Novel role of mortalin in attenuating HIV-1 Tat-mediated astrogliosis

Abstract

Background In human immunodeficiency virus-1 (HIV-1) infection, activation of astrocytes induces imbalance in physiological functions due to perturbed astrocytic functions that unleashes toxicity on neurons. This leads to inflammatory response finally culminating into neurocognitive dysfunction. In neuroAIDS, HIV-1 protein, transactivator of transcription (Tat) is detected in the cerebrospinal fluid of infected patients. Mortalin, a multifunctional protein, has anti-inflammatory role following its activation in various stress conditions. Recent studies demonstrate downregulation of mortalin in neurodegenerative diseases. Here, we explored the mechanisms of mortalin in modulating HIV-1 Tat-mediated neuroinflammation. Methods Expression of mortalin in autopsy section in normal and diseased individuals were examined using immunohistochemistry. To decipher the role of mortalin in HIV-1 Tat-induced activation, human fetal brain-derived astrocytes were transiently transfected with Tat and mortalin using expression vectors. HIV-1 Tat-mediated damage was analyzed using RT-PCR and western blotting. Modulatory role of mortalin was examined by coexpressing it with Tat, followed by examination of mitochondrial morphodynamics using biochemical assay and confocal and electron microscopy. Extracellular ATP release was monitored using luciferase assay. Neuroinflammation in astrocytes was examined using flow cytometry, dye based study, immunocytochemistry, immunoprecipitation, and western blotting. Indirect neuronal damage was also analyzed. Results HIV-1 Tat downregulates the expression of mortalin in astrocytes, and this is corroborated with autopsy sections of HIV-1 patients. We found that overexpression of mortalin with Tat reduced inflammation and also rescued astrocytic-mediated neuronal death. Using bioinformatics, we discovered that binding of mortalin with Tat leads to Tat degradation and rescues the cell from neuroinflammation. Blocking of proteosomal pathway rescued the Tat degradation and revealed the ubiquitination of Tat. Conclusion Overall, our data demonstrated the protective role of mortalin in combating HIV-1 Tat-mediated damage. We also showed that mortalin could degrade Tat through direct binding with HIV-1 Tat. Overexpression of mortalin in the presence of Tat could significantly reduce cytotoxic effects of Tat in astrocytes. Indirect neuronal death was also found to be rescued. Our in vitro findings were validated as we found attenuated expression of mortalin in the autopsy sections of HIV-1 patients.

Cal‘MAM’ity at the Endoplasmic Reticulum-Mitochondrial Interface: A Potential Therapeutic Target for Neurodegeneration and Human Immunodeficiency Virus-Associated Neurocognitive Disorders

“…For example, in astrocytes exposed to HIV-1, blocking mitochondrial Ca 2+ uptake by targeting MCU (Natarajaseenivasan et al, 2018), VDAC (Fatima et al, 2017), or cytosolic Ca 2+ (Nooka and Ghorpade, 2017) can prevent astrocyte mitochondrial dysfunction and reverse astrocyte-mediated neurotoxicity. The same outcome was achieved by overexpression of mortalin/grp75 (Priyanka et al, 2020). However, this was due to a direct interaction of mortalin/grp75 with HIV-1 Tat leading to Tat degradation rather than modulating ER-mitochondrial Ca 2+ transfer.…”

“…Moreover, knockdown of MCU to reduce mitochondrial Ca 2+ uptake during exposure to HIV-1 Tat and/or cocaine reverses astrocyte mitochondrial dysfunction and metabolic switching to restore a neuroprotective phenotype ( Natarajaseenivasan et al, 2018 ). Manipulation of VDAC1 (the OMM Ca 2+ channel within the MAM interface) or “mortalin” (aka grp75, the scaffolding protein between IP 3 R on the ER membrane and VDAC on the mitochondria) is also able to rescue neurons from HIV-1 Tat-induced astrocyte-mediated neurotoxicity ( Fatima et al, 2017 ; Priyanka et al, 2020 ). Specifically, HIV-1 Tat expressing primary human astrocytes trigger neuronal death by excessive ATP release, a mechanism that was counteracted by repression of VDAC1 ( Fatima et al, 2017 ).…”

“…Neuroinflammation and glutamate excitotoxicity are additional mechanisms for which astrocytes inflict neuronal damage during HAND, as previously reviewed by our research team ( Cisneros and Ghorpade, 2012 ; Borgmann and Ghorpade, 2015 ). Overexpression of mortalin/grp75 protects neurons from astrocyte-mediated neurotoxicity by reversing HIV-1 Tat-induced astrocyte mitochondrial dysfunction and fragmentation while also reducing the release of excess ATP, inflammatory cytokines, and extracellular glutamate ( Priyanka et al, 2020 ). Altogether, these findings strongly support MAM-mediated Ca 2+ transfer as a pivotal regulator of astrocyte-mediated neurotoxicity during HAND pathogenesis.…”

“…In astrocytes, both HIV-1 infection and external HIV-1 exposure perturb mitochondrial integrity ( Nooka and Ghorpade, 2017 ; Natarajaseenivasan et al, 2018 ; Ojeda et al, 2018 ; Priyanka et al, 2020 ). Unlike neurons, astrocytes increase their metabolic activity and ATP production ( Natarajaseenivasan et al, 2018 ; Swinton et al, 2019 ).…”

“…For example, HIV-1 Tat provokes astrocytes to undergo a distinct metabolic shift from glucose to fatty acid oxidation, which restricts astrocyte provision of lactate to neurons ( Natarajaseenivasan et al, 2018 ). In addition to decreasing release of neurotrophic factors when ‘activated’ by HIV-1 Tat, astrocytes increase their release of neurotoxic factors including excessive ATP ( Fatima et al, 2017 ; Priyanka et al, 2020 ), ROS ( Natarajaseenivasan et al, 2018 ; Priyanka et al, 2020 ), and inflammatory cytokines ( Natarajaseenivasan et al, 2018 ). As briefly discussed in the previous section, blocking ER-mitochondrial Ca 2+ transfer via VDAC (on the OMM) ( Fatima et al, 2017 ), MCU (on the IMM) ( Natarajaseenivasan et al, 2018 ), or mortalin/grp75 (scaffold between IP 3 R and VDAC) ( Priyanka et al, 2020 ) restored an astrocyte neurotrophic phenotype, highlighting a probable MAM interplay in astrocyte-mediated neurotoxicity during HAND.…”