HIV-1 Tat protein inhibits neurosecretion by binding to phosphatidylinositol 4,5-bisphosphate

Abstract

Summary HIV-1 transcriptional activator (Tat) enables viral transcription and is also actively released by infected cells. Extracellular Tat can enter uninfected cells and affect some cellular functions. Here, we examine the effects of Tat protein on the secretory activity of neuroendocrine cells. When added to the culture medium of chromaffin and PC12 cells, Tat was actively internalized and strongly impaired exocytosis as measured by carbon fiber amperometry and growth hormone release assay. Expression of Tat mutants that do not bind to phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P 2 ] did not affect secretion, and overexpression of phosphatidylinositol 4-phosphate 5-kinase (PIP5K), the major PtdIns(4,5)P 2 synthesizing enzyme, significantly rescued the Tat-induced inhibition of neurosecretion. This suggests that the inhibition of exocytosis may be the consequence of PtdIns(4,5)P 2 sequestration. Accordingly, expression of Tat in PC12 cells interfered with the secretagogue-dependent recruitment of annexin A2 to the plasma membrane, a PtdIns(4,5)P 2 -binding protein that promotes the formation of lipid microdomains that are required for exocytosis. In addition Tat significantly prevented the reorganization of the actin cytoskeleton necessary for the movement of secretory vesicles towards plasma membrane fusion sites. Thus, the capacity of extracellular Tat to enter neuroendocrine cells and sequester plasma membrane PtdIns(4,5)P 2 perturbs several PtdIns(4,5)P 2 -dependent players of the exocytotic machinery, thereby affecting neurosecretion. We propose that Tat-induced inhibition of exocytosis is involved in the neuronal disorders associated with HIV-1 infection.

HIV-Tat induces a decrease in I Kr and I Ks via reduction in phosphatidylinositol-(4,5)-bisphosphate availability

“…In neurosecretory cells, it has been proposed that Tat perturbs the recruitment of several PIP 2 -dependent key players of the exocytotic machinery [32]. Combined with this previous investigation, our work suggests a general remodeling of PIP 2 -dependent processes by HIV via Tat.…”

“…cotransfection of hERG with either WT-or W11Y-Tat/GFP bidirectional plasmids. They confirmed that WT-Tat localizes at the COS-7 plasma membrane, through its interaction with PIP 2 [32]. Unlike WT-Tat, W11Y-Tat localized diffusely in the cytosol, its lack of affinity for PIP 2 preventing its plasma membrane localization ( Figure 3A & B).…”

“…We first studied the effect of Tat 24-h incubation in HEK293 cells stably expressing hERG channels. We chose a 24-h incubation since in the different biological assays in which Tat had been tested (neurosecretion [32] and phagocytosis [33]), it was found to require some time (3-4 hours) for Tat, after initial internalization, to accumulate on PIP 2 and to perturb processes relying on this phosphoinositide. Moreover such a 24-h incubation, and not acute application, had already been associated with a decrease in hERG current density in this model [16].…”

HIV-1 Tat inhibits phagocytosis by preventing the recruitment of Cdc42 to the phagocytic cup

“…It should be noted that Tat targets in these biological systems are different, Annexin A2 (ref. 54) for neurons/exocytosis and Cdc42 (this study) for macrophages/ phagocytosis. The capacity of Tat to perturb these activities is also different since subnanomolar concentrations of Tat are sufficient to efficiently inhibit phagocytosis, while 5-10 nM of Tat are required to affect the neurosecretory activity 54 .…”

“…Tat is an original toxin in this field since it affects Cdc42 function by preventing its recruitment to the phagocytic cup, thereby preventing its activation. Hence, Tat is a PI(4,5)P 2 targeting toxin that can affect the neurosecretory 54 , the phagocytic (this study) and probably other PI(4,5)P 2 -dependent membrane traffic machineries. It should be noted that Tat targets in these biological systems are different, Annexin A2 (ref.…”