FAK/mTOR信号通在肿瘤细胞中的调控作用

粘附斑激酶（focal adhesion kinase,FAK）是一种胞质非受体酪氨酸激酶,是整合素信号通路里一个重要的调节因子,在肿瘤细胞中高表达,与细胞迁移、粘附和侵袭有关。mTOR (mammalian target of rapamycin)是非典型性的Ser/Thr激酶,属于PIKK（phosphatidylinositol kinase related kinase）超家族,介导营养信号调控细胞生长、分化及代谢等生理过程。近年的研究发现FAK通过三条途径与mTOR相关联,组成FAK/mTOR信号通路,在肿瘤细胞的增殖、迁移及肿瘤微环境中发挥着重要的调控作用。本文综述了FAK、mTOR和FAK/mTOR信号通路的特点及对肿瘤细胞调控作用的研究概况,为肿瘤治疗提供参考。     

ILK—整合素信号传导通路中的关键激酶

胸外基质与细胞的相互作用主要由整合素介导。由整合素介导的机械和生化信号调控胞浆激酶、生长因子受体、离子通道的活性并掏胞内肌动蛋白细胞骨架的组装。众多由整合素介导的信号传导通路最后可归于对细胞周期的调节、决定细胞存活或死亡、增殖或者退出细胞周期和分化。近年的研究发现,整合素连接激酶（inte-grin-linked kinase,ILK）在这些信号传导通路中具有关键作用。     

蛋白酪氨酸磷酸酶SHP-2与疾病的相关性研究进展

蛋白酪氨酸磷酸酶家族由130多种蛋白酪氨酸磷酸酶组成,它们和蛋白质酪氨酸激酶家族一起调控蛋白质中酪氨酸残基的磷酸化以及去磷酸化的动态平衡,它们的活性直接决定细胞内蛋白质的磷酸化水平的高低。SHP-2是蛋白酪氨酸磷酸酶家族的一员,在各种细胞和组织中均有广泛的表达,参与多个信号传导通路,介导细胞的生长、分化、迁移、粘附及凋亡等。SHP-2的表达异常会导致多种疾病的产生,但是相关综述较少,同时未见文献报道其在胶质瘤中的作用,因此本文简要介绍SHP-2的结构、功能、信号传导,并阐述了SHP-2与常见疾病的关系。     

粘着斑激酶

粘着斑激酶（ＦＡＸ）是一种非受体型酪氨酸激酶,在整合素介导的信号传导中起重要作用;与细胞的粘附,迁移、增殖和凋亡密切相关,在血管成型术后再狭窄,动脉粥样硬化和肿瘤的转移中起一定的作用     

Wnt/β-catenin信号通路对细胞凋亡和坏死的调控研究进展

Wnt信号通路是一条与细胞增殖分化和机体平衡密切相关且高度保守的信号通路,主要包括Wnt/β-catenin信号通路、Wnt-Ca2+信号通路和平面细胞极性信号通路。其中,以经典Wnt/β-catenin信号炎性反应和细胞命运方面的研究最为深入。现已证实,Wnt/β-catenin信号对细胞命运的调控作用具有两面性,不仅通过调节Survivin、Cyclin、C-myc等基因的表达抑制一些肿瘤细胞凋亡,而且可通过上调促凋亡蛋白BIM、Bax和下调抗凋亡蛋白Mcl-1、Bcl-xl的表达量来促进细胞凋亡。同时,该信号还可以通过抑制某些炎性因子的过度分泌,并下调活性氧(reactive oxygen species,ROS)的含量及坏死相关蛋白PARP-1的表达来抑制细胞坏死。该文对Wnt/β-catenin信号对细胞凋亡和坏死的调控研究进展进行综述。     

人参皂苷-Rh2诱导肿瘤细胞凋亡信号转导通路研究进展

人参皂苷-Rh2诱导肿瘤细胞凋亡机制复杂,且不同机制间相互联系。它主要通过调节细胞周期蛋白依赖性激酶复合物特异性信号传递引发细胞周期阻滞诱导细胞凋亡,或通过调控细胞信号转导通路,如ROS、Ca2+、PKC、JNK介导的线粒体信号通路和TRAIL-R1(DR4)介导的死亡受体信号通路等诱导肿瘤细胞凋亡。     

JNK激酶介导mGluR1对细胞凋亡的不同效应

代谢型谷氨酸受体1（mGluR1）可以通过激活多条信号通路促进或抑制细胞凋亡.然而,导致这种生理功能差异的机制尚不明确.本研究选用两种细胞系,即大鼠神经胶质瘤细胞系（C6）和人胚胎肾细胞（HEK293）分别研究内源性和外源转染的mGluR1的激活对细胞凋亡的影响及其调节机制. 结果显示,内源性mGluR1的活化能够激活PI3K/ERK/JNK通路,抑制凋亡试剂STS诱导的细胞凋亡;而外源转染的mGluR1的活化能够分别激活PI3K/ERK和JNK通路,同时促进STS诱导的应激损伤. HEK293细胞中,应用JNK通路抑制剂SP600125,能够部分抑制由mGluR1激活介导的caspase 3的剪切和细胞凋亡;而在C6细胞中阻断JNK通路,则加剧了由mGluR1活化而引起的细胞凋亡. 本文结果提示：mGluR1通过不同信号通路影响细胞凋亡,其中JNK通路可能是调控细胞凋亡的关键途径.本文为受体激活对细胞凋亡能够产生不同的调控作用提供了相应的证据.     

黏着斑激酶与肿瘤侵袭转移

黏着斑激酶（focal adhesion kinase,FAK）是一种非受体型蛋白酪氨酸激酶,在肿瘤细胞的侵袭和转移中起着重要的作用。FAK是整合素介导的或生长因子受体诱导的调节细胞迁移的信号通路的关键组分。FAK通过与相关分子作用可以调节细胞骨架重构、胞外基质降解、细胞黏附更新以及质膜突出,进而参与肿瘤细胞的运动等多个过程,所以FAK与肿瘤发展的关系已经越来越受到重视。     

神经酰胺诱导肿瘤细胞凋亡的机制研究进展

神经酰胺(ceramide,Cer)作为一种神经鞘磷脂分子,不仅是细胞膜的组成成分,而且可以作为各种信号转导途径的第二信使,参与细胞增殖、分化、衰老和凋亡等生命活动的调节。Cer的合成、代谢及信号转导在肿瘤发生发展甚至耐药和抵抗放射治疗中有着密切的关系。Cer可以被诸如肿瘤坏死因子α(Tumor necrosis factorα,TNF-α)、激素、电离辐射和化疗药物等细胞外信号和受体激活,其主要可以通过内源性凋亡途径和外源性凋亡途径诱导肿瘤细胞凋亡的发生。在肿瘤细胞凋亡发生过程中,Cer通过激活Jun氨基末端激酶(JNKs)、有丝分裂原活化蛋白激酶/细胞外信号调节蛋白激酶(MAPK/ERK)和P38等信号通路以及蛋白激酶、组织蛋白酶D、蛋白磷酸酶1(Protein phosphatase1,PP1)和蛋白磷酸酶2A(Protein phosphatase2A,PP2A)等效应分子介导肿瘤细胞凋亡。本文综述近年来有关Cer在应激反应级联以及肿瘤细胞凋亡中的作用的研究进展。     

Characterization of Nuclear Localization Signal in the N Terminus of Integrin-linked Kinase-associated Phosphatase (ILKAP) and Its Essential Role in the Down-regulation of RSK2 Protein Signaling

Integrin-linked kinase-associated phosphatase (ILKAP) is a serine/threonine (S/T) phosphatase that belongs to the protein phosphatase 2C (PP2C) family. Many previous studies have demonstrated that ILKAP plays key roles in the regulation of cell survival and apoptosis. Researchers have thus far considered ILKAP a cytoplasmic protein that negatively regulates integrin signaling by interacting with and phosphorylating integrin-linked kinase 1 (ILK1). In this study, we found that both endogenous and tagged ILKAP mainly localize to the nucleus and that the nuclear transport of ILKAP is nuclear localization signal (NLS) importin-mediated. The ILKAP protein interacts directly with importin α1, α3, and α5. The NLS in ILKAP is located in the N-terminal region between amino acids 71 and 86, and the NLS-deleted ILKAP protein was distributed in the cytoplasm. In addition, we show that Lys-78 and Arg-79 are critical for the binding of ILKAP to importin α. We also found that nuclear ILKAP interacts with ribosomal protein S6 kinase-2 (RSK2) and induces apoptosis by inhibiting RSK2 activity and down-regulating the expression level of the RSK2 downstream substrate cyclin D1. These results indicate that ILKAP is a nuclear protein that regulates cell survival and apoptosis through the regulation of RSK2 signaling.     

Probing protein phosphatase substrate binding: affinity pull-down of ILKAP phosphatase 2C with phosphopeptides

Proteomics and high throughput analysis for systems biology can benefit significantly from solid-phase chemical tools for affinity pull-down of proteins from complex mixtures. Here we report the application of solid-phase synthesis of phosphopeptides for pull-down and analysis of the affinity profile of the integrin-linked kinase associated phosphatase (ILKAP), a member of the protein phosphatase 2C (PP2C) family. Phosphatases can potentially dephosphorylate these phosphopeptide substrates but, interestingly, performing the binding studies at 4 °C allowed efficient binding to phosphopeptides, without the need for phosphopeptide mimics or phosphatase inhibitors. As no proven ILKAP substrates were available, we selected phosphopeptide substrates among known PP2Cδ substrates including the protein kinases: p38, ATM, Chk1, Chk2 and RSK2 and synthesized directly on PEGA solid supports through a BAL type handle. The results show that phosphopeptides tethered to a flexible solid support bind with high affinity and specificity to ILKAP, which is pulled down from lysates of cells transfected with ILKAP cDNA. Phosphorylation on Ser or Thr residues is important for binding of ILKAP, but sequences around the phosphorylated residue are important for the binding affinity of ILKAP. We conclude that solid-phase affinity pull-down of proteins from complex mixtures can be applied in phosphoproteomics and systems biology.     

Modulation of integrin signal transduction by ILKAP, a protein phosphatase 2C associating with the integrin-linked kinase, ILK1

ILKAP, a protein serine/threonine (S/T) phosphatase of the PP2C family, was isolated in a yeast two-hybrid screen baited with integrin-linked kinase, ILK1. Association of ILK1 and ILKAP was independent of the catalytic activity of either partner, as assayed in co-precipitation and two-hybrid experiments. Condi tional expression of ILKAP in HEK 293 cells resulted in selective inhibition of ECM- and growth factor-stimulated ILK1 activity, but did not inhibit Raf-1 kinase activity. A catalytic mutant of ILKAP, H154D, did not inhibit ILK1 kinase activity. Two cellular targets of ILK1, glycogen synthase kinase 3 beta (GSK3beta) and protein kinase B (PKB)/AKT, were differentially affected by ILKAP-mediated inhibition of ILK1. Catalytically active, but not mutant ILKAP, strongly inhibited insulin-like growth factor-1-stimulated GSK3beta phosphorylation on Ser9, but did not affect phosphorylation of PKB on Ser473, suggesting that ILKAP selectively affects ILK-mediated GSK3beta signalling. Consistent with this, active, but not H154D mutant or the related PP2Calpha, selectively inhibited transactivation of a Tcf/Lef reporter gene, TOPFlash, in 293 cells. We propose that ILKAP regulates ILK1 activity, targeting ILK1 signalling of Wnt pathway components via modulation of GSK3beta phosphorylation.     

Palladin is a novel binding partner of ILKAP in eukaryotic cells

Palladin was a novel binding partner of ILKAP in eukaryotic cells. Palladin’s C-terminal fragment including only its last three Ig domains (residues 710–1106) and the PP2C domain of ILKAP (residues 108–392) were necessary and sufficient for their interaction. The biological significance of the interaction between palladin and ILKAP was that palladin recruited the cytoplasmic ILKAP to initiate ILKAP-induced apoptosis. Our results suggested that palladin played a specific role in modulating the subcellular localization of the cytoplasmic ILKAP and promoting the ILKAP-induced apoptosis.     

Socs3 induction by PPARγ restrains cancer-promoting inflammation

Integrin-linked kinase (ILK) plays key roles in a variety of cell functions, including cell proliferation, adhesion and migration. Within the cell, ILK localizes to multiple sites, including the cytoplasm, focal adhesion complexes that mediate cell adhesion to extracellular substrates, as well as cell-cell junctions in epidermal keratinocytes. Central to understanding ILK function is the elucidation of the mechanisms that regulate its subcellular localization. We now demonstrate that ILK is imported into the nucleus through sequences in its N-terminus, via active transport mechanisms that involve nuclear pore complexes. In addition, nuclear ILK can be rapidly exported into the cytoplasm through a CRM1-dependent pathway, and its export is enhanced by the type 2C protein phosphatase ILKAP. Nuclear localization of ILK in epidermal keratinocytes is associated with increased DNA synthesis, which is sensitive to inhibition by ILKAP. Our studies demonstrate the importance for keratinocyte proliferation of ILK regulation through changes in its subcellular localization, and establish ILKAP and CRM1 as pivotal modulators of ILK subcellular distribution and activity in these cells.     

Role of type 2C protein phosphatases in growth regulation and in cellular stress signaling

A number of interesting features, phenotypes, and potential clinical applications have recently been ascribed to the type 2C family of protein phosphatases. Thus far, 16 different PP2C genes have been identified in the human genome, encoding (by means of alternative splicing) for at least 22 different isozymes. Virtually ever since their discovery, type 2C phosphatases have been predominantly linked to cell growth and to cellular stress signaling. Here, we provide an overview of the involvement of type 2C phosphatases in these two processes, and we show that four of them (PP2Calpha, PP2Cbeta, ILKAP, and PHLPP) can be expected to function as tumor suppressor proteins, and one as an oncoprotein (PP2Cdelta /Wip1). In addition, we demonstrate that in virtually all cases in which they have been linked to the stress response, PP2Cs act as inhibitors of cellular stress signaling. Based on the vast amount of experimental evidence obtained thus far, it therefore seems justified to conclude that type 2C protein phosphatases are important physiological regulators of cell growth and of cellular stress signaling.     

Role of Type 2C Protein Phosphatases in Growth Regulation and in Cellular Stress Signaling

ABSTRACT

A number of interesting features, phenotypes, and potential clinical applications have recently been ascribed to the type 2C family of protein phosphatases. Thus far, 16 different PP2C genes have been identified in the human genome, encoding (by means of alternative splicing) for at least 22 different isozymes. Virtually ever since their discovery, type 2C phosphatases have been predominantly linked to cell growth and to cellular stress signaling. Here, we provide an overview of the involvement of type 2C phosphatases in these two processes, and we show that four of them (PP2Cα, PP2Cβ, ILKAP, and PHLPP) can be expected to function as tumor suppressor proteins, and one as an oncoprotein (PP2Cδ /Wip1). In addition, we demonstrate that in virtually all cases in which they have been linked to the stress response, PP2Cs act as inhibitors of cellular stress signaling. Based on the vast amount of experimental evidence obtained thus far, it therefore seems justified to conclude that type 2C protein phosphatases are important physiological regulators of cell growth and of cellular stress signaling.     

不同磷源对金龟子绿僵菌CQMa102生物量及生成胞外磷酸酶的影响(英文)

金龟子绿僵菌Metarhizium anisopliae是一种广泛分布于世界各地土壤中的重要的昆虫病原真菌。已有研究表明,胞外磷酸酶在绿僵菌侵染并致死寄主过程中发挥了重要作用。利用摇瓶培养方法探究了无机磷(KH2PO4)、简单有机磷(植酸钠、磷酸苯二钠)和蛋白有机磷(酪蛋白)为单独磷源条件下,绿僵菌生物量、产胞外酸性磷酸酶以及酪氨酸蛋白磷酸酶、丝/苏氨酸蛋白磷酸酶活性的变化。实验结果显示:加入酪蛋白的培养基,最有利于绿僵菌生长、胞外蛋白的分泌和产酸性磷酸酶;其次为加入KH2PO4和磷酸苯二钠的培养基;加入植酸钠的培养基不利于绿僵菌的生长代谢。然而,加入磷酸苯二钠的培养基,最有利于酪氨酸蛋白磷酸酶、丝/苏氨酸蛋白磷酸酶的分离纯化。     

Protein phosphatase 2C is involved in the cAMP-dependent ciliary control in Paramecium caudatum

Forward swimming of the Triton-extracted model of Paramecium is stimulated by cAMP. Backward swimming of the model induced by Ca(2+) is depressed by cAMP. Cyclic AMP and Ca(2+) act antagonistically in setting the direction of the ciliary beat. Some ciliary axonemal proteins from Paramecium caudatum are phosphorylated in a cAMP-dependent manner. In the presence of cAMP, axonemal 29- and 65-kDa polypeptides were phosphorylated by endogenous A-kinase in vitro. These phosphoproteins, however, were not dephosphorylated after in vitro phosphorylation, presumably because of the low endogenous phosphoprotein phosphatase activity associated with isolated axonemes. We purified the protein phosphatase that specifically dephosphorylated the 29- and 65-kDa phosphoproteins from Paramecium caudatum. The molecular weight of the protein phosphatase was 33 kDa. The protein phosphatase had common characteristics as protein phosphatase 2C (PP2C). The characteristics of the protein phosphatase were the same as those of the PP2C from Paramecium tetraurelia (PtPP2C) [Grothe et al., 1998: J. Biol. Chem. 273:19167-19172]. We concluded that the phosphoprotein phosphatase is the PP2C from Paramecium caudatum (PcPP2C). The PcPP2C markedly accelerated the backward swimming of the Triton-extracted model in the presence of Ca(2+). On the other hand, the PcPP2C slightly depressed the forward swimming speed. This indicates that the PP2C plays a role in the cAMP-dependent regulation of ciliary movement in Paramecium caudatum through dephosphorylation of 29- and/or 65-kDa regulatory phosphoproteins by terminating the action of cAMP.     

Carboxyl methylation regulates phosphoprotein phosphatase 2A by controlling the association of regulatory B subunits

Phosphoprotein phosphatase 2A (PP2A) is a major phosphoserine/threonine protein phosphatase in all eukaryotes. It has been isolated as a heterotrimeric holoenzyme composed of a 65 kDa A subunit, which serves as a scaffold for the association of the 36 kDa catalytic C subunit, and a variety of B subunits that control phosphatase specificity. The C subunit is reversibly methyl esterified by specific methyltransferase and methylesterase enzymes at a completely conserved C-terminal leucine residue. Here we show that methylation plays an essential role in promoting PP2A holoenzyme assembly and that demethylation has an opposing effect. Changes in methylation indirectly regulate PP2A phosphatase activity by controlling the binding of regulatory B subunits to AC dimers.