Ras蛋白信号途径及其对线虫生长发育的调控作用

Ras蛋白是一个分子质量为21 kD左右的单体GTP酶,具有两种构象:GTP结合构象(Ras.GTP)及GDP结合构象(Ras.GDP),这两种构象在一定条件下可发生互变.由生长因子介导的Ras信号传导途径是诸多信号途径中与细胞增殖、分化密切相关的重要信号途径.受体型TPK/Ras/MAPK信号转导途径是是目前研究的最为清楚的受Ras蛋白调节的信号传导途径,该途径包括受体型酪氨酸蛋白激酶(RTK)、接头蛋白、鸟苷酸释放因子(GNEF)、Ras蛋白以及MAPK级联反应体系.目前,TPK/Ras/MAPK信号转导途径在秀丽杆线虫(Caenorhabolitis elegans中研究的最为清楚:Ras信号途径对于许多发育进程是必需的,包括阴门、子宫、交合刺、P12以及排泄管细胞的诱导分化;控制着性肌原细胞迁移、轴突导向;对细胞减数分裂粗线期具有促进作用.对C.elegans的研究加深了对TPK/Ras/MAPK信号途径结构、突变体表型以及与其他信号途径的互作的了解,将会促进Ras信号途径对植物寄生线虫调控作用的研究.     

靶向作用小G蛋白在骨肉瘤中的研究进展

骨肉瘤(osteosarcoma,OS)是原发性骨肉瘤,具有很强的侵袭性质,虽然患者的生活质量得到了改善,但肢体截肢、肺转移和药物毒性的持续高发率仍未得到解决。小G蛋白(Small GTPase)能通过影响细胞行为的多个方面影响癌症发生发展的进程。实际上,某些小G蛋白如Ras和Rho等家族成员,已被证实其相关生物抑制剂或相关药物在癌症进展中发挥着积极作用。因此靶向调控相关小G蛋白可能成为治疗癌症的潜在策略。本文综述了小G蛋白在骨肉瘤的基本生物机制和多种功能,同时讨论研究结合新的治疗方法来靶向小G蛋白治疗骨肉瘤。     

RIN1蛋白在肿瘤发病中的作用研究进展

RIN1蛋白是人体内第一个特征性Ras效应蛋白,不仅可以通过调节Ras与Rab5蛋白的结合介导EGFR、TβR等受体的内吞作用,还可以直接激活ABL酪氨酸激酶活性。已有研究发现RIN1在膀胱癌、结肠癌、慢性粒细胞白血病等多种肿瘤的发生发展中发挥了重要作用。本文就此进行简要综述。     

PCR法检测水稻中存在Ras类蛋白基因家族(简报)

Ras类蛋白家族是普遍存在于动物及低等真核生物细胞中的一类单亚基GTP结合蛋白(20—29kDa),它们在一系列细胞过程中起重要作用。它们有GTP结合蛋白共有的作用机制。在刺激物诱导下,结合GTP成为活化状态,与效应子作用产生一定效应;GTP水解成GDP后恢复GDP结合状态,即非活化状态。它们通过GTPase驱动的构象变化的循环,作为“分子开关”起调控作用。目前已有40多种Ras类蛋白被发现,按结构可分为Ras、     

植物小G 蛋白功能的研究进展

近年来,小G蛋白的调控途径已经成为人们研究细胞信号转导过程的热点问题。小G蛋白家族包括Ras、Rab、Rho、Arf和Ran亚家族,它们起着许多不同的重要细胞生理作用,例如基因表达、细胞骨架重组装、微管的形成以及囊泡和核孔运输机制。这些小G蛋白作为重要的分子开关,具有一个非常保守的功能区域,即I-IV结构区,它起着关键性作用。从拟南芥(Arabidopsis thaliana)基因组预测分析得出,拟南芥含有93个小G蛋白同源序列,包含Rab、Rho、Arf和Ran亚家族,但没有Ras亚家族。本文主要阐述了迄今在植物中研究小G蛋白各个亚家族功能的最新进展,并对植物、酵母和动物相关的同源蛋白的生理功能进行比较和推测。     

RASSF1A在细胞自噬及凋亡中的功能

肿瘤抑制因子Ras相关结构域家族成员1A（Ras association domain family 1A，RASSF1A）是Ras超家族蛋白重要的下游效应因子，具有调控自噬及凋亡的作用。自噬及凋亡是影响机体生存发育的重要生命过程，其调节紊乱与肿瘤的发生发展密切相关。本文针对RASSF1A对自噬及凋亡的调节机制及其与肿瘤发生发展之间的关系展开综述，分析翻译后修饰对于RASSF1A调节自噬及凋亡过程中功能切换的作用，探讨自噬及凋亡在肿瘤发生中的调节作用，以期为RASSF1A启动子高甲基化型肿瘤的治疗提供新思路。     

植物小G蛋白功能的研究进展

近年来,小G蛋白的调控途径已经成为人们研究细胞信号转导过程的热点问题.小G蛋白家族包括Ras、Rab、Rho、Arf和Ran亚家族,它们起着许多不同的重要细胞生理作用,例如基因表达、细胞骨架重组装、微管的形成以及囊泡和核孔运输机制.这些小G蛋白作为重要的分子开关,具有一个非常保守的功能区域,即I-Ⅳ结构区,它起着关键性作用.从拟南芥(Arabidopsisthaliana)基因组预测分析得出,拟南芥含有93个小G蛋白同源序列,包含Rab、Rho、Arf和Ran亚家族,但没有Ras亚家族.本文主要阐述了迄今在植物中研究小G蛋白各个亚家族功能的最新进展,并对植物、酵母和动物相关的同 源蛋白的生理功能进行比较和推测.     

RhoA的泛素化降解机制及功能

Rho小G蛋白(Ras homology frowth-related,Rho G)家族作为分子开关(molecular switch)在GTP结合的激活形式和GDP结合的非激活形式之间转换,发挥着重要的生物学功能,细胞内Rho小G蛋白的含量可由泛素–蛋白酶体系统(ubiquitin-proteasome system,UPS)降解途径来调控。Rho A(Ras homolog gene family member A,Rho A)是Rho小G蛋白家族成员,其功能涉及细胞极性、细胞迁移、细胞周期调控、神经系统发育等,通过UPS途径对该蛋白在细胞内的含量进行调控,可保证细胞的相关正常生理功能。在Rho A泛素化降解过程中,不同的泛素连接酶(ubiquintin ligases,E3)发挥了重要的作用。该文将简单介绍UPS的过程和Rho A蛋白质的结构、功能,详细论述Rho A泛素化降解过程的分子机制和生物学功能。     

SOS1蛋白病理生理功能研究进展

SOS1(son of sevenless homolog 1)蛋白是一种在细胞中广泛表达的调控蛋白。作为信号通路中的关键蛋白,SOS1在细胞内许多信号转导通路中起着重要的调控作用,例如参与调控Ras和Rac信号通路。SOS1的异常表达或突变与临床疾病的发生密切相关。本文对SOS1的功能及其在生理学、病理生理学中的作用的研究进展进行综述。     

Ras蛋白在地塞米松体外诱导大鼠胚胎垂体生长激素细胞分化过程中的作用

目的:探究Ras蛋白在地塞米松体外诱导大鼠胚胎垂体生长激素细胞分化过程中的作用。方法:本课题利用大鼠胚胎垂体细胞的无血清原代细胞培养技术,在地塞米松诱导生长激素细胞分化的过程中,加入蛋白Ras的抑制剂Manumycin,利用免疫荧光、western-blot、放射免疫分析和MTT等技术对Ras蛋白在糖皮质激素体外诱导生长激素细胞分化中的作用进行研究。结果:地塞米松能够显著提高生长激素阳性细胞百分比和生长激素的含量(P0.01)。加入不同浓度的Manumycin后,地塞米松诱导的生长激素阳性细胞百分比显著降低(P0.01),生长激素的含量亦出现降低(P0.05)。结论:Ras蛋白在地塞米松诱导垂体生长激素细胞分化过程中发挥重要作用。     

Mortalin-MPD (mevalonate pyrophosphate decarboxylase) interactions and their role in control of cellular proliferation

Mortalin (mot-2/GRP75/PBP74/mthsp70) is a member of the hsp70 family of proteins and is differentially distributed in normal and immortal cells. It was shown to be involved in pathways to cell senescence and immortalization. To elucidate its functional aspects, a yeast interactive screen for mortalin (mot-2) binding proteins was performed. Mevalonate pyrophosphate decarboxylase (MPD) was identified as one of the mortalin binding partners. The interactions were confirmed in mammalian cells by two-hybrid assay and in vivo coimmunoprecipitation. MPD is known to furnish prenyl groups required for prenylation, protein modification that is essential for the activity of many proteins including p21(Ras) (Ras). We have examined the effect of MPD-mot-2 interactions on the level and activity of p21(Ras) and its downstream effectors, p44 and p42 MAP kinases (ERK1/ERK2), in Ras-Raf pathway. An overexpression of mot-2 resulted in reduced level of Ras and phosphorylated ERK2. These were rescued by co-expression of MPD from an exogenous promoter demonstrating a functional link between mot-2, MPD, and Ras. Ras and its oncogenic forms act as key players in controlling proliferation of normal and cancerous cells. Assigning mot-2 upstream of p21(Ras) offers an important mechanism for influence over cell proliferation.     

Kinases KISS and tell

The Ras oncoprotein is a key driver of cancer. However, Ras also provokes senescence, which serves as a major barrier to Ras-driven transformation. Ras senescence pathways remain poorly characterized. NORE1A is a novel Ras effector that serves as a tumor suppressor. It is frequently inactivated in tumors. We show that NORE1A is a powerful Ras senescence effector and that down-regulation of NORE1A suppresses senescence induction by Ras and enhances Ras transformation. We show that Ras induces the formation of a complex between NORE1A and the kinase HIPK2, enhancing HIPK2 association with p53. HIPK2 is a tumor suppressor that can induce either proapoptotic or prosenescent posttranslational modifications of p53. NORE1A acts to suppress its proapoptotic phosphorylation of p53 but enhance its prosenescent acetylation of p53. Thus, we identify a major new Ras signaling pathway that links Ras to the control of specific protein acetylation and show how NORE1A allows Ras to qualitatively modify p53 function to promote senescence.     

Diverting a protein from its cellular location by intracellular antibodies. The case of p21Ras.

We describe the use of phage libraries to derive new antibodies against p21Ras to be used for intracellular expression in mammalian cells. A panel of single-chain antibody fragments, binding to Ras, were analyzed and characterized for their capacity to interfere in vitro with (a) the intrinsic GTPase activity of Ras and (b) the binding of Ras to its effector Raf, and were found not to neutralize its function, according to these biochemical criteria. When expressed intracellularly in mouse 3T3 K-Ras transformed cells all the anti-Ras single-chain variable fragments (scFv) tested inhibited cell proliferation, as assessed by bromodeoxyuridine incorporation. Double immunofluorescence analysis of transfected cells using confocal microscopy confirmed that anti-Ras antibody fragments colocalize with endogenous Ras, at subcellular locations where the protein Ras is not normally found. These data suggest that the ability of phage-derived anti-Ras scFv fragments to inhibit the function of Ras in vivo is a rather general and frequent property and that the range of antibodies that can be successfully used for intracellular inhibition studies is much greater than anticipated, exploiting the mode of action of diverting protein traffic.     

Ras-induced cellular events

Ras is a crucial regulator of cell growth in eukaryotic cells. Activated Ras can stimulate signal transduction cascades, leading to cell proliferation, differentiation or apoptosis. It is also one of the most commonly mutated genes in both solid tumours and haematologic neoplasias. In leukaemia and tumours, aberrant Ras signalling can be induced directly by Ras mutation or indirectly by altering genes that associate with Ras or its signalling pathways. A requisite for Ras function is localization to the plasma membrane, which is induced by the post-translational modification farnesylation. Molecules that interfere with this Ras modification have been used as antitumour agents. Ras is emerging as a dual regulator of cell functions, playing either positive or negative roles in the control of proliferation or apoptosis. The diversity of Ras-mediated effects may be related in part to the differential involvement of Ras homologues in distinct cellular processes or to the expanding array of Ras effectors.     

Overexpressed galectin-3 in pancreatic cancer induces cell proliferation and invasion by binding ras and activating ras signaling

Pancreatic cancer (PDAC) is a lethal disease with a five-year survival of 3-5%. Mutations in K-Ras are found in nearly all cases, but K-Ras mutations alone are not sufficient for the development of PDAC. Additional factors contribute to activation of Ras signaling and lead to tumor formation. Galectin-3 (Gal-3), a multifunctional β-galactoside-binding protein, is highly expressed in PDAC. We therefore investigated the functional role of Gal-3 in pancreatic cancer progression and its relationship to Ras signaling. Expression of Gal-3 was determined by immunohistochemistry, Q-PCR and immunoblot. Functional studies were performed using pancreatic cell lines genetically engineered to express high or low levels of Gal-3. Ras activity was examined by Raf pull-down assays. Co-immunoprecipitation and immunofluorescence were used to assess protein-protein interactions. In this study, we demonstrate that Gal-3 was highly up-regulated in human tumors and in a mutant K-Ras mouse model of PDAC. Down-regulation of Gal-3 by lentivirus shRNA decreased PDAC cell proliferation and invasion in vitro and reduced tumor volume and size in an orthotopic mouse model. Gal-3 bound Ras and maintained Ras activity; down-regulation of Gal-3 decreased Ras activity as well as Ras down-stream signaling including phosphorylation of ERK and AKT and Ral A activity. Transfection of Gal-3 cDNA into PDAC cells with low-level Gal-3 augmented Ras activity and its down-stream signaling. These results suggest that Gal-3 contributes to pancreatic cancer progression, in part, by binding Ras and activating Ras signaling. Gal-3 may therefore be a potential novel target for this deadly disease.     

Ras-induced cellular events (review)

Ras is a crucial regulator of cell growth in eukaryotic cells. Activated Ras can stimulate signal transduction cascades, leading to cell proliferation, differentiation or apoptosis. It is also one of the most commonly mutated genes in both solid tumours and haematologic neoplasias. In leukaemia and tumours, aberrant Ras signalling can be induced directly by Ras mutation or indirectly by altering genes that associate with Ras or its signalling pathways. A requisite for Ras function is localization to the plasma membrane, which is induced by the post-translational modification farnesylation. Molecules that interfere with this Ras modification have been used as antitumour agents. Ras is emerging as a dual regulator of cell functions, playing either positive or negative roles in the control of proliferation or apoptosis. The diversity of Ras-mediated effects may be related in part to the differential involvement of Ras homologues in distinct cellular processes or to the expanding array of Ras effectors.     

Dominant inhibitory Ras mutants selectively inhibit the activity of either cellular or oncogenic Ras.

Two dominant inhibitory Ras mutant proteins were analyzed by microinjection. One, [Asn-17]Ras, had a substitution in the putative Mg(2+)-binding site of Ha-Ras. The other, RAST, had a mutation in a yeast RAS protein that impaired its GTPase activity and increased its affinity for GAP. RAST also had a mutation that blocked its localization to the plasma membrane. In NIH 3T3 cells [Asn-17]Ras inhibited the function of normal Ras much more efficiently than that of oncogenic Ras. In contrast, RAST interfered with the transforming activity of oncogenic Ras more efficiently than that of normal Ras. These conclusions were based on two separate types of analysis. The inhibitory Ras mutant proteins were first microinjected into cells stably transformed either by oncogenic Ras or by high levels of expression of cellular Ras. Results obtained in stably transformed cells were then verified by coinjection of the inhibitory Ras mutant proteins together with transforming concentrations of either oncogenic or normal Ras protein. Whereas RAST was active in soluble form. [Asn-17]Ras required membrane localization for activity. Furthermore, mutations in the GAP/effector-binding domain reduced or eliminated the inhibitory activity of RAST but had no detectable effect on [Asn-17]Ras. These results are consistent with the possibility that [Asn-17]Ras functions by blocking the activation of endogenous Ras proteins, while RAST functions by blocking the ability of activated Ras to stimulate a downstream target within the cells. The properties of RAST suggest that interference with the GAP/effector-binding function of RAS represents a strategy for the preferential inactivation of oncogenic Ras in cells.     

Enhancing FTS (Salirasib) efficiency via combinatorial treatment

The Ras oncogene transmits signals, which regulate various cellular processes including cell motility, differentiation, growth and death. Since Ras signalling is abnormally activated in more than 30% of human cancers, Ras and its downstream signalling pathways are considered good targets for therapeutic interference. Ras is post‐translationally modified by the addition of a farnesyl group, which permits its attachment to the plasma membrane. Exploiting this knowledge, a synthetic Ras inhibitor, S‐trans, trans‐farnesylthiosalicylic acid (FTS; Salirasib), was developed. FTS resembles the farnesylcysteine group of Ras, and acts as an effective Ras antagonist. In the present review, the effect of FTS in combination with various other drugs, as tested in vitro and in vivo, and its therapeutic potential are discussed. As reviewed, FTS cooperates with diverse therapeutic agents, which significantly improves treatment outcome. Therefore, combinations of FTS with other agents have a potential to serve as anti‐cancer or anti‐inflammatory therapies.     

Functional crosstalk between AKT/mTOR and Ras/MAPK pathways in hepatocarcinogenesis: Implications for the treatment of human liver cancer

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death worldwide, with limited treatment options. AKT/mTOR and Ras/MAPK pathways are frequently deregulated in human hepatocarcinogenesis. Recently, we generated an animal model characterized by the co-expression of activated forms of AKT and Ras in the mouse liver. We found that concomitant activation of AKT/mTOR and Ras/MAPK cascades leads to rapid liver tumor development in AKT/Ras mice, mainly through mTORC1 induction. To further define the role of mTORC1 cascade in AKT/Ras induced HCC development, the mTORC1 inhibitor Rapamycin was administered to AKT/Ras mice at the time when small tumors started to emerge in the liver. Of note, Rapamycin treatment significantly delayed hepatocarcinogenesis in AKT/Ras mice. However, some microscopic lesions persisted in the livers of AKT/Ras mice despite the treatment and rapidly gave rise to HCC following Rapamycin withdrawal. Mechanistically, Rapamycin inhibited mTORC1 and mTORC2 pathways, lipogenesis and glycolysis, resulting in inhibition of proliferation in the treated livers. However, activated ERK and its downstream effectors, Mnk1 and eIF4E, were strongly upregulated in the residual lesions. Concomitant suppression of AKT/mTOR and Ras/MAPK pathways was highly detrimental for the growth of AKT/Ras cells in vitro. The study indicates the existence of a complex interplay between AKT/mTOR and Ras/MAPK pathways during hepatocarcinogenesis, with important implications for the understanding of HCC pathogenesis as well as for its prevention and treatment.