1. The Potential Role Of Human-specific Acetylcholine Receptor Gene CHRFAM7A In Mental And Cognitive Functions. 2. The Protective Effect And Mechanism Of The Interaction Between Ecrg4 And Enolase In Myocardial Ischemia

Objective: ?7-acetylcholine receptor(?7-nAChR),a ligand-gated ion channel,is expressed in neurons,microglia and astrocytes of regions of the brain affecting cognition and higher order of cortical functions.It has been implicated in cognition,memory,and many neuropsychiatric disorders including schizophrenia,autism,Alzheimer's disease(AD),attention deficit hyperactivity disorder(ADHD),major depressive disorder,anxiety disorder,addiction,pain,and Parkinson disease(PD)among others.Therefore,?7-nAChR,highly conserved among species,has been considered an attractive potential therapeutic target for neuropsychiatric diseases.Many ?7-nAChR agonists and positive allosteric modulators(PAMs)have been successfully passed the pre-clinical animal studies and are advanced to clinical trials for the treatment of different neuropsychiatric disorders.Unfortunately,most lead compounds in clinical trials were not as effective as that observed in the preclinical animal models.This failure may,at least partially,be attributed to the emergence of a human-specific CHRFAM7 A gene during evolution.The uniquely human ?7-nAChR gene(CHRFAM7A)is evolved from the fusion of two partially duplicated genes,FAM7 and ?7-nAChR gene(CHRNA7),and is inserted on same chromosome 15,5'end of the CHRNA7 gene.Transcription of CHRFAM7 A gene produces a 1256-bp open reading frame encoding dup-?7-nAChR,where a 27-aminoacid residues from FAM7 replaced the146-aminoacid residues of the N-terminal extracellular ligand binding domain of ?7-nAChR.In in vitro experiment,dup-?7-nAChR has been shown to form hetero-pentamer with ?7-nAChR and dominant-negatively regulates the channel functions of ?7-nAChR.However,the contribution of CHRFAM7 A gene to the biology of ?7-nAChR in the brain in vivo remains largely a matter of conjecture.The purpose of this study was to study the regulation of?7-nAChR by CHRFAM7 A transgenic mice in vivo,so as to provide a new theoretical basis for the treatment of neuropsychiatric diseases related to?7-nAChR.Methods: CHRFAM7 A transgenic mouse was created and differentially expressed proteins were profiled from the whole brain using i TRAQ-2D-LC-MS/MS proteomic technology.Proteins with a fold change of?1.2 or ?0.83 and p < 0.05 were considered to be statistically significant.Results: Bioinformatics analysis showed that over-expression of the CHRFAM7 A gene significantly modulated the expression of proteins commonly involved in the signaling pathways of ?7-nAChR-mediated neuropsychiatric disorders including Parkinson's disease,Alzheimer's disease,Huntington's disease,and alcoholism.Conclusion: CHRFAM7 A gene contributes to the pathogenesis of neuropsychiatric disorders most likely through fine-tuning the functions of?7-nAChR in the brain.Objective: ECRG4 is a tumor suppressor that has been shown to protect cardiomyocytes from ischemia-reperfusion injury.However,the molecular mechanisms responsible for this protection remain uncertain.In order to understand the mechanism related to the myocardial protective effect of ECRG4 in ischemia,the purpose of this experiment was to identify ECRG4 interacting proteins and study their molecular mechanisms leading to ischemia pretection.Methods: Two plasmids p LVX-IRES-Zs Green1-2FLAG and p LVX-IRES-Zs Green1-ECRG4-2FLAG were constructed by molecular cloning technology;the plasmids were then transiently transfected into 293 T cells and were confirmed to express FLAG or ECRG4-FLAG respectively;lantiviruses were packaged and transduced into neonatal rat cardiomyocytes,which were confirmed by GFP-positive cells;transduced cardiomyocytes were then lysed,which was then processed sequentially by pulled-down using anti-FLAG-conjugated beads,SDS-PAGE,and silver-staining;the diffenentially pulled-down bands were then harvested,sequenced,and the potential interacting protein was identified.The potential interacting protein would be further confirmed by co-immunoprecipitation and immunofluorence methods,and the the molecular mechanisms leading to cardiac protection would be further explored.Results: 1.Two plasmids p LVX-IRES-Zs Green1-2FLAG and p LVX-IRES-Zs Green1-ECRG4-2FLAG were successfully constructed and could be successfully over-expressed in 293 T cells.2.p LVX-IRES-Zs Green1-ECRG4-2FLAG virus was successfully transduced into neonatal rat cardiomyocytes.3.Enolase was identified to interact with Ecrg4 in cardiomyocytes.Conclusion: Enolase may interact with Ecrg4 in cardiomyocytes resulting in Ecrg4-mediated protection of cardiac ischemia.