泛素/26S蛋白酶体系统介导的细胞程序化死亡

在一定的生理或者病理条件下,细胞为了自身发育或者抵御不良刺激,会采取细胞程序化死亡（programmed cell death,PCD）的方式结束生命。泛素/26S蛋白酶体系统（ubiquitin-26S proteasome system,UPS）作为生物体中重要的翻译后蛋白质调节系统,对PCD起着关键的调节作用。该文介绍UPS通过两条细胞凋亡信号转导通路以及天冬氨酸特异性半胱氨酸蛋白酶来调控PCD的研究进展。     

Induction of autophagy supports the bioenergetic demands of quiescent muscle stem cell activation

The exit of a stem cell out of quiescence into an activated state is characterized by major metabolic changes associated with increased biosynthesis of proteins and macromolecules. The regulation of this transition is poorly understood. Using muscle stem cells, or satellite cells (SCs), we found that autophagy, which catabolizes intracellular contents to maintain proteostasis and to produce energy during nutrient deprivation, was induced during SC activation. Inhibition of autophagy suppressed the increase in ATP levels and delayed SC activation, both of which could be partially rescued by exogenous pyruvate as an energy source, suggesting that autophagy may provide nutrients necessary to meet bioenergetic demands during this critical transition from quiescence to activation. We found that SIRT1, a known nutrient sensor, regulates autophagic flux in SC progeny. A deficiency of SIRT1 led to a delay in SC activation that could also be partially rescued by exogenous pyruvate. These studies suggest that autophagy, regulated by SIRT1, may play an important role during SC activation to meet the high bioenergetic demands of the activation process.     

Activation—induced cell death in B lymphocytes

Upon encountering the antigen(Ag),the immune system can either develop a specific immune response of enter a specific state of unresponsiveness,tolerance.The response of B cells to their specific Ag can be activation and proliferation,leading to the immune response,or anergy and activation-induced cell death(AICD),leading to tolerance.AICD in B lymphocytes is a highly regulated event initiated by crosslinking of the B cell receptor (BCR).BCR engagement initiates several signaling events such as activation of PLCγ,Ras,and PI3K,which generally speaking,lead to survival.However,in the absence of survival signals(CD40 or IL-4R engagement),BCR crosslinking can also promote apoptotic signal transduction pathways such as activation of effector caspases,expression of pro-apoptotic genes,and inhibition of pro-survival genes.The complex interplay between survival and death signals determines the B cell fate and, consequently,the immune response.     

A cellular suicide strategy of plants: vacuole-mediated cell death

Programmed cell death (PCD) occurs in animals and plants under various stresses and during development. Recently, vacuolar processing enzyme (VPE) was identified as an executioner of plant PCD. VPE is a cysteine protease that cleaves a peptide bond at the C-terminal side of asparagine and aspartic acid. VPE exhibited enzymatic properties similar to that of a caspase, which is a cysteine protease that mediates the PCD pathway in animals, although there is limited sequence identity between the two enzymes. VPE and caspase-1 share several structural properties: the catalytic dyads and three amino acids forming the substrate pockets (Asp pocket) are conserved between VPE and caspase-1. In contrast to such similarities, subcellular localizations of these proteases are completely different from each other. VPE is localized in the vacuoles, while caspases are localized in the cytosol. VPE functions as a key molecule of plant PCD through disrupting the vacuole in pathogenesis and development. Cell death triggered by vacuolar collapse is unique to plants and has not been seen in animals. Plants might have evolved a VPE-mediated vacuolar system as a cellular suicide strategy.     

A possible intermediate step during apoptotic execution

Many proteases are known to be involved in apoptosis. Among them, interleukin-1beta converting enzyme (ICE) and its family proteases, which are called caspases, play critical roles in the execution stage of apoptosis. We previously reported that a proteasome-inhibitor, benzyloxycarbonyl Leu-Leu-leucinal (ZLLLal), induced apoptosis in MOLT-4 cells. In the present study, in order to analyze the detailed mechanism of ZLLLal-induced apoptosis, we examined the effect of a caspase-inhibitor, acetyl(Ac)-Tyr-Val-Ala-Asp-chloromethyl ketone (AcYVADcmk), on ZLLLal-induced apoptosis in the cells. Agarose gel electrophoresis revealed that low concentrations of AcYVADcmk efficiently suppressed apoptotic DNA fragmentation. However, the cells presented morphology different from normal, apoptotic or necrotic cells, although DNA fragmentation was suppressed. The same examination was performed on the cells with anti-Fas antibody-induced apoptosis, and the same results were obtained. Some cells with a similar morphology were found even without the caspase-inhibitor in the early stage of anti-Fas antibody-induced physiological apoptosis. In addition, apoptotic cascade was reactivated by washing out the caspase inhibitor from the DNA degradation-suppressed cells. Therefore, this newly found morphological feature shows the presence of a step prior to caspase activation in the cells, and this is the first report presenting the pre-caspase-activated step in the apoptotic cascade.     

Activation of multiple caspases and modification of cell surface fas (CD95) in proteasome inhibitor-induced apoptosis of rat natural killer cells

The proteasome is a multi-subunit protease complex that is involved in intracellular protein degradation in eukaryotes. Previously, we have reported that selective, synthetic chymotryptic proteasome inhibitors inhibit A-NK cell-mediated cytotoxicity by approximately 50%; however, the exact role of the proteasome in NK cell-mediated cytotoxicity remains unknown. Herein, we report that proteasome inhibitors, MG115 and MG132, decreased the proteasome chymotrypsin-like activity in the rat natural killer cell line RNK16 by 85% at a concentration of 5 microM. The viability of RNK16 cells was also reduced in the presence of these inhibitors. Both inhibitors induced the apoptosis of RNK16 cells, as shown by DNA fragmentation, caspase-3 activation and the appearance of sub-G-cell populations. An increase in the fraction of apoptotic cells was observed in a dose- and time-dependent manner in our studies. In addition, the activity of caspase-1, -2, -6, -7, -8, and -9, was increased following the treatment of RNK16 cells with these inhibitors. Further investigation revealed that the expression of Fas (CD95) protein on the RNK16 cell surface was increased after the treatment by MG115 or MG132, indicating that apoptosis induced by proteasome inhibitors in RNK16 cells might be mediated through the Fas (CD95)-mediated death pathway as well. Our studies indicate, for the first time, that proteasomal chymotryptic inhibitors can reduce natural killer cell viability and therefore indirectly inhibit cell-mediated cytotoxicity via the apoptosis-inducing properties of these agents.     

Active participation of cell death in development and organismal homeostasis

The precise, regional execution of programmed cell death is required for the proper patterning and sculpting of the embryonic primordium during animal development. In addition, cell death that is not directly involved in sculpting is also widely observed. The most abundant morphological form of programmed cell death in developing animals is apoptosis, and identification of the apoptotic genetic pathways has enabled the study of apoptosis' regulation and roles during development. Genetic and bio-imaging studies have permitted the study of the active roles of cell death in development and organismal homeostasis.     

Caspase-3-mediated degradation of condensin Cap-H regulates mitotic cell death

Mitotic death is a major form of cell death in cancer cells that have been treated with chemotherapeutic drugs. However, the mechanisms underlying this form of cell death is poorly understood. Here, we report that the loss of chromosome integrity is an important determinant of mitotic death. During prolonged mitotic arrest, caspase-3 is activated and it cleaves Cap-H, a subunit of condensin I. The depletion of Cap-H results in the loss of condensin I complex at the chromosomes, thus affecting the integrity of the chromosomes. Consequently, DNA fragmentation by caspase-activated DNase is facilitated, thus driving the cell towards mitotic death. By expressing a caspase-resistant form of Cap-H, mitotic death is abrogated and the cells are able to reenter interphase after a long mitotic delay. Taken together, we provide new insights into the molecular events that occur during mitotic death.     

Proteasomal inhibition and apoptosis regulatory changes in human isolated lymphocytes: the synergistic role of dopamine

Abnormal deposition of protein aggregates and increased susceptibility to apoptotic cell death may result from defects in the activity of the ubiquitin-proteasome system (UPS); neurotoxicity related to UPS defects seems to require dopamine to be fully expressed. The aim of this study was to investigate the pro-apoptotic effects caused by proteasomal activity inhibition, as well as the synergistic effect of dopaminergic stimulation in human lymphocytes isolated from healthy volunteers. Cells were incubated 20 h at 37 degrees C, with: (1) lactacystin, (2) increasing concentrations of dopamine or (3) mixture of dopamine and lactacystin. Activities of proteasome 20S and pro-apoptotic caspases-3 and -9 and levels of anti-apoptotic Bcl-2 were measured with fluorimetric or immunochemical assays, while a "DNA diffusion" assay was used to determine the apoptosis. Incubation of lymphocytes with lactacystin, which caused reduction of proteasomal activity, was associated with activation of caspases. A clear, dose-dependent reduction of proteasomal activity was also seen in the presence of increasing doses of dopamine, which was accompanied by a slight dose-dependent increase of caspases activities and Bcl-2 levels. Both effects on proteasome and caspase activities were enhanced when cells were simultaneously exposed to lactacystin and elevated concentrations of dopamine. Apoptosis was detected in all treated samples, but not in controls, without significant differences among the treatment groups; however, the association of dopamine and lactacystin induced a clear reduction in the number of cells being analyzed, pointing to marked cytotoxicity. Our data confirm the potentiation of cytotoxicity related to proteasome inhibition, in the presence of dopaminergic stimulation.     

Programmed cell death and the proteasome

A characteristic feature of apoptotic cell death is the activation of a cascade of cytoplasmic proteases that results in the cleavage of a limited number of target proteins. A central role in these proteolytic events has been assigned to members of the capase family. However, the use of low molecular weight proteasomal inhibitors has also demonstrated that protein degradation or processing by the ubiquitin-proteasome system of the cell has a decisive impact on cell survival and death as well, depending on the cell type and/or the proliferative status of the cells studied. Treatment of proliferating cells with proteasome inhibitors leads to cell death, potentially involving an internal signalling conflict between accumulating levels of the cdk inhibitor p27Kip1 and c-myc. In contrast, in terminally differentiated cells the same compounds have the opposite effect of blocking apoptosis, possibly by preventing proteasome-mediated degradation of a capase inhibitor. In this review the role of proteasome-mediated proteolysis in the dying cell is discussed and apparently conflicting results are integrated into a working hypothesis which functionally locates the proteasome upstream of capase3-like enzymes.     

Chemical Inactivation of Lipase in Organic Solvent: A Lipase from Pseudomonas aeruginosa TE3285 is More Like a Typical Serine Enzyme in an Organic Solvent than in Aqueous Media

A microbial lipase from Pseudomonas aeruginosa TE3285 was treated in anhydrous diisopropyl ether with three kinds of serine-reactive reagents, ethyl p-nitrophenyl methylphosphonate (ENMP), diisopropyl fluorophosphate (DFP), and phenylmethylsulfonyl fluoride (PMSF) to lose its catalytic activity for both transesterification in an organic solvent and ester hydrolysis in aqueous system. In contrast with the facile inactivation in an organic solvent, no or very slow inactivation was observed in an aqueous solution. The lipase was shown to behave more like a typical serine enzyme in an organic solvent than in aqueous solution with regard to the chemical inactivation by serine-reactive reagents. The unique behavior of the lipase in an organic solvent may be associated with inferfacial activation of the lipase, which is one of the most distinct characteristics of the lipase family, and the activiation of lipase could be induced by a hydrophobic interaction with an organic solvent.     

The role of vacuole in plant cell death

Almost all plant cells have large vacuoles that contain both hydrolytic enzymes and a variety of defense proteins. Plants use vacuoles and vacuolar contents for programmed cell death (PCD) in two different ways: for a destructive way and for a non-destructive way. Destruction is caused by vacuolar membrane collapse, followed by the release of vacuolar hydrolytic enzymes into the cytosol, resulting in rapid and direct cell death. The destructive way is effective in the digestion of viruses proliferating in the cytosol, in susceptible cell death induced by fungal toxins, and in developmental cell death to generate integuments (seed coats) and tracheary elements. On the other hand, the non-destructive way involves fusion of the vacuolar and the plasma membrane, which allows vacuolar defense proteins to be discharged into the extracellular space where the bacteria proliferate. Membrane fusion, which is normally suppressed, was triggered in a proteasome-dependent manner. Intriguingly, both ways use enzymes with caspase-like activity; the membrane-fusion system uses proteasome subunit PBA1 with caspase-3-like activity, and the vacuolar-collapse system uses vacuolar processing enzyme (VPE) with caspase-1-like activity. This review summarizes two different ways of vacuole-mediated PCD and discusses how plants use them to attack pathogens that invade unexpectedly.     

Parkin attenuates manganese-induced dopaminergic cell death

Manganese as environmental factor is considered to cause parkinsonism and induce endoplasmic reticulum stress-mediated dopaminergic cell death. We examined the effects of manganese on parkin, identified as the gene responsible for familial Parkinson's disease, and the role of parkin in manganese-induced neuronal cell death. Manganese dose-dependently induced cell death of dopaminergic SH-SY5Y and CATH.a cells and cholinergic Neuro-2a cells, and that the former two cell types were more sensitive to manganese toxicity than Neuro-2a cells. Moreover, manganese increased the expression of endoplasmic reticulum stress-associated genes, including parkin, in SH-SY5Y cells and CATH.a cells, but not in Neuro-2a cells. Treatment with manganese resulted in accumulation of parkin protein in SH-SY5Y cells and its redistribution to the perinuclear region, especially aggregated Golgi complex, while in Neuro-2a cells neither expression nor redistribution of parkin was noted. Manganese showed no changes in proteasome activities in either cell. Transient transfection of parkin gene inhibited manganese- or manganese plus dopamine-induced cell death of SH-SY5Y cells, but not of Neuro-2a cells. Our results suggest that the attenuating effects of parkin against manganese- or manganese plus dopamine-induced cell death are dopaminergic cell-specific compensatory reactions associated with its accumulation and redistribution to perinuclear regions but not with proteasome system.     

Mechanism of basic fibroblast growth factor-induced cell death

Basic fibroblast growth factor (bFGF) is a potent mitogen for a number of different cell types. Its over-expression has been implicated in transformation and malignant progression. The use of bFGF to treat malignancy is therefore counterintuitive. However, recent studies have shown bFGF-induces cell death in some tumour types. This mini review will summarise the most recent findings on bFGF-induced cell death and discuss its potential mechanism of action.     

Definition and stabilisation of the quiescent centre in rice roots

The definition of a quiescent centre (QC) in Arabidopsis has been adequately demonstrated. However, the QC structure of rice has not yet been described in detail. In this research, using histological and marker gene expression analysis, we concluded that the rice QC is very small, and is similar to that of Arabidopsis. Next we investigated the stability of the rice QC during nutrient deficiencies or external hormone treatments, and found that nutrient deficiencies, auxin treatment and cytokinin treatment did not change the cell patterns of the QC. However, ethylene induced irregular transverse cell divisions in the QC and changed formative cell divisions of the ground tissue stem cells (GTSCs) in rice.     

Caspases: potential targets for regulating cell death

While in multicellular organisms all cells inexorably die, there are several different ways provided for the realization of cell death. One of them, apoptosis, represents a universal energy-dependent and tightly regulated physiologic process of cell death in both normal and pathologic tissues. The execution of apoptosis appears to be uniformly mediated through consecutive activation of the members of a caspase family. This review briefly summarizes current knowledge on the molecular mechanisms of caspase activation and the inhibitory components of caspase cascades. The suitability of caspases as a new potential therapeutic target is discussed next. Particular attention is focused on two broad categories of caspase-directed compounds: highly specific caspase inhibitors that distinctly block the progress of apoptosis and caspase activators that selectively induce cell death in a variety of in vitro and in vivo systems. These agents promise to be useful clinically, either alone or in combination with more conventional therapeutics.