Mutation of Tyrosine 470 of Human Dopamine Transporter is Critical for HIV-1 Tat-Induced Inhibition of Dopamine Transport and Transporter Conformational Transitions

Abstract

HIV-1 Tat protein plays a crucial role in perturbations of the dopamine (DA) system. Our previous studies have demonstrated that Tat decreases DA uptake, and allosterically modulates DA transporter (DAT) function. In the present study, we have found that Tat interacts directly with DAT, leading to inhibition of DAT function. Through computational modeling and simulations, a potential recognition binding site of human DAT (hDAT) for Tat was predicted. Mutation of tyrosine470 (Y470H) attenuated Tat-induced inhibition of DA transport, implicating the functional relevance of this residue for Tat binding to hDAT. Y470H reduced the maximal velocity of [3H]DA uptake without changes in the Km and IC50 values for DA inhibition of DA uptake but increased DA uptake potency for cocaine and GBR12909, suggesting that this residue does not overlap with the binding sites in hDAT for substrate but is critical for these inhibitors. Furthermore, Y470H also led to transporter conformational transitions by affecting zinc modulation of DA uptake and WIN35,428 binding as well as enhancing basal DA efflux. Collectively, these findings demonstrate Tyr470 as a functional recognition residue in hDAT for Tat-induced inhibition of DA transport and transporter conformational transitions. The consequence of mutation at this residue is to block the functional binding of Tat to hDAT without affecting physiological DA transport.

Protective effects and mechanisms of Ndfipl on SH-SY5Y cell apoptosis in an in vitro Parkinson’s disease model

“…It can penetrate the blood brain barrier and convert to toxic MPP + under the action of monoamine oxidase B. MPP + is very similar to dopamine in chemical structure, and can act on SNpc through dopamine transporter to further recruit MPP + to the mitochondria; thus, impeding the role of respiratory chain enzyme complex I, energy crisis, and cell death (Midde et al, 2013;Velázquez-Sánchez et al, 2013;Dehnes et a., 2014;Schwarz et al, 2014). Additionally, MPP + is able to inhibit the activity of tyrosine hydroxylase, thus reducing the synthesis of dopamine.…”

Exposure to HIV-1 Tat in brain impairs sensorimotor gating and activates microglia in limbic and extralimbic brain regions of male mice

“…Prepulse inhibition is known to be attenuated by dopamine agonists in mice, particularly those with preference for D1-like receptors [46]. Tat is known to allosterically modulate the dopamine transporter [47], elevating synaptic dopamine concentrations [48] and increasing the efficacy of binding ligands [49]. Tat-induced enhancement of dopamine levels could further promote the deficits observed in PPI presently.…”

Modeling Deficits in Attention, Inhibition, and Flexibility in HAND

“…Reduced DA transporter (DAT) levels in the putamen and ventral striatum are found in HIV-1+ individuals with cognitive impairment (Wang et al 2004; Chang et al , 2008). In vitro studies have shown that DAT is targeted by HIV-1 proteins Tat and gp120, resulting in transporter impairment (Aksenov et al 2008; Ferris et al 2009; Midde et al 2013; Zhu et al 2009; Zhu et al 2011), due to direct protein-protein interactions (Zhu et al 2009) involving an allosteric modulation of DAT by the Tat protein (Zhu et al 2011). In addition, DA-dependent signaling has been identified as a mechanism of HIV-1 protein neurotoxicity (Aksenova et al 2006; Silvers et al 2007; Wallace et al 2006).…”