本文選題：前列腺癌 + 核纖層蛋白A/C�。� 參考：《首都醫(yī)科大學學報》2017年06期

【摘要】：目的探討前列腺癌中核纖層蛋白A/C(lamin A/C,LMNA)基因第七外顯子點突變與LMNA表達差異的相關(guān)性。方法采用高通測序分析了永生化正常前列腺上皮細胞(RWPE-1)、低侵襲力前列腺癌上皮細胞(PC-3M-2B4)及高侵襲力前列腺癌上皮細胞(PC-3M-1E8)LMNA基因的12個外顯子,識別出在RWPE-1與PC-3M-1E8細胞系中C.1159CCA(p.L387LI)點突變,定位在1號染色體156106006。為了驗證點突變在前列腺癌患者中的分布情況,Pub Med GEO數(shù)據(jù)庫下載了3組數(shù)據(jù),分別是100例前列腺癌患者石蠟包埋標本的總RNA測序數(shù)據(jù)、20例前列腺癌和10例配對正常組織的轉(zhuǎn)錄組數(shù)據(jù)及21種前列腺正常及癌細胞株的測序數(shù)據(jù)。在1號染色體上選取156105950到156106055范圍,設(shè)置比對序列CTACGCCTG或者NTACGCCTGTCCCCCAGCCC,每次分析比對長度為50 nt。同時,利用GEO數(shù)據(jù)庫分析了突變點周圍序列的特點與功能。最后,轉(zhuǎn)染突變質(zhì)粒,蛋白質(zhì)電泳分析LMNA點突變對LMNA蛋白質(zhì)表達的影響。結(jié)果在所有分析的樣品中,1號染色體LMNA外顯子7從156106006到156106011的區(qū)域內(nèi),存在2種錯義突變形式:C/CA(p.L387LI)、C/CG(p.L387LV)和4種同義突變形式:C/CT(p.L387L)、C/CA(p.R388R)、C/CT(p.R388R)、C/CG(p.R388R)。前列腺癌按格里森(Gleason score,GS)評分系統(tǒng)分組,錯義突變的總發(fā)生率分別占40%(正常)、11%(GS 5-6)、2%(GS 7)和6%(GS 8-10)。在16種前列腺癌細胞系和5種前列腺良性細胞系中,錯義突變的總發(fā)生率分別占31%和60%。此外,還發(fā)現(xiàn)1號染色體上從156106008到156106066是轉(zhuǎn)錄因子結(jié)合位點,常見轉(zhuǎn)錄因子為PAX5、HEN1(NHLH1)、HTF、P53、MIF1、COMP1和NGFIC(NGF4)。通過功能聚類分析,這些轉(zhuǎn)錄因子的功能主要集中在染色質(zhì)重組及調(diào)控、RNA代謝過程的正向調(diào)節(jié)、組蛋白修飾的調(diào)控以及程序性細胞死亡的負調(diào)節(jié)等方面。高表達突變LMNA的細胞系LMNA及磷酸化LMNA蛋白質(zhì)表達下調(diào)。結(jié)論 156106006位點突變多發(fā)生在前列腺正常組織和前列腺良性細胞株,與LMNA蛋白質(zhì)表達密切相關(guān),可能是lamin蛋白質(zhì)異源性表達的機制之一。
[Abstract]:Objective to investigate the relationship between the point mutation of the seventh exon of nuclear laminin A/C(lamin A / C rmna gene and the difference of LMNA expression in prostate cancer.Methods the 12 exons of RWPE-1, PC-3M-2B4 and PC-3M-1E8 LMNA gene in immortalized normal prostatic epithelial cells and low invasive prostate cancer epithelial cells were analyzed by high pass sequencing. The mutation of L387LI gene was identified in RWPE-1 and PC-3M-1E8 cell lines.Located on chromosome 1 156106006.To verify the distribution of point mutations in prostate cancer patients and download three sets of data from the Pub Med GEO database,The total RNA sequencing data were obtained from paraffin-embedded paraffin embedded specimens of 100 prostate cancer patients respectively. The transcriptome data of 20 cases of prostate cancer and 10 cases of matched normal tissues and 21 kinds of prostate normal and cancer cell lines were sequenced.From 156105950 to 156106055 on chromosome 1, the alignment sequence CTACGCCTG or NTACGCCTGTCCCCCAGCCC was set, and the length of each analysis was 50 NT.At the same time, the characteristics and functions of the sequence around the mutation point are analyzed by using GEO database.Finally, the effect of point mutation of LMNA on the expression of LMNA protein was analyzed by protein electrophoresis.Results in all the samples analyzed, there were two missense mutations in the region of exon 7 of chromosome 1 from 156106006 to 156106011. There were two missense mutations: C / CAp.L387LI / L / L ~ (387L) and four synonyms: C / C / C / C / L ~ (387L) / C / C / C / C / R ~ (388R).According to the Gleason scoreGSscoring system, the total incidence of missense mutation in prostate cancer was 40% (normal 11 GS 5-6% GS7) and 6%(GS 8-10%.In 16 prostate cancer cell lines and 5 benign prostate cell lines, the total incidence of missense mutation was 31% and 60%, respectively.It was also found that the transcription factor binding sites on chromosome 1 were from 156106008 to 156106066, and the common transcription factors were PAX5, HEN1, NHLHH1, HTF5, P53, MIF1, COMP1 and NGFICF4, respectively.By functional cluster analysis, these transcription factors mainly focus on the positive regulation of chromatin recombination and regulation of RNA metabolism, the regulation of histone modification and the negative regulation of programmed cell death.The expression of LMNA and phosphorylated LMNA protein was down-regulated in the cell line with high expression of mutant LMNA.Conclusion the 156106006 mutation occurs in normal prostate tissues and benign prostate cell lines, which is closely related to the expression of LMNA protein, which may be one of the mechanisms of heterogenous expression of lamin protein.
【作者單位】： 首都醫(yī)科大學基礎(chǔ)醫(yī)學院生物化學與分子生物學系;首都醫(yī)科大學生物醫(yī)學工程學院生物醫(yī)學信息學系;
【基金】：國家自然科學基金(81272406,81672834) 北京市教育委員會科技計劃面上項目(KM201510025009)~~
【分類號】：R737.25

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