AIDS also have increased barriers to smoking cessation, which adds to the risk for COPD over time. 30 Prior pulmonary infection with Pneumocystis jiroveci or other pathogens may also influence lung function.…”