The Role Of PSAP On SOD1^G93A-Induced Apoptosis

ALS?Amyotrophic Lateral Syndrome?is a progressive neural degenerative disease that accumulates damage from upper motor neurons to lower motor neurons.The pathogenesis is that death motor neurons prevent the brain from controlling the movement of muscles and muscles finally shrink as the result of lacking of exercise.Patients in late stage will lose their mobility completely and eventually die of respiratory failure caused by respiratory muscle paralysis.ALS is divided into two categories which include familial ALS?FALS?and sporadic ALS?SALS?.The percentages of FASL and SALS account for the total number of the cases are 5%¹0%and 90%respectively.Mutations in the SOD1 gene were found in both 20%of FLAS and 5%of SALS.SOD1?Cu,Zn-SOD?which is one of the important antioxidant enzyme mainly presents in cytoplasm and SOD1^G93A93A mutant is significantly used in ALS cell and mouse model.Mitochondrial degradation and structural damage are both of the earliest onsets of ALS.Mitochondria as a highly energized organelle,its dysfunction is likely to cause the lack of antioxidant capacity.Triggering tanslocation of BAX from mitochondrial tunnel and leading to the release of Cyt C and inducing cell apoptosis eventually.There was a direct bearing between Death receptor 6?DR6?and ALS,which has been reported before.DR6 was up-regulated both in the spinal cords of ALS patients and in ALS model mice using immunohistochemical analysis and western analysis.High expression of DR6 caused the death of motor neurons.Our previous research found that in the process of DR6-induced apoptosis,Pesenilin-1 associated protein?PSAP?also called mitochondrial apoptotic precursor molecule was required basically.In other words,the knockdown of PSAP can inhibit DR6-induced apoptosis.The purpose of our research was to figure out the role of PSAP in SOD1^G93A-induced apoptosis.Our study demonstrated that overexpression of SOD1^G93A93A in NSC-34 cell?mouse motor neuron?can decrease cellular activity?CCk-8 assay?,the result of Annexin V/PI staining assay and western blot showed that overexpression of SOD1^G93A93A could induce cell apoptosis.The knockdown of PSAP or BAX using siRNA interference can block the apoptosis induced by SOD1^G93A93A overexpression,at the same time,we found that inhibited PSAP/BAX expression can decrease the activation of Caspase-3 and increase cell activity.We proofed that PSAP and BAX made an important effect on theapoptosisofcelldeathinducedbyoverexpressionofSOD1^G93A.Immunoprecipitation?IP?experiments showed that SOD1/SOD1^G93A93A can form protein complex with BAX,and PSAP also had interaction with BAX during SOD1^G93A93A overexpression.How SOD1-BAX complex formed was confirmed by using mitochondrial isolation kit to compare the amount of BAX in cytosolic extracts and mitochondria-containing membrane.Furthermore,we proved that the formation of PSAP-BAX complex using BMH crosslinker,we found that Bax can form polymer with itself during the apoptosis induced by SOD1^G93A93A transfection.Overall,PSAP has an important influence on ALS associated with SOD1 mutant,the declination of PSAP expression can increase cell activity and alleviate intensity of cell apoptosis.PSAP can interact with other apoptosis proteins like BAX which is also a mitochondrial protein and this protein is also significant to ALS pathogenesis.This paper provides a theoretical basis for revealing the pathogenesis of ALS and it also supplies new ideas for drug development and clinical application about ALS.