【作者】 谭震球；

【导师】 赵颖俊；

【作者基本信息】 厦门大学 ， 生理学， 2021， 博士

【摘要】 溶质转运蛋白 SLC25A38(Solute Carrier Family 25 Member 38)属于线粒体载体家族,由细胞核基因编码表达。有研究表明SLC25A38在血红素生物合成过程中发挥了重要作用。在阿尔茨海默病(Alzheimer’s Disease,AD)和其它相关的tau疾病病理发展过程中,SLC25A38的增加可激活caspase-3、使其剪切tau蛋白并加速tau的沉积,增强其神经毒性。然而目前并不清楚SLC25A38的神经生理学功能及其在突触损伤中的作用。在本研究中,我们构建了 Slc25a38敲除小鼠模型,发现Slc25a38完全缺失导致小鼠胚胎死亡,而Slc25a38单倍体缺失小鼠(Slc25a38+/-,HET)具有正常的生长和发育能力。行为学实验结果表明,与对照小鼠(Slc25a38+/+,WT)相比,HET小鼠具有正常的学习能力,但记忆消退减缓;电生理实验结果表明Slc25a38单倍体缺失不影响急性海马脑片的长时程增强(Long-term potentiation,LTP),但是降低了 NMDAR(N-methyl-D-aspartate receptor)依赖性的长时程抑制(Long-term depression,LTD);此外,与WT小鼠相比,经NMDA处理的HET小鼠急性海马脑片和神经元细胞表面的AMPAR(α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor)去磷酸化减弱、内吞作用受损。免疫沉淀质谱分析鉴定了 190多个可能与SLC25A38具有相互作用的蛋白,进一步KEGG通路分析表明SLC25A38潜在相互作用蛋白参与神经退行性疾病病变、细胞凋亡以及钙离子信号通路等生物学过程,基因本底分析表明多个SLC25A38潜在相互作用蛋白在突触传递调节功能通路富集,该通路包含蛋白磷酸酶 2B(protein phosphatase 2B,PP2B,又名 calcineurin)的亚基 PPP3CB(Protein phosphatase 3 catalytic subunit beta 和 PPP3CA(Protein phosphatase 3 catalytic subunit alpha);免疫共沉淀实验结果显示SLC25A38能与PP2B相互作用并影响PP2B在线粒体的分布,提示SLC25A38可能通过PP2B调控AMPAR的转运,进而调节突触功能。在AD的病变过程中,可溶性Aβ(amyloid-β)寡聚体能增强LTD、损伤突触结构和认知功能。在本研究中,我们发现Slc25a38的单倍体缺失能够缓解Aβ诱导的小鼠空间学习/记忆损伤,进一步的研究表明Slc25a38单倍体缺失能够抑制Aβ诱导的NMDAR-LTD以及突触结构损伤,但是并没有改变APP(Amyloid precursor protein)蛋白的淀粉样水解和Aβ水平。综合以上研究结果,我们首次发现SLC25A38可能通过与PP2B相互作用调节AMPAR的去磷酸化和胞内转运,进而调控NMDAR依赖性的LTD和小鼠记忆消退;此外,Slc25a38单倍体缺失能够缓解Aβ诱导的认知功能以及突触损伤,Slc25a38或许可以成为AD的一个潜在治疗靶点。总之,我们的研究发现了一种新的调控记忆消退和Aβ突触毒性的机制。更多还原

【Abstract】 Solute transport protein SLC25A38(solute carrier family 25 member 38)belongs to mitochondrial carrier family and is encoded by nucleus genome.SLC25A38 plays an important role in heme biosynthesis.Additionally,SLC25A38 can regulate caspase-3-mediated neurotoxicity and tau cleavage in the pathogenesis of Alzheimer’s disease(AD)and other related tauopathies.However,the detailed role of SLC25A38 in neurophysiological function and synaptic impairment remains unclear.In the present study,we find that deficiency of Slc25a38(Slc25a38-/-,K0 mice)resulted in embryonic lethality at E17.5.The SLC25A38 haploinsufficient mice(Slc25a38+/-,HET)grow normally.Interestingly,we found that HET mice exhibited normal learning ability but slower memory decay,when compared to wild type mice(Slc25a38+/+,WT).Results from electrophysiological recordings showed that SLC25A38 haplodeficiency reduced NMDAR(N-methyl-D-aspartate receptor)-dependent long term depression(LTD),but did not affect long term potentiation(LTP).In addition,NMDA-induce d AMPAR(α-amino-3-hydroxy-5-methyl-4-isoxazole-propionicacid receptor)dephosphorylation and endocytosis were impaired acute hippocampal slices and cultured primary neurons derived from HET mice.To further determine how SLC25A38 regulates synaptic plasticity,we carried out immunoprecipitation-mass spectrometry and analyzed the SLC25A38 interactome data.One hundred ninety potential interacting components were specifically identified by proteomic analysis;KEGG(Kyoto Encyclopedia of Genes and Genomes)pathway analysis indicates a number of components related to neurodegeneration as well as apoptosis and calcium signaling.G0(Gene ontology)analysis for related biological process(BP)shows that many potential SLC25A38 interacting proteins are enriched in the“regulation of synaptic transmission”category,including PP2B subunits PPP3CB and PPP3CA.The interaction between SLC25A38 and PP2B was confirmed by co-immunoprecipitation assay.In addition,biochemical fractionation assays showed that SLC25A38 can regulate distribution of PP2B to mitochondria.Together,these results suggest that SLC25A38 may regulate AMPAR dephosphorylation and trafficking,and synaptic plasticity via its interaction with PP2B.During AD pathogenesis,soluble Aβ oligomers can enhance LTD and impair cognitive function and synaptic structures.Here,we found that oligomeric Aβ-induced spatial memory deficit was partially rescued in HET mice.Further,the enhancement of NMDAR-dependent LTD and the impairment of synaptic structures triggered by oligomeric Aβ treatment was abolished by Slc25a38 haploinsufficieny.However,Slc25a38 haploinsufficieny doesn’t alter amyloidogenic proteolysis of beta amyloid precursor protein(APP)and Aβ levels.In sum,our present work uncovers that SLC25A38 may regulate AMPAR dephosphorylation and trafficking through interacting with PP2B,and subsequently modulate NMDAR-dependent LTD and memory decay.In addition,Slc25a38 haploinsufficieny can protect against Aβ-induced synaptic and cognitive deficits,suggesting that SLC25A38 may be a potential therapeutic target for AD.Our study uncovers a novel mechanism for memory extinction and Aβ-induced synaptotoxicity.更多还原