The Role of Human Dopamine Transporter in NeuroAIDS

HIV-associated neurocognitive disorder (HAND) remains highly prevalent in HIV infected individuals and represents a special group of neuropathological disorders, which are associated with HIV-1 viral proteins, such as transactivator of transcription (Tat) protein. Cocaine abuse increases the incidence of HAND and exacerbates its severity by enhancing viral replication. Perturbation of dopaminergic transmission has been implicated as a risk factor of HAND. The presynaptic dopamine (DA) transporter (DAT) is essential for DA homeostasis and dopaminergic modulation of the brain function including cognition. Tat and cocaine synergistically elevate synaptic DA levels by acting directly on human DAT (hDAT), ultimately leading to dysregulation of DA transmission. Through integrated computational modeling and experimental validation, key residues have been identified in hDAT that play a critical role in Tat-induced inhibition of DAT and induce transporter conformational transitions. This review presents current information regarding neurological changes in DAT-mediated dopaminergic system associated with HIV infection, DAT-mediated adaptive responses to Tat as well as allosteric modulatory effects of novel compounds on hDAT. Understanding the molecular mechanisms by which Tat induces DAT-mediated dysregulation of DA system is of great clinical interest for identifying new targets for an early therapeutic intervention for HAND

A Comparative Study of Survival, Metabolism, Immune Indicators, and Proteomics, in Five Batches of Japanese Scallop Mizuhopecten yessoensis under Short-Term High Temperature Stress

Five batches of the Japanese scallop Mizuhopecten pyessoensis were tested for survival rate, oxygen consumption, catalase (CAT) and superoxide dismutase (SOD) activities, total antioxidant capacities (T-AOC) contents, and proteomics under short-term high temperature conditions. The five batches, (W1, W2, W3, W4, W5) selected from the established 21 ‘ivory white’ M. yessoensis batches, had higher survival rates than the other batches after one year of culture. Initial rearing water temperature of 15°C was increased by 1°C per day with a cooling and heating system. The temperature was raised until over 50% of the scallops from 3 batches died. This occurred at 30°C. The higher than normal culture temperature conditions showed significant or highly significant differences in the responses of some of the batches. Some showed significantly higher survival rates and significantly different rates of oxygen consumption. CAT activity, SOD activity and T-AOC content was similar in the five batches, and all three indices were significantly lower in W3 and W5 than in the other batches (P<0.01). Expression patterns of MDA content were opposite to those of CAT activity, SOD activity and T-AOC content. Protein profiles of all five batches were similar; the sizes of the predominant bands ranged from 20-110 kDa. We identified twenty-eight proteins with high scores in the database. These included heat shock proteins (HSPs), glucose-regulated protein 94, and arginine kinase

Regulating Rumination by Anger: Evidence for the Mutual Promotion and Counteraction (MPMC) Theory of Emotionality

Unlike the strategy of cognitive regulation that relies heavily on the top-down control function of the prefrontal cortex (PFC), which was recently found may be critically impaired in stressful situations, traditional Chinese philosophy and medicine views different types of emotionality as having mutual promotion and counteraction (MPMC) relationships, implying a novel approach that requires less cognition to emotional regulation. Actually, our previous studies have indicated that anger responses could be successfully regulated via the induction of sadness, and this efficiency could not be influenced by stress, thus providing evidences for the hypothesis of “sadness counteracts anger” (SCA) proposed by the MPMC theory of emotionality (Zhan et al., 2015, 2017). In this study, we experimentally examined the MPMC hypothesis that “anger counteracts rumination” (ACR) which postulates that rumination may be alleviated by the anger emotion. In Study 1, all participants were initially caused state rumination and then induced anger, joy or neutral mood, the results showed that the rumination-related affect was alleviated after anger induction relative to that after joy or neutral mood induction. In Study 2, female participants with high trait rumination were recruited and divided into two groups for exposure to an anger or neutral emotion intervention, the result indicated that the anger intervention group exhibited a greater decline in trait rumination than the neutral emotion intervention group. These findings provided preliminary evidence supporting the hypothesis of ACR, which suggested a new strategy that employs less cognitive resources to regulating state and trait rumination by inducing anger

Role of Histidine 547 of Human Dopamine Transporter in Molecular Interaction with HIV-1 Tat and Dopamine Uptake

HIV-1 Tat plays an important role in HIV-associated neurocognitive disorders (HAND) by disrupting neurotransmission including dopamine uptake by human dopamine transporter (hDAT). Previous studies have demonstrated that HIV-1 Tat directly binds to hDAT and some amino-acid mutations that attenuate the hDAT-Tat binding also significantly decreased dopamine uptake activity of hDAT. This combined computational-experimental study demonstrates that histidine-547 (H547) of hDAT plays a crucial role in the hDAT-Tat binding and dopamine uptake by hDAT, and that the H547A mutation can not only considerably attenuate Tat-induced inhibition of dopamine uptake, but also significantly increase the Vmax of hDAT for dopamine uptake. The finding of such an unusual hDAT mutant capable of both increasing the Vmax of hDAT for dopamine uptake and disrupting the hDAT-Tat binding may provide an exciting knowledge basis for development of novel concepts for therapeutic treatment of the HAND

Molecular Mechanism: The Human Dopamine Transporter Histidine 547 Regulates Basal and HIV-1 Tat Protein-Inhibited Dopamine Transport

Abnormal dopaminergic transmission has been implicated as a risk determinant of HIV-1-associated neurocognitive disorders. HIV-1 Tat protein increases synaptic dopamine (DA) levels by directly inhibiting DA transporter (DAT) activity, ultimately leading to dopaminergic neuron damage. Through integrated computational modeling prediction and experimental validation, we identified that histidine547 on human DAT (hDAT) is critical for regulation of basal DA uptake and Tat-induced inhibition of DA transport. Compared to wild type hDAT (WT hDAT), mutation of histidine547 (H547A) displayed a 196% increase in DA uptake. Other substitutions of histidine547 showed that DA uptake was not altered in H547R but decreased by 99% in H547P and 60% in H547D, respectively. These mutants did not alter DAT surface expression or surface DAT binding sites. H547 mutants attenuated Tat-induced inhibition of DA transport observed in WT hDAT. H547A displays a differential sensitivity to PMA- or BIM-induced activation or inhibition of DAT function relative to WT hDAT, indicating a change in basal PKC activity in H547A. These findings demonstrate that histidine547 on hDAT plays a crucial role in stabilizing basal DA transport and Tat-DAT interaction. This study provides mechanistic insights into identifying targets on DAT for Tat binding and improving DAT-mediated dysfunction of DA transmission

Allosteric Modulatory Effects of SRI-20041 and SRI-30827 on Cocaine and HIV-1 Tat Protein Binding to Human Dopamine Transporter

Dopamine transporter (DAT) is the target of cocaine and HIV-1 transactivator of transcription (Tat) protein. Identifying allosteric modulatory molecules with potential attenuation of cocaine and Tat binding to DAT are of great scientific and clinical interest. We demonstrated that tyrosine 470 and 88 act as functional recognition residues in human DAT (hDAT) for Tat-induced inhibition of DA transport and transporter conformational transitions. Here we investigated the allosteric modulatory effects of two allosteric ligands, SRI-20041 and SRI-30827 on cocaine binding on wild type (WT) hDAT, Y470 H and Y88 F mutants. Effect of SRI-30827 on Tat-induced inhibition of [3H]WIN35,428 binding was also determined. Compared to a competitive DAT inhibitor indatraline, both SRI-compounds displayed a similar decrease (30%) in IC50 for inhibition of [3H]DA uptake by cocaine in WT hDAT. The addition of SRI-20041 or SRI-30827 following cocaine slowed the dissociation rate of [3H]WIN35,428 binding in WT hDAT relative to cocaine alone. Moreover, Y470H and Y88F hDAT potentiate the inhibitory effect of cocaine on DA uptake and attenuate the effects of SRI-compounds on cocaine-mediated dissociation rate. SRI-30827 attenuated Tat-induced inhibition of [3H]WIN35,428 binding. These observations demonstrate that tyrosine 470 and 88 are critical for allosteric modulatory effects of SRI-compounds on the interaction of cocaine with hDAT

Mutational Effects of Human Dopamine Transporter at Tyrosine88, Lysine92, and Histidine547 on Basal and HIV-1 Tat-Inhibited Dopamine Transport

Dysregulation of dopaminergic system induced by HIV-1 Tat protein-mediated direct inhibition of the dopamine transporter (DAT) has been implicated as a mediating factor of HIV-1 associated neurocognitive disorders. We have reported that single point mutations on human DAT (hDAT) at tyrosine88 (Y88F), lysine92 (K92M), and histidine547 (H547A) differentially regulate basal dopamine uptake but diminish Tat-induced inhibition of dopamine uptake by changing dopamine transport process. This study evaluated the effects of double (Y88F/H547A) and triple (Y88F/K92M/H547A) mutations on basal dopamine uptake, Tat-induced inhibition of DAT function, and dynamic transport process. Compared to wild-type hDAT, the Vmax values of [3H]Dopamine uptake were increased by 96% in Y88F/H547A but decreased by 97% in Y88F/K92M/H547A. [3H]WIN35,428 binding sites were not altered in Y88F/H547A but decreased in Y88F/K92M/H547A. Y88F/H547A mutant attenuated Tat-induced inhibition of dopamine uptake observed in wild-type hDAT. Y88F/H547A displayed an attenuation of zinc-augmented [3H]WIN35,428 binding, increased basal dopamine efflux, and reduced amphetamine-induced dopamine efflux, indicating this mutant alters transporter conformational transitions. These findings further demonstrate that both tyrosine88 and histidine547 on hDAT play a key role in stabilizing basal dopamine transport and Tat-DAT integration. This study provides mechanistic insights into developing small molecules to block multiple sites in DAT for Tat binding