目的采用蛋白质组学方法,对长滩军团菌血清1型临床分离株(LL-1)与环境分离株(CD-1)全菌蛋白进行比较和分析,探索军团菌临床株与环境分离株是否存在蛋白表达差异,为进一步认识、研究军团菌提供依据。方法培养、收集CD-1与LL-1菌株,用超声兼尿素-3-[(3-胆酰胺丙基)-乙二胺]-1丙磺酸(CHAPS)-二硫苏糖醇(DTT)裂解法提取全菌蛋白,SDS-PAGE及Western blot对二者全菌蛋白进行初步比较,确定二者全菌蛋白表达存在差异后,采用双向电泳(2-DE)联合MALDI-TOF-TOF质谱深入研究,并对质谱鉴定的差异点进行Western blot验证。结果初步SDS-PAGE及Western blot试验表明,CD-1与LL-1存在蛋白表达差异。进一步的2-DE研究结果共获得142个蛋白。CD-1与LL-1相比,18个蛋白点表达上调、19个蛋白表达下调。对其中显著差异的四个点进行质谱分析、Mascot软件查询、蛋白数据库搜库后证实CD-1表达下调的为热休克蛋白70(Chaperone protein Dna K)和30 s核糖体蛋白S1(30S ribosomal protein S1);CD-1表达上调的蛋白为假想蛋白TP04_0359(hypothetical protein TP04_0359)和一个未知蛋白。对进行质谱分析的四个蛋白点进行Western blot验证确实是军团菌蛋白。结论 LL-1与CD-1蛋白表达存在差异,此试验为进一步研究军团菌毒力和致病及疫苗研制奠定了基础。
The aim of the present study was to investigate how the miR-451 a can regulate resistance to tamoxifen(TAM) of breast cancer cells by modulating the expression of estrogen receptor α(ERa) and 14-3-3ζ.The effects of ...
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The aim of the present study was to investigate how the miR-451 a can regulate resistance to tamoxifen(TAM) of breast cancer cells by modulating the expression of estrogen receptor α(ERa) and 14-3-3ζ.The effects of miR-451 a and R18,an inhibitory peptide of 14-3-3 protein,on growth and apoptosis of MCF-7 and LCC2 cells were detected by MTT assay,AnnexinV-FITC binding assay,and Hochest 33258 *** expressions of ERa,14-3-3ζ,and miR-451 a were measured by qRT-PCR and Western blot *** interaction of 14-3-3ζ and ERa was investigated by *** was shown that over-expression of miR-451 a can enhance the TAM sensitivity in MCF-7 and LCC2 *** effects were elicitedby knocking down miR-451 a in these *** results also showed that the TAM treatment could up-regulate 14-3-3ζ,and down-regulate ERa expression in a time-dependent *** mRNA expression of 14-3-3ζ and miR-451 a were inversely correlated,while the mRNA expression of ERa and miR-451 a were positively ***,the 14-3-3ζ protein and ERa protein can ***-expression of miR-451 a can decrease 14-3-3ζ expression,and increase ERa expression,which consequently suppressed cell proliferation,increased apoptosis,and reduced activation of p-AKT and p-mTOR.R18 can significantly decrease cell prolif-eration,and increase apoptosis.R18 and si 14-3-3ζ can inhibit the effects of down-regulation of ERa after knocking down of miR-451 *** conclusion,overexpression of miR-451 a can enhance the sensitivity of breast cancer cells to TAM by down-regulating 14-3-3ζ and up-regulating ERa,which suggested it to be a potential target for restoring ERa expression and reversing resistance to antiestrogen therapy.
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