The Point Mutation Of Nogo-C And Its Apoptosis Mechnism

Nogo (Neurite Outgrowth Inhibitor) was identified by Strittmatter SM and Schwab ME et al, who reported it in Nature simultaneously in 2000. Nogo is a member of Rticulums (RTN4/Nogo) and characterized by a～200-amino-acid C-terminal domain, including two hydrophobic sequence. In mammalian, Nogo mainly encodes three transcripts: Nogo-A,Nogo-B,Nogo-C. Nogo-A can inhibit neurite growth through Nogo receptor NgR. Recently, it has been reported that Nogo-B may be related with cancer cell apoptosis, and be a regulator of vascular remodeling. Besides neurite inbition, Nogo-C has been recently showed to interact with BACE (bindingβ-amloid converting enzyme 1) and regulate it reversely, also it can interacted with Nogo-A,Nogo-B in some cell surface for certain function. However, the relation between Nogo-C and Hepatocellular carcinoma has not ever been reported.Hepatocellular carcinoma (HCC) is one of tumours with high mortality rate. Some areas with high incidences of HCC in China include Qidong,Fusui,Haimen,Chongming, which provide much materias for HCC pathogeny study. The cancerogenic factors inducing HCC consist of hepatitis virus, chemical substance and abnormal genes et al. So far many genes related with HCC have been cloned, including Ras,c-myc,IGF-II,c-erbB-2,c-fos,Bcl-2,MDM2,C-ets2,P53,P73,Rb,p16,PTEN,P27,FHIT et al, which revealed that multi-gene activation is the molecular basis for HCC occurence. The invistigation about HCC in Qidong showed that hepatitis virus and aflatoxin B1 contamination iduced HCC with p53 gene mutation in the 249th codon of extron 7. However, HCC suffers a muti-gene, muti-step and complicated mechanism.Nogo-C may be related with Hepatocellular caicinona (HCC) by bioimformatics analysis firstly. Our study before has also indicated Nogo-C expressed abnormally in HCC, and then in this paper Nogo-C cellular function and its relation with HCC were furtherly studied. The main results include:1. Nogo-C expressed differently at transcriptional level in HCC and its relative normal tissues. Nogo-C was positively detected in 16/42 (38 % ) cases, and the cases of patients with tumor diameter more than 5 cm and Edomondson's III grade showed downregulated expression of Nogo-C.2. Three new point mutations of Nogo-C were found in HCC cases: A172G(Thr58Ala); A340G(Argll4Gly); A571G(Ile191Val). In 6 cases from Qidong, 5 cases showed Nogo-C mutation or incomplated mutation. The accesstion number of mutant Nogo-C submitted in NCBI is DQ778739.3. Our study showed mutant Nogo-C promoted HEK293 cell apotosis through JNK-c-Jun pathway. To detect endogeneous and exogenous Nogo-C expression, we prepared specific Nogo-C antiserum. By detection of signalling molecules and using JNK specific inhibitor SP600125, Nogo-C apoptotic mechanism were confirmed.4. Mutant Nogo-C was transfected with SMMC-7721, and induced cell growth inhibition and apoptosis. The ICC showed mutant p53 protein transferred from nucleus to cytoplasma and Hsp70 expreeion decreased and focused in cytoplasma. The mechanism still needs to be clarified.5. To further study the relation between Nogo-C and HCC, the wide and mutant Nogo/Ad were constructed.The above indicated Nogo-C missense mutation in HCC patients, and Nogo-C could inhibit SMMC-7721 cell growth and promote cell apoptosis through mutant p53 probably; also mutant Nogo-C could promote HEK293 cell apoptosis through JNK-c-Jun pathway.