蛋白丝氨酸/苏氨酸磷酸酶PP1在HIV Tat诱导的神经毒性过程中的应答机制

We are seeing an ever increasing number of HIV-associated neurocognitive disorders (HAND) due to the life lengthening of the people infected by HIV-1. However, the mechanism of HIV-1 Tat in inducing neurological dysfunction in more details remains unclear. N-Methyl-D-Aspartate receptors (NMDAR) is the major receptor involved in Tat-mediated neuronal toxicity and its activation can lead to function changes of several cellular proteins. As the Ca2+ modulator, PP1 can modulate synaptic plasticity through the receptors of cell membrane, therefore exerts the neuroprotective functions. We hypothesize that Tat may have an effect on the neurons through interaction with PP1 as well as regulation of PP1 activity. Our preliminary results showed that Tat interacts with PP1 in neuron cells of SH-SY5Y and affects the degradation, splicing or modification of PP1. In addition, the location of subtype B Tat was different from subtype C Tat in SH-SY5Y cells. Our results indicated that Tat may interact with PP1 and then change its location and function. Moreover, it exhibited a different pattern between subtype B Tat and subtype C Tat in modulating PP1 activity. The molecular mechanisms in the pathway of PP1 activity regulated by Tat need to be explored further. This study will reveal the mechanisms of neurotoxicity induced by Tat and provide strategy for developing new drugs of blocking HAND.

随着HIV感染者寿命不断延长，HIV相关性神经认知障碍（HAND）的发病率越来越高。HIV Tat蛋白在诱发神经功能障碍中发挥重要作用，但缺乏进一步分子机制研究。作为介导Tat引发神经毒性的主要受体—NMDARs的活化参与调控多种细胞蛋白的活性。其中对神经元有保护作用的Ca2+调控蛋白—PP1通过调控细胞膜受体功能影响突触可塑性。我们推测Tat可能通过调控PP1活性对神经元细胞功能产生影响。本室前期工作发现在神经元细胞SH-SY5Y中Tat与PP1相互作用，影响PP1剪切、降解或修饰；同时B、C亚型Tat在细胞中定位不同。因此Tat可能通过与PP1相互作用影响其细胞定位进而影响其功能，并且B、C亚型Tat对PP1调控存在差异。本研究在此基础上，进一步明确Tat对PP1调控的具体通路，探索其分子机制。从而阐明HIV Tat诱导神经毒性的分子机制，也为控制HAND的药物研发提供新的策略。

HIV相关性神经认知障碍（HAND）严重影响HIV感染者的生存质量，而HIV Tat蛋白在诱发神经功能障碍和神经元损伤中发挥重要作用，且具体分子机制复杂多样。本研究证实重组Tat1-72可以引起原代胎鼠神经元损伤，Tat1-72主要引起神经元树突长度变短，数目减少。Tat1-72可以调节PP1的表达，并通过降低Thr320磷酸化的水平来增强PP1活性。Tat1-72处理原代神经元细胞24小时，可以降低PP1底物CREB的活性，并降低CREB下游靶基因的转录水平（BDNF, c-fos, Egr-1）。综上，Tat1-72可能通过调控PP1的活性和CREB/BDNF信号通路损伤神经元。此外，使用Tat1-72蛋白处理小鼠的原代神经元细胞，通过基因芯片技术检测miRNA的变化。通过生物信息学分析发现，在差异表达的miRNA中，显著上调miR-128-3p与miR-181a-5p。使用多个数据库预测了两者共同的靶基因有9个，并通过蛋白互作分析锁定Tgfbr1，实验验证Tat蛋白能够引起神经元中Tgfbr1的mRNA与蛋白水平的下调，Tat蛋白通过下调的TGFBR1影响神经元活性。本研究丰富了HIV Tat诱导神经毒性的分子机制，也为控制HAND的药物研发提供新的策略。