徐云峰刘松坚
(1.广州军区机关门诊部检验科,510080;2.广州军区疗养院检验科,510515)
细胞凋亡是一种程序性死亡,其形态学特征是质膜出泡、细胞皱缩、染色体浓缩、染色体DNA裂解[1]。有两种途径可导致细胞凋亡:①外源性凋亡途径,又被称为死亡受体通路,可由胞外肿瘤坏死因子受体(TNFR)或Fas受体(FasR)引发。②内源性凋亡途径,又被称为线粒体/细胞色素C介导的通路,可通过Bcl-2家族成员调控。Bcl-2家族是细胞凋亡信号转导途径中关键的凋亡调节因子,现就近年来Bcl-2家族的结构、家族成员之间的相互作用、信号通路的调控以及与肿瘤的关系等作一综述。
根据在细胞凋亡过程中发挥的生物学效应不同,Bcl-2家族可分为3类[2],第1类是抗凋亡蛋白,主要有Bcl-2、Bcl-xl、Bcl-w和Mcl-1等;第2类是促凋亡蛋白,主要包括Bax、Bak和Bok等;第3类是促凋亡蛋白中的特殊成员,主要有Bad、Bid、Bim、Bik、Puma和Noxa等。大部分Bcl-2家族蛋白主要由两大结构域构成,即位于羧基末端的跨膜结构域(TM)和不同数量的Bcl-2同源结构域(BH)。抗凋亡蛋白和促凋亡蛋白的结构中均含有BH1-4结构域[3],促凋亡蛋白中的特殊成员仅含有BH3结构域,又被称为BH3-only成员[4]。
Bcl-2家族是细胞凋亡信号转导途径中关键的凋亡调节因子,它们共同参与一个非常复杂的相互作用机制以调控细胞凋亡。
Bcl-2家族在线粒体介导的细胞凋亡中发挥重要作用。Bcl-2家族的抗凋亡蛋白Bcl-2、Bcl-xl主要分布于线粒体膜内外侧,其中Bcl-2还存在于核膜以及内质网膜上,通常在线粒体外膜发挥抗凋亡作用,以维持膜的完整性。Bcl-2家族的促凋亡蛋白Bax一般出现在胞浆中,当细胞响应损伤或刺激等凋亡信号后,Bax将重新定位于线粒体表面,通过破坏线粒体膜的完整性发挥作用[5]。
BH3-only蛋白是细胞应对外界凋亡信号的最主要方式,当BH3-only成员包括Bad、Bid、Bim、Bik和Puma等接收到凋亡信号后,BH3-only蛋白的表达增多,且发生翻译后修饰,通过两种机制发挥促凋亡作用[6-8]:一种是与Bcl-2家族中的抗凋亡蛋白形成对抗[9-11],一种是激活促凋亡蛋白Bax和Bak[12-15]。
不仅Bcl-2家族成员的表达可调控细胞凋亡,Bcl-2家族成员的翻译后修饰也可调节细胞凋亡。Bcl-2家族成员受到许多翻译后修饰,尤其是磷酸化和泛素化,但是修饰后的生物学功能受到争议[16]。例如:抗凋亡蛋白在丝氨酸或苏氨酸残基处发生磷酸化后,可上调也可下调其活性和稳定性,也可诱导细胞周期停滞以及改变细胞内定位等[16-17]。近期研究发现,纺锤体抑制剂可诱导Bcl-2在Ser70的磷酸化,促进Bcl-2与Bak、Bim的结合,从而提高细胞的化疗耐受力[18]。
Mcl-1蛋白在转录、翻译以及蛋白质转换水平均受到调控。非泛素依赖途径可降解Mcl-1,而去泛素化酶可提高Mcl-1的稳定性[19-20]。最新研究还发现,Mcl-1的N端也可影响其稳定性[19]。
BH3-only蛋白可通过不同途径进行调控。当细胞内DNA发生损伤,P53可被ATM和ATR信号通路激活,进而作为转录因子参与调控Puma的转录;Puma蛋白在多个位点如Ser10处发生磷酸化可诱导其蛋白发生蛋白酶体降解,从而下调其表达[21];另外研究还发现,胞浆中非磷酸化的Bad可与膜上的Bcl-2或Bcl-xl形成杂二聚体诱导细胞凋亡,当被14-3-3蛋白磷酸化后,Bad则被封闭在胞浆中[22]。
肿瘤的发生通常与Bcl-2家族成员的表达异常有关。研究发现,过表达抗凋亡蛋白Bcl-2与包括淋巴瘤、慢性淋巴细胞白血病等多种恶性肿瘤的发生发展有关[23-24];敲除促凋亡基因如Bim[25]、Puma[26-27]、Bad[28]和Bax[29]等可诱导多种肿瘤的发生。
在90%的人的滤泡性B细胞淋巴瘤中,t(14;18)染色体发生易位,引起Bcl-2基因过表达从而抑制淋巴细胞的凋亡[30];在多种肿瘤细胞中,Mcl-1和Bcl-xl同样过表达[31];另外,在多种恶性肿瘤中,Bim和Puma的启动子发生高度甲基化,其蛋白表达降低,从而抑制肿瘤细胞的凋亡[26,32]。
近期研究还发现,Bcl-2家族成员的表达与肿瘤细胞对化疗药物的耐受性密切相关。例如,Bcl-2基因的过表达可使肺小细胞癌和白血病/淋巴瘤细胞对化疗药阿糖胞苷产生耐药性[33];Puma、Noxa和Bim的过表达可降低淋巴瘤细胞对DNA损伤-诱导药物的耐药性[34];Bim的过表达可诱导糖皮质激素[35]、紫杉醇[36]杀死肿瘤细胞。因此,提高BH3-only蛋白的表达可能有助于肿瘤的治疗。
近年来,有关Bcl-2家族调控肿瘤细胞凋亡的机制研究越来越多,但是有关在凋亡过程中Bcl-2家族成员之间相互作用机制、膜结构如何调节各成员之间的相互作用等还有待进一步研究。
大量有关细胞凋亡的分子机制研究表明Bcl-2家族可用于肿瘤的靶向治疗。因此,我们可以设计一些高效且特异性好的抗肿瘤药物,通过抑制抗凋亡蛋白的表达或激活促凋亡蛋白的表达以治疗恶性肿瘤。
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