Expression Profile And Tissue-Specific Distribution Of COMMD10 In BALB/c Mice And Human And Bioinformatics Analysis Of COMMD10

Background and PurposeNowadays,cancer has posed a great threat on human health and social development.In recent years,the incidence of cancer shows an increasing trend worldwide.With a rather high cancer incidence,the number of newly diagnosed cancer cases every year in China has reached 21.8%of total cancer cases worldwide.Obviously,the prevention and treatment of cancer has become the public concern.The development of cancer is complicated and made up of multiple procedures.Every step is modulated by various genes and proteins.Changes including mutation,depletion,overexpression and epigenetic regulation of key molecules will make a big difference.In order to have a better understanding of mechanisms underlying cancer and to precisely prevent cancer development,a comprehensive knowledge of the cancer-related genes is necessary.Recently,a protein family named COMMD was discovered characteristic of a COMM domain located in carboxy-terminal.The family contains 10 members and they are demonstrated to be involved in many biological processes,including the regulation of NF-?B(nuclear factor-KB)pathway.COMMD 10 has been proved to target p65 NF-?B subunit and reduce its nuclear translocation,thereby leading to the inactivation of NF-?B pathway and suppression of CRC invasion and metastasis.However,the distribution and expression of COMMD 10 in human normal tissues as well as tumor tissues are not clear.This study will start from the distribution and expression of COMMD10 in BALB/c Mice and human tissues.Furthermore,the biological function of COMMD 10 in human tumors and the involved mechanisms will be explored using bioinformatics methods.Methods1.Expression and distribution of COMMD10 in normal tissues of BALB/c mice1.1 Ten kinds of fresh frozen normal tissues including heart,lung,brain,stomach,small intestine,spleen,liver,kidney,pancreas,and thymus were harvested from BALB/c mice.And expression of COMMD10 mRNA in different tissues of BALB/c mice was detected by qPCR.1.2 Twelve kinds of normal tissues including heart,lung,brain,trachea,esophagus,stomach,small intestine,spleen,liver,kidney,pancreas and thymus were harvested from BALB/c mice and made into paraffin sections.The expression of COMMD 10 protein in different tissues of BALB/c mice was detected by immunohistochemical staining.2.The expression and distribution of COMMD10 in human normal tissues and common tumor tissues2.1 Sixteen kinds of human normal tissues including lung,esophagus,stomach,liver,kidney,colon,rectum,breast,placenta,prostate,skin,thyroid,bladder,cervix,spleen and uterus were collected.And twenty-two kinds of tumor tissues including hepatocellular carcinoma,cholangiocellular carcinoma,colon adenocarcinoma,rectal adenocarcinoma,breast invasive ductal carcinoma,renal urothelial carcinoma,renal clear cell carcinoma,renal chromophobe,uterine fibroids,endometrial cancer,gastric cancer,esophageal squamous cell carcinoma,thyroid papillary carcinoma,prostate cancer,nasopharyngeal carcinoma,gastrointestinal stromal tumor,breast small cell carcinoma,breast phyllode tumor,lung adenocarcinoma,thymoma,cervical cancer and bladder urothelial carcinoma were collected.The expression of COMMD10 protein in these human tissues was detected by immunohistochemical staining.2.2 The expression of COMMD10 in different tumor tissues and matched normal tissues was analyzed using TCGA data.The correlations between expression of COMMD10 and patients prognosis were also analyzed using TCGA data.3.Bioinformatics analysis of COMMDIO3.1 The basic information of COMMDIO was analyzed using UCSC database(http://genome.ucsc.edu/)and the COMMD10 interacting proteins were analyzed using the String database(https://string-db.org/).3.2 The COMMDIO and its interacting proteins were analyzed from the following aspects,biological processes,cell composition,molecular function,and KEGG pathways using DAVID database(https://david.abcc.ncifcrf.gov/).3.3 Databases including miRDB(http://www.mirdb.org/),miRWalk(http://zmf.umm.uni-heidelberg.de/),Target Scan(http://www.targetscan.org/)and microRNA(http://34.236.212.39/microRNA/home.do)were used to predict miRNAs that might regulate COMMDIO expression and the results were intersected to obtain the target miRNA.Results1.Expression and distribution of COMMD10 in normal tissues of BALB/c mice1.1 In ten kinds of BALB/c mice tissues including heart,lung,brain,stomach,small intestine,liver,spleen,kidney,pancreas and thymus,COMMD10 mRNA has different expression levels.Among them,the expression of COMMD10 in lung and spleen tissues was significantly higher than that in other tissues.The expression of COMMD10 in heart and brain tissues was significantly lower than that in other tissues.1.2 COMMD10 protein showed no expression in the thymus tissue of BALB/c mice.COMMDIO protein expression differed in the other eleven kinds of tissues such as heart,lung,brain,trachea,esophagus,stomach,small intestine,spleen,liver,kidney and pancreas.And COMMD10 protein was mainly located in the cytoplasm.The common expression of COMMD10 protein was found in the vascular endothelial cells and smooth muscle cells of different tissues.2.The expression and distribution of COMMD10 in human normal tissues and tumor tissues2.1 COMMD10 protein showed no expression in skin,thyroid,bladder,cervix and spleen.COMMD10 protein had different expression in lung,esophagus,stomach,liver,kidney,colon,rectum,breast,placenta,prostate and uterus.Among them,high expression of COMMD10 was observed in stomach,liver and kidney tissues.The COMMDIO protein was mainly located in the cytoplasm,but expressed in both cytoplasm and nucleus in hepatocytes.COMMD10 protein showed no expression in six types of tumor tissues including breast small cell carcinoma,breast phyllode tumor,lung adenocarcinoma,thymoma,cervical cancer and bladder urothelial carcinoma.High expression of COMMDIO was observed in nasopharyngeal carcinoma and gastrointestinal stromal tumor.Low expression of COMMDIO was observed in hepatocellular carcinoma,cholangiocellular carcinoma,colon adenocarcinoma,rectal adenocarcinoma,breast invasive ductal carcinoma,renal urothelial carcinoma,renal clear cell carcinoma,renal chromophobe,prostate cancer,uterine fibroids,endometrial cancer,gastric cancer,esophageal squamous cell carcinoma and thyroid papillary carcinoma.The COMMD10 protein was localized in the cytoplasm of tumor cells.2.2 The expression of COMMDIO in invasive breast cancer tissues was up-regulated,while the expression of COMMD10 was down-regulated in lung squamous cell carcinoma and renal clear cell carcinoma.In renal clear cell carcinoma,the overall survival of patients with high expression of COMMD10 was significantly higher than that of patients with low expression of COMMD10.The low expression of COMMD10 predicts poor prognosis of patients with renal clear cell carcinoma.3.Bioinformatics analysis of COMMD103.1 STRING database was used to do the protein interaction network analysis.The results showed that 41 genes are likely to interact with COMMD10.These genes are RBX1,ATP7B,SLC31A2,C14orfl66,TCEB2,COMMD9,CP,COMMD4,COMMD10,COMMD7,ATP11A,ZER1,ARAP2,COMMD5,MORN4,COMMD1,DSCR3,ATOX1,ASB4,PSMG1,PNRC2,WARS,COMMD6,NPEPL1,CCDC151,MCMBP,FAM45A,SLC31A1,TFDP1,IARS,CCDC22,CCDC93,COMMD3,COMMD8,VPS29,ENSG00000215700,RELA,C16orf62,SH3GLB1,COMMD2 and CUL2.3.2 COMMD10 and its interaction genes were clustered using the DAVID database(https://david.abcc.ncifcrf.gov/).The results showed that COMMD10 and its interaction genes are mainly involved in the formation of cellular components such as Cullin-RING ubiquitin ligase complexes,endosomes and so on.The clustering of molecular functions mainly points to the binding and transport of copper ions,binding to ubiquitinated protein ligase and regulating NF-?B pathway.The related biological processes mainly include the transport and steady-state maintenance of copper ions,transcription,translation,and transport of proteins,and negative regulation of NF-?B transcription factors.COMMD10 and its interaction genes are analyzed in the KEGG database.It has been found to be related to HIF-1 signaling pathway,ubiquitination-mediated proteolysis and pathways in renal cell carcinoma.3.3 Hsa-miR-590-3p was predicted simultaneously by four miRNA prediction database to regulate the expression of COMMD10.Conclusion1.COMMD10 mRNA was differently expressed in normal tissues of BALB/c mice,with high expression in spleen and lung tissues and lowest expression in cardiac tissues.2.COMMD10 protein showed no expression in the thymus tissue of BALB/c mice and was expressed in other important tissues.It was mainly located in the cytoplasm.3.COMMD10 protein was highly expressed in human normal stomach,liver and kidney tissues,and is commonly expressed in vascular endothelium and smooth muscle tissue.4.COMMD10 predicts a good prognosis of renal clear cell carcinoma patients.Innovation points of this paper1.The expression and distribution of COMMDIO in different tissues of BALB/c mice were elucidated from mRNA and protein levels for the first time.2.It is the first time that COMMDIO has been suggested as a tumor suppressor in renal clear cell carcinoma.3.The possible biological function and underlying mechanism of COMMD10 in renal clear cell carcinoma was predicted using bioinformatics methods.