细胞色素c能诱导植物细胞编程性死亡

以悬浮培养的胡萝卜（ＤａｕｃｕｓｃａｒｏｔａＬ．）与烟草（ＮｉｃｏｔｉａｎａｔａｂａｃｕｍＬ．ｃｖ．ＢＹ２）细胞原生质体为材料,加入一定浓度的细胞色素ｃ和ｄＡＴＰ。不同取样时间的ＤＡＰＩ荧光染色与电镜超薄切片观察的结果显示染色质发生凝集、趋边化,最终形成凋亡小体。核酸电泳显示ＤＮＡ发生特异降解并形成电泳“阶梯”（ＤＮＡｌａｄｄｅｒ）。用末端脱氧核糖核酸转移酶介导的ｄＵＴＰ切口末端标记方法（ＴＵＮＥＬ）检测发现ＤＮＡ的３＇ＯＨ断端被原位特异标记。以上结果说明：细胞色素ｃ能诱导植物细胞发生典型的凋亡。     

B-Raf Inhibits Programmed Cell Death Downstream of Cytochrome c Release from Mitochondria by Activating the MEK/Erk Pathway

Growth factor-dependent kinases, such as phosphatidylinositol 3-kinase (PI 3-kinase) and Raf kinases, have been implicated in the suppression of apoptosis. We have recently established Rat-1 fibroblast cell lines overexpressing B-Raf, leading to activation of the MEK/Erk mitogen-activated protein kinase pathway. Overexpression of B-Raf confers resistance to apoptosis induced by growth factor withdrawal or PI 3-kinase inhibition. This is accompanied by constitutive activation of Erk without effects on the PI 3-kinase/Akt pathway. The activity of MEK is essential for cell survival mediated by B-Raf overexpression, since either treatment with the specific MEK inhibitor PD98059 or expression of a dominant inhibitory MEK mutant blocks the antiapoptotic activity of B-Raf. Activation of MEK is not only necessary but also sufficient for cell survival because overexpression of constitutively activated MEK, Ras, or Raf-1, like B-Raf, prevents apoptosis after growth factor deprivation. Overexpression of B-Raf did not interfere with the release of cytochrome c from mitochondria after growth factor deprivation. However, the addition of cytochrome c to cytosols of cells overexpressing B-Raf failed to induce caspase activation. It thus appears that the B-Raf/MEK/Erk pathway confers protection against apoptosis at the level of cytosolic caspase activation, downstream of the release of cytochrome c from mitochondria.     

死亡信号传递: 叶绿体与线粒体间信号交流调控植物程序性细胞死亡

程序性细胞死亡(PCD)是生物体受遗传调控的自主细胞死亡现象, 在植物生长发育和抵抗环境胁迫中起重要作用。PCD的发生可受线粒体中活性氧(ROS)诱导。中国科学院遗传与发育生物学研究所李家洋研究组早期的研究发现了1个拟南芥(Arabidopsis thaliana)细胞死亡突变体mod1, 并暗示植物细胞中存在叶绿体与线粒体之间的信号交流调控PCD, 但其中的具体作用机制尚不清楚。最近, 他们通过大规模筛选mod1突变体的抑制突变体, 克隆了3个新的抑制基因plNAD- MDH、DiT1和mMDH1。此3个基因分别编码质体定位的NAD依赖的苹果酸脱氢酶、叶绿体被膜定位的二羧酸转运蛋白1和线粒体定位的苹果酸脱氢酶1, 突变后都可抑制mod1中ROS的积累及PCD的发生。通过对这些基因进行深入的功能分析, 他们论证了苹果酸从叶绿体到线粒体的转运对线粒体中ROS的产生及随后PCD的诱导起重要作用。该研究拓展了我们对植物细胞中细胞器间交流的认识, 为我们深入理解植物PCD发生机制提供了新线索, 是该领域的一项突破性进展。     

New Arabidopsis thaliana Cytochrome c Partners: A Look Into the Elusive Role of Cytochrome c in Programmed Cell Death in Plants

Programmed cell death is an event displayed by many different organisms along the evolutionary scale. In plants, programmed cell death is necessary for development and the hypersensitive response to stress or pathogenic infection. A common feature in programmed cell death across organisms is the translocation of cytochrome c from mitochondria to the cytosol. To better understand the role of cytochrome c in the onset of programmed cell death in plants, a proteomic approach was developed based on affinity chromatography and using Arabidopsis thaliana cytochrome c as bait. Using this approach, ten putative new cytochrome c partners were identified. Of these putative partners and as indicated by bimolecular fluorescence complementation, nine of them bind the heme protein in plant protoplasts and human cells as a heterologous system. The in vitro interaction between cytochrome c and such soluble cytochrome c-targets was further corroborated using surface plasmon resonance. Taken together, the results obtained in the study indicate that Arabidopsis thaliana cytochrome c interacts with several distinct proteins involved in protein folding, translational regulation, cell death, oxidative stress, DNA damage, energetic metabolism, and mRNA metabolism. Interestingly, some of these novel Arabidopsis thaliana cytochrome c-targets are closely related to those for Homo sapiens cytochrome c (Martínez-Fábregas et al., unpublished). These results indicate that the evolutionarily well-conserved cytosolic cytochrome c, appearing in organisms from plants to mammals, interacts with a wide range of targets on programmed cell death. The data have been deposited to the ProteomeXchange with identifier PXD000280.Programmed cell death (PCD)¹ is a fundamental event for the development of multicellular organisms and the homeostasis of their tissues. It is an evolutionarily conserved mechanism present in organisms ranging from yeast to mammals (1–3).In mammals, cytochrome c (Cc) and dATP bind to apoptosis protease-activating factor-1 (Apaf-1) in the cytoplasm, a process leading to the formation of the Apaf-1/caspase-9 complex known as apoptosome. This apoptosome subsequently activates caspases-3 and -7 (4, 5). In other organisms, such as Caenorhabditis elegans or Drosophila melanogaster, however, Cc is not essential for the assembly and activation of the apoptosome (6) despite the presence of proteins homologous to Apaf-1—cell death abnormality-4 (CED-4) in C. elegans and Drosophila Apaf-1-related killer (Dark) in D. melanogaster—which have been found to be essential for caspase cascade activation. Furthermore, other organisms such as Arabidopsis thaliana lack Apaf-1 (7). In fact, only highly distant caspase homologues (metacaspases) (8, 9), serine proteases (saspases) (10), phytaspases (11) and VEIDases (12–14) with caspase-like activity have been detected in plants; however, their targets remain veiled and whether they are activated by Cc remains unclear.Intriguingly, the release of Cc from mitochondria into the cytoplasm during the onset of PCD is an evolutionarily conserved event found in organisms ranging from yeast (15) and plants (16) to flies (17), and mammals (18). However, understanding of the roles of this phenomenon in different species can be said to be uneven at best. In fact, the release of Cc from mitochondria has thus far been considered a random event in all organisms, save mammals. Thus, the participation of Cc in the onset and progression of PCD needs to be further elucidated.Even in the case of mammals, the role(s) of Cc in the cytoplasm during PCD remain(s) controversial. Recently, new putative functions of Cc, going beyond the already-established apoptosome assembly process, have been proposed in the nucleus (19, 20) and the endoplasmic reticulum (21–23). Neither these newly proposed functions nor other arising functions, such as oxidative stress (24), are as yet fully understood. This current state of affairs demands deeper exploration of the additional roles played by Cc in nonmammalian species.In this study, putative novel Cc-partners involved in plant PCD were identified. For this identification, a proteomic approach was employed based on affinity chromatography and using Cc as bait. The Cc-interacting proteins were identified using nano-liquid chromatography tandem mass spectrometry (NanoLC-MS/MS). These Cc-partners were then further confirmed in vivo through bimolecular fluorescence complementation (BiFC) in A. thaliana protoplasts and human HEK293T cells, as a heterologous system. Finally, the Cc-GLY2, Cc-NRP1 and Cc-TCL interactions were corroborated in vitro using surface plasmon resonance (SPR).These results indicate that Cc is able to interact with targets in the plant cell cytoplasm during PCD. Moreover, they provide new ways of understanding why Cc release is an evolutionarily well-conserved event, and allow us to propose Cc as a signaling messenger, which somehow controls different essential events during PCD.     

植物细胞的编程性死亡

植物细胞受基因调控死亡,这种编程性定位的细胞死亡具有积极的生理功能,形成有利于自身发展的结构。基因调控细胞内的酶活动,细胞器产生变化,导致细胞死亡。外部因素可调节植物基因及基因调控产物的表达。     

胱冬肽酶非依赖性的细胞程序性死亡

在多细胞有机体的组织内稳态维持和正常发育过程中,细胞程序性死亡发挥着重要的作用。细胞程序性死亡有多种形式(如细胞凋亡、类细胞凋亡和类坏死等),其中了解较清楚的是细胞凋亡。一直以来,胱冬肽酶(caspase)被认为是细胞凋亡发生中关键的一种蛋白酶。但是最近的研究表明,包括细胞凋亡在内的一些细胞程序性死亡可以以一种不依赖胱冬肽酶的方式发生。细胞程序性死亡与胱冬肽酶之间存在非依赖性关系。     

Akt/Protein Kinase B Inhibits Cell Death by Preventing the Release of Cytochrome c from Mitochondria

Growth factors signaling through the phosphoinositide 3-kinase/Akt pathway promote cell survival. The mechanism by which the serine/threonine kinase Akt prevents cell death remains unclear. We have previously shown that Akt inhibits the activity of DEVD-targeted caspases without changing the steady-state levels of Bcl-2 and Bcl-x(L). Here we show that Akt inhibits apoptosis and the processing of procaspases to their active forms by delaying mitochondrial changes in a caspase-independent manner. Akt activation is sufficient to inhibit the release of cytochrome c from mitochondria and the alterations in the inner mitochondrial membrane potential. However, Akt cannot inhibit apoptosis induced by microinjection of cytochrome c. We also demonstrated that Akt inhibits apoptosis and cytochrome c release induced by several proapoptotic Bcl-2 family members. Taken together, our results show that Akt promotes cell survival by intervening in the apoptosis cascade before cytochrome c release and caspase activation via a mechanism that is distinct from Bad phosphorylation.     

细菌的细胞程序性死亡

细胞凋亡 (ａｐｏｐｔｏｓｉｓ)也称为细胞程序性死亡 (ｐｒｏｇｒａｍｍｅｄｃｅｌｌｄｅａｔｈ ,ＰＣＤ) ,是由细胞自身的程序性自杀机制激活的细胞死亡现象。在多细胞真核生物中 ,程序性的细胞死亡是由细胞表面的死亡受体介导 ,通过一系列的半胱氨酸蛋白酶 (ｃａｓｐａｓｅｓ)作用而启动[1] ,维系个体结构稳定、功能平衡和生长发育所必需的基本生物学过程。一般存在于个体的发育过程中 ,导致细胞功能和形态学上的改变 ,如蛋白质的水解、ＤＮＡ和ＲＮＡ的降解、细胞的收缩 ,以及细胞碎裂形成凋亡小体 (ａｐｏｐｔｏｔｉｃｂｏｄｉｅｓ)等 …     

细胞程序性死亡与帕金森病的相关研究进展

帕金森病发病机制至今未明,近几年研究发现,线粒体依赖性PCD通路的激活在PD发病过程中是不可缺少的,不同形态学表现的细胞死亡形式在帕金森病发病过程中可以共同存在,而所有的这些细胞死亡都归因于PCD共同的上游通路的激活。PCD通路不仅仅是指线粒体介导的caspase依赖性凋亡,还包括非caspase依赖性细胞非凋亡性死亡,比如细胞坏死。这不仅仅是概念上的延伸,更为我们在帕金森病神经保护性治疗上提供了更多的靶点,有助于寻求神经保护的新方法和延缓神经退行性疾病的进程.抗凋亡治疗已经成为帕金森病等神经退行性疾病治疗的新热点,已经证实,caspase抑制剂能够通过抑制caspase的激活,阻止细胞退行性病变。那么将位于caspase执行者上游的Bax作为靶点,抑制Bax的激活与转位,能够产生更为持久显著的神经保护作用。本文综述了近年来相关研究进展。     

植物中的细胞程序性死亡

细胞程序性死亡(PCD)对于维持植物的正常生长发育非常重要,目前已成为植物学研究的一个热点。本文综合评述了近年来植物PCD研究的某些进展,包括植物PCD的特征,植物的营养生长、生殖生长以及与环境互作过程中存在的各种PCD及其证据,植物PCD发生的分子机制及其调控等等。对植物PCD研究中有待进一步解决的问题和可能意义提出了自己的见解。     

细胞死亡的基因调节

GeneRegulationofProgrammedCellDeathYangJianminLiuLianrui(InstituteofGenetics,AcademiaSinica,Beijing100101)细胞死亡与细胞生长、分化和增殖一样是维持生物体平衡的重要环节。它是生命过程一个重要组成部分,只在近年来才引起生物学家和医学家的重视．细胞死亡现象称为Apoptosis,或称Programmedcelldeath（细胞程序化死亡）,也有称为Suicide（细胞自杀现象）。细胞死亡（apoptosis）是在细胞内和细胞外因子的严格控制下～种有步骤有活性的生理性自行消亡过程,正在死亡的细胞形态上发生明显的变化,如胞浆浓缩,原生质膜形…     

细胞程序性死亡的非Caspase依赖型途径

程序性细胞死亡是一种程序化的主动性细胞死亡,半胱胺酸天冬氨酸特异性蛋白酶家族(在该过程中起着不可忽视的作用.基于Caspase在程序性细胞死亡过程中所起的作用,将程序性细胞死亡分为两大类:Caspase依赖型和Caspase非依赖型.前者即典型的凋亡,后者包括自体吞噬、副凋亡、有丝分裂灾变、凋亡样程序性死亡、坏死样程序性死亡等.这些Caspase非依赖型的细胞程序性死亡途径与生理及病理现象密切相关.     

Epigenetics and Cell Death: DNA Hypermethylation in Programmed Retinal Cell Death

Background

Vertebrate genomes undergo epigenetic reprogramming during development and disease. Emerging evidence suggests that DNA methylation plays a key role in cell fate determination in the retina. Despite extensive studies of the programmed cell death that occurs during retinal development and degeneration, little is known about how DNA methylation might regulate neuronal cell death in the retina.

Methods

The developing chicken retina and the rd1 and rhodopsin-GFP mouse models of retinal degeneration were used to investigate programmed cell death during retinal development and degeneration. Changes in DNA methylation were determined by immunohistochemistry using antibodies against 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC).

Results

Punctate patterns of hypermethylation paralleled patterns of caspase3-dependent apoptotic cell death previously reported to occur during development in the chicken retina. Degenerating rd1 mouse retinas, at time points corresponding to the peak of rod cell death, showed elevated signals for 5mC and 5hmC in photoreceptors throughout the retina, with the most intense staining observed in the peripheral retina. Hypermethylation of photoreceptors in rd1 mice was associated with TUNEL and PAR staining and appeared to be cCaspase3-independent. After peak rod degeneration, during the period of cone death, occasional hypermethylation was observed in the outer nuclear layer.

Conclusion

The finding that cell-specific increases of 5mC and 5hmC immunostaining are associated with the death of retinal neurons during both development and degeneration suggests that changes in DNA methylation may play a role in modulating gene expression during the process of retinal degeneration. During retinal development, hypermethylation of retinal neurons associates with classical caspase-dependent apoptosis as well as caspase-3 independent cell death, while hypermethylation in the rd1 mouse photoreceptors is primarily associated with caspase-3 independent programmed cell death. These findings suggest a previously unrecognized role for epigenetic mechanisms in the onset and/or progression of programed cell death in the retina.     

植物衰老中的编程性细胞死亡

本文通过对植物衰老和动植物中编程性细胞死亡(PCD)的研究,阐述了植物衰老中PCD存在的依据,澄清了植物衰老和PCD的关系,提出了植物衰老中可能的PCD发生途径,为调控植物衰老的遗传操作提供依据.     

植物衰老中的编程性细胞死亡

本文通过对植物衰老和动植物中编程性细胞死亡(PCD)的研究,阐述了植物衰老中PCD存在的依据,澄清了植物衰老和PCD的关系,提出了植物衰老中可能的PCD发生途径,为调控植物衰老的遗传操作提供依据。     

细胞程序性死亡的判定方法

胞程序性死亡有别于一般意义上的死亡──坏死,所以对其判定显得格外重要．国内外目前常从细胞形态、细胞膜完整性、DNA及某些重要生化指标等方面进行判定．