甲硫氨酸腺苷转移酶1A/2A平衡与肝细胞癌

肝细胞癌是一种死亡率极高的癌症,大多数病人发现时已属晚期.甲硫氨酸腺苷转移酶(MAT)是细胞生命活动的关键酶,可以通过催化甲酼氨酸和三磷酸腺苷(ATP)结合,促进生物甲基供体S-腺苷甲酼氨酸(SAMe)的生物合成.正常肝细胞中MAT1A与MAT2A存在动态平衡,共同维持细胞内SAMe稳态;肝细胞癌中MAT1A转变成MAT2A,会使SAMe生物合成减少,为癌细胞生长提供有利条件,故MAT1A表达降低而MAT2A增高.因此,促进MAT2A向MAT1A转化,进而提高MAT1A/MAT2A的比值可能成为治疗肝细胞癌的关键靶点之一.本文就MAT1A/MAT2A平衡在肝细胞癌中的重要作用作一综述,旨在为寻找肝细胞癌防治靶点提供新的思路.     

氨基酸代谢重编程在肿瘤发生发展中的作用

肿瘤的发生发展不仅取决于基因的突变或缺失，还随着肿瘤细胞的代谢重塑或异常改变而发生改变。在营养缺乏的条件下，肿瘤细胞的代谢重编程赋予癌细胞快速增殖的能力。其中，氨基酸代谢重编程是肿瘤代谢异常改变的重要特征之一。研究发现，氨基酸不仅能够作为氮供体为肿瘤细胞的增殖、侵袭和免疫逃逸过程提供核苷酸等生物大分子的合成原料，而且还是肿瘤微环境中免疫细胞活化和发挥抗肿瘤作用的重要代谢物质。氨基酸代谢的异常改变与肿瘤的发生发展和肿瘤免疫密切相关，其代谢途径中的部分关键蛋白质或关键酶可作为肿瘤诊断和治疗的生物标志物。因此，本文围绕氨基酸转运体对癌细胞增殖的影响和肿瘤代谢循环过程中的谷氨酰胺、天冬酰胺、丝氨酸和甘氨酸等氨基酸代谢的异常改变进行总结，介绍了氨基酸代谢与肿瘤细胞mTOR信号通路、肿瘤微环境和免疫细胞功能的相关性，对靶向氨基酸代谢的肿瘤治疗药物进行了分析和展望。期望该工作为深入了解氨基酸代谢对肿瘤发生发展的调控及其可能存在的肿瘤治疗靶点提供有用的参考。     

腺苷甲硫氨酸合成酶的基因及结构研究进展

腺苷甲硫氨酸合成酶催化ATP和L-甲硫氨酸合成腺苷甲硫氨酸,在不同生物体和不同组织中腺苷甲硫氨酸合成酶的存在形式和编码酶的基因都有差别,本文综述了不同生物的腺苷甲硫氨酸合成酶的基因、酶结构、酶反应动力学及应用前景。     

多胺生物合成途径中两个关键酶基因研究进展

多胺是一类小分子生物活性物质,广泛存在于生物体内,与植物的生长发育、衰老及抗逆性都有着密切的联系。就多胺合成途径中的两个关键酶基因,即S-腺苷甲硫氨酸合成酶基因(SAMS)和S-腺苷甲硫氨酸脱羧酶基因(SAMDC)的克隆、表达,以及转S-腺苷甲硫氨酸合成酶基因(SAMS)和转S-腺苷甲硫氨酸脱羧酶基因(SAMDC)表达调控等方面的研究进行回顾总结,并对其应用前景进行展望。     

硫腺苷甲硫氨酸合成酶基因的克隆及其在盐胁迫条件下的不同表达

硫腺苷甲硫氨酸作为甲基供体在转甲基反应中起到重要作用.为了解硫腺苷甲硫氨酸在盐地碱蓬(Suaedasalsa (L.)Pall)耐盐中的作用,我们对可能编码硫腺苷甲硫氨酸合成酶的基因(SsSAMS2)进行了分析.该基因在经400 mmol/L NaCl处理的盐地碱蓬地上部分的λ-Zap cDNA文库中克隆到,其插入片段全长1 531 bp,包含一个395个氨基酸的开放阅读框架,该基因推断的分子量约为43 kD.SsSAMS2与长春花(Catharanthus roseus)的SAMS2在氨基酸水平上的一致性为93%.Southern杂交显示,SsSAMS2在盐地碱蓬基因组中可能是两个拷贝.Northern分析显示硫腺苷甲硫氨酸合成酶基因受NaCl等胁迫的正调控.酶活性检测表明,NaCl胁迫条件下该酶活性增强.     

微生物源甲硫氨酸γ-裂解酶的研究进展

甲硫氨酸γ-裂解酶(methionineγ-lyase,MGL)催化甲硫氨酸γ位C-S键的裂解反应,生成等摩尔的α-酮丁酸、甲基硫醇和氨。MGL降低胞内甲硫氨酸浓度,显著抑制恶性肿瘤细胞的生长和迁移,激发正常细胞的抗氧化反应,开发高效的MGL已成为肿瘤治疗和抗衰老研究的热点。MGL广泛存在于微生物中,而在哺乳动物中不存在,MGL是开发抗致病微生物感染药物的重要靶标。产物甲基硫醇及其衍生物是构成食品香味的主体成分,其组分和浓度决定了食品整体香味的形成,系统阐明MGL的催化机制和活性调节机制将推动食品品质及其稳定性的精准控制。本文总结了微生物源MGL的挖掘、催化机制和改造方面的最新进展,讨论了MGL在癌症治疗、抗衰老、抗致病微生物感染以及食品香味合成和制造领域的应用情况,展望了MGL的发展前景与挑战。     

硫腺苷甲硫氨酸合成酶基因的克隆及其在盐胁迫条件下的不同表达

硫腺苷甲硫氨酸作为甲基供体在转甲基反应中起到重要作用。为了解硫腺苷甲硫氨酸在盐地碱蓬(Suaeda salsa (L.) Pall)耐盐中的作用,我们对可能编码硫腺苷甲硫氨酸合成酶的基因(SsSAMS2)进行了分析.该基因在经400mmol/L NaCl处理的盐地碱蓬地上部分的λ-Zap cDNA文库中克隆到,其播入片段全长1531bp,包含一个395个氨基酸的开放阅读框架,该基因推断的分子量约为43kD.SsSAMS2与长春花(Catharanthus roseus)的SAMS2在氨基酸水平上的一致性为93%.Southern杂交显示,SsSAMS2在盐地碱蓬基因组中可能是两个拷贝.Northern分析显示硫腺苷甲硫氨酸合成酶基因受NaCl等胁迫的正调控.酶活性检测表明,NaCl胁迫条件下该酶活性增强.     

肿瘤细胞代谢异常及其调控机制

肿瘤细胞的异常代谢已成为肿瘤研究领域的共识.肿瘤细胞代谢重编程的发生是为了维持在恶劣微环境中的存活和无限增殖.因此,肿瘤细胞异常的代谢通路、代谢调控蛋白及代谢酶可能是肿瘤治疗的关键靶点.本文旨在介绍近年来肿瘤代谢的研究进展及肿瘤代谢研究对癌症治疗的意义.     

强化表达SAM合成酶促进SAM在毕赤酵母中累积

S 腺苷甲硫氨酸 (S adenosyl L methionine ,SAM)是生物体硫代谢的重要中间代谢物质 ,在体内起着转甲基、转硫基、转氨丙基的作用 ,具有重要的药用和保健价值。将酿酒酵母来源的SAM合成酶 2基因置于GAP启动子调控下 ,构建胞内组成型表达质粒 ,并电转化至毕赤酵母菌株GS115。经Zeocin抗性和培养筛选到一株高产SAM的重组菌。对重组菌表达工艺的研究表明 ,碳源、氮源、pH和溶解氧对SAM的累积有较大影响。在优化条件下 ,重组细胞培养 3天 ,SAM累积量可达 2 .49g/L。     

Mondo蛋白在肿瘤相关代谢中的研究进展

转录因子Mondo蛋白家族包括MondoA和ChREBP(MondoB)两个家族成员,是葡萄糖介导的基因转录调控的关键调控因子,可直接调控糖酵解和脂肪酸生成相关基因的表达,在细胞代谢与能量平衡中发挥重要作用。细胞代谢改变是肿瘤的重要特征之一,为肿瘤细胞生长及恶性进展创造了有利条件。近年来,研究发现,Mondo蛋白在肿瘤细胞糖酵解、脂肪酸合成和谷氨酰胺利用等代谢通路中发挥着重要作用,而且Mondo蛋白调控肿瘤细胞的代谢,其在肿瘤细胞生长、增殖和侵袭等过程中的作用值得肿瘤研究者关注。因此,更好地认识Mondo蛋白调控肿瘤细胞代谢的机制,将为癌症的治疗提供新的方向。本文对Mondo蛋白家族成员的分子特征、表达调控、组织特异性功能及其在肿瘤代谢重编程和细胞增殖中的最新研究进行综述,为肿瘤的防治提供新思路。     

Methylthioadenosine phosphorylase regulates ornithine decarboxylase by production of downstream metabolites

The gene encoding methylthioadenosine phosphorylase (MTAP), the initial enzyme in the methionine salvage pathway, is deleted in a variety of human tumors and acts as a tumor suppressor gene in cell culture (Christopher, S. A., Diegelman, P., Porter, C. W., and Kruger, W. D. (2002) Cancer Res. 62, 6639-6644). Overexpression of the polyamine biosynthetic enzyme ornithine decarboxylase (ODC) is frequently observed in tumors and has been shown to be tumorigenic in vitro and in vivo. In this paper, we demonstrate a novel regulatory pathway in which the methionine salvage pathway products inhibit ODC activity. We show that in Saccharomyces cerevisiae the MEU1 gene encodes MTAP and that Meu1delta cells have an 8-fold increase in ODC activity, resulting in large elevations in polyamine pools. Mutations in putative salvage pathway genes downstream of MTAP also cause elevated ODC activity and elevated polyamines. The addition of the penultimate salvage pathway compound 4-methylthio-2-oxobutanoic acid represses ODC levels in both MTAP-deleted yeast and human tumor cell lines, indicating that 4-methylthio-2-oxobutanoic acid acts as a negative regulator of polyamine biosynthesis. Expression of MTAP in MTAP-deleted MCF-7 breast adenocarcinoma cells results in a significant reduction of ODC activity and reduction in polyamine levels. Taken together, our results show that products of the methionine salvage pathway regulate polyamine biosynthesis and suggest that MTAP deletion may lead to ODC activation in human tumors.     

Molecular cloning of the human methylthioadenosine phosphorylase processed pseudogene and localization to 3q28

Human methylthioadenosine phosphorylase (MTAP) is a purine and methionine metabolic enzyme present ubiquitously in all normal tissues, but often deleted in many types of cancer. The gene for this enzyme maps to chromosome 9 at band p21 where the cyclin-dependent kinase inhibitor genes for p16 and p15 also reside. During our efforts to clone this gene we also isolated a phage clone containing a processed pseudogene of MTAP. The sequence is 92% homologous to the MTAP cDNA, is flanked at its 3′ end by a repetitive element, but does not possess a poly(A) stretch. We localized this processed pseudogene to band 28 on the long arm of chromosome 3 by fluorescence in situ hybridization. All 22 malignant cell lines with deletions at 9p21 screened possessed the pseudogene.     

Expression and Function of Methylthioadenosine Phosphorylase in Chronic Liver Disease

To study expression and function of methylthioadenosine phosphorylase (MTAP), the rate-limiting enzyme in the methionine and adenine salvage pathway, in chronic liver disease.

Design

MTAP expression was analyzed by qRT-PCR, Western blot and immunohistochemical analysis. Levels of MTA were determined by liquid chromatography-tandem mass spectrometry.

Results

MTAP was downregulated in hepatocytes in murine fibrosis models and in patients with chronic liver disease, leading to a concomitant increase in MTA levels. In contrast, activated hepatic stellate cells (HSCs) showed strong MTAP expression in cirrhotic livers. However, also MTA levels in activated HSCs were significantly higher than in hepatocytes, and there was a significant correlation between MTA levels and collagen expression in diseased human liver tissue indicating that activated HSCs significantly contribute to elevated MTA in diseased livers. MTAP suppression by siRNA resulted in increased MTA levels, NFκB activation and apoptosis resistance, while overexpression of MTAP caused the opposite effects in HSCs. The anti-apoptotic effect of low MTAP expression and high MTA levels, respectively, was mediated by induced expression of survivin, while inhibition of survivin abolished the anti-apoptotic effect of MTA on HSCs. Treatment with a DNA demethylating agent induced MTAP and reduced survivin expression, while oxidative stress reduced MTAP levels but enhanced survivin expression in HSCs.

Conclusion

MTAP mediated regulation of MTA links polyamine metabolism with NFκB activation and apoptosis in HSCs. MTAP and MTAP modulating mechanisms appear as promising prognostic markers and therapeutic targets for hepatic fibrosis.     

Methylthioadenosine phosphorylase gene expression is impaired in human liver cirrhosis and hepatocarcinoma

Methylthioadenosine phosphorylase (MTAP) is a key enzyme in the methionine and adenine salvage pathways. In mammals, the liver plays a central role in methionine metabolism, and this essential function is lost in the progression from liver cirrhosis to hepatocarcinoma. Deficient MTAP gene expression has been recognized in many transformed cell lines and tissues. In the present work, we have studied the expression of MTAP in human and experimental liver cirrhosis and hepatocarcinoma. We observe that MTAP gene expression is significantly reduced in human hepatocarcinoma tissues and cell lines. Interestingly, MTAP gene expression was also impaired in the liver of CCl4-cirrhotic rats and cirrhotic patients. We provide evidence indicating that epigenetic mechanisms, involving DNA methylation and histone deacetylation, may play a role in the silencing of MTAP gene expression in hepatocarcinoma. Given the recently proposed tumor suppressor activity of MTAP, our observations can be relevant to the elucidation of the molecular mechanisms of multistep hepatocarcinogenesis.     

Growth and metastases of human lung cancer are inhibited in mouse xenografts by a transition state analogue of 5'-methylthioadenosine phosphorylase

The S-adenosylmethionine (AdoMet) salvage enzyme 5'-methylthioadenosine phosphorylase (MTAP) has been implicated as both a cancer target and a tumor suppressor. We tested these hypotheses in mouse xenografts of human lung cancers. AdoMet recycling from 5'-methylthioadenosine (MTA) was blocked by inhibition of MTAP with methylthio-DADMe-Immucillin-A (MTDIA), an orally available, nontoxic, picomolar transition state analogue. Blood, urine, and tumor levels of MTA increased in response to MTDIA treatment. MTDIA treatment inhibited A549 (human non-small cell lung carcinoma) and H358 (human bronchioloalveolar non-small cell lung carcinoma cells) xenograft tumor growth in immunodeficient Rag2(-/-)γC(-/-) and NCr-nu mice. Systemic MTA accumulation is implicated as the tumor-suppressive metabolite because MTDIA is effective for in vivo treatment of A549 MTAP(-/-) and H358 MTAP(+/+) tumors. Tumors from treated mice showed increased MTA and decreased polyamines but little alteration in AdoMet, methionine, or adenine levels. Gene expression profiles of A549 tumors from treated and untreated mice revealed only modest alterations with 62 up-regulated and 63 down-regulated mRNAs (≥ 3-fold). MTDIA antitumor activity in xenografts supports MTAP as a target for lung cancer therapy.     

Methionine metabolism is essential for SIRT1‐regulated mouse embryonic stem cell maintenance and embryonic development

Methionine metabolism is critical for epigenetic maintenance, redox homeostasis, and animal development. However, the regulation of methionine metabolism remains unclear. Here, we provide evidence that SIRT1, the most conserved mammalian NAD⁺‐dependent protein deacetylase, is critically involved in modulating methionine metabolism, thereby impacting maintenance of mouse embryonic stem cells (mESCs) and subsequent embryogenesis. We demonstrate that SIRT1‐deficient mESCs are hypersensitive to methionine restriction/depletion‐induced differentiation and apoptosis, primarily due to a reduced conversion of methionine to S‐adenosylmethionine. This reduction markedly decreases methylation levels of histones, resulting in dramatic alterations in gene expression profiles. Mechanistically, we discover that the enzyme converting methionine to S‐adenosylmethionine in mESCs, methionine adenosyltransferase 2a (MAT2a), is under control of Myc and SIRT1. Consistently, SIRT1 KO embryos display reduced Mat2a expression and histone methylation and are sensitive to maternal methionine restriction‐induced lethality, whereas maternal methionine supplementation increases the survival of SIRT1 KO newborn mice. Our findings uncover a novel regulatory mechanism for methionine metabolism and highlight the importance of methionine metabolism in SIRT1‐mediated mESC maintenance and embryonic development.     

PRMT5 silencing selectively affects MTAP-deleted mesothelioma: In vitro evidence of a novel promising approach

Malignant mesothelioma (MM) is an aggressive asbestos-related cancer of the serous membranes. Despite intensive treatment regimens, MM is still a fatal disease, mainly due to the intrinsic resistance to current therapies and the lack of predictive markers and new valuable molecular targets. Protein arginine methyltransferase 5 (PRMT5) inhibition has recently emerged as a potential therapy against methylthioadenosine phosphorylase (MTAP)-deficient cancers, in which the accumulation of the substrate 5'-methylthioadenosine (MTA) inhibits PRMT5 activity, thus sensitizing the cells to further PRMT5 inhibition. Considering that the MTAP gene is frequently codeleted with the adjacent cyclin-dependent kinase inhibitor 2A (CDKN2A) locus in MM, we assessed whether PRMT5 could represent a therapeutic target also for this cancer type. We evaluated PRMT5 expression, the MTAP status and MTA content in normal mesothelial and MM cell lines. We found that both administration of exogenous MTA and stable PRMT5 knock-down, by short hairpin RNAs (shRNAs), selectively reduced the growth of MTAP-deleted MM cells. We also observed that PRMT5 knock-down in MTAP-deficient MM cells reduced the expression of E2F1 target genes involved in cell cycle progression and of factors implicated in epithelial-to-mesenchymal transition. Therefore, PRMT5 targeting could represent a promising new therapeutic strategy against MTAP-deleted MMs.