發(fā)布時(shí)間：2018-04-14 16:31

  本文選題：前列腺癌 + 核纖層蛋白A/C�。� 參考：《首都醫(yī)科大學(xué)學(xué)報(bào)》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(diǎn)突變與LMNA表達(dá)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(xì)胞(RWPE-1)、低侵襲力前列腺癌上皮細(xì)胞(PC-3M-2B4)及高侵襲力前列腺癌上皮細(xì)胞(PC-3M-1E8)LMNA基因的12個(gè)外顯子,識別出在RWPE-1與PC-3M-1E8細(xì)胞系中C.1159CCA(p.L387LI)點(diǎn)突變,定位在1號染色體156106006。為了驗(yàn)證點(diǎn)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(biāo)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(xì)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(shí),利用GEO數(shù)據(jù)庫分析了突變點(diǎn)周圍序列的特點(diǎn)與功能。最后,轉(zhuǎn)染突變質(zhì)粒,蛋白質(zhì)電泳分析LMNA點(diǎn)突變對LMNA蛋白質(zhì)表達(dá)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(cuò)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(cuò)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(xì)胞系和5種前列腺良性細(xì)胞系中,錯(cuò)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(diǎn),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(xì)胞死亡的負(fù)調(diào)節(jié)等方面。高表達(dá)突變LMNA的細(xì)胞系LMNA及磷酸化LMNA蛋白質(zhì)表達(dá)下調(diào)。結(jié)論 156106006位點(diǎn)突變多發(fā)生在前列腺正常組織和前列腺良性細(xì)胞株,與LMNA蛋白質(zhì)表達(dá)密切相關(guān),可能是lamin蛋白質(zhì)異源性表達(dá)的機(jī)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(xué)基礎(chǔ)醫(yī)學(xué)院生物化學(xué)與分子生物學(xué)系;首都醫(yī)科大學(xué)生物醫(yī)學(xué)工程學(xué)院生物醫(yī)學(xué)信息學(xué)系;
【基金】：國家自然科學(xué)基金(81272406,81672834) 北京市教育委員會科技計(jì)劃面上項(xiàng)目(KM201510025009)~~
【分類號】：R737.25

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