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Cell & JCI: 乳酰化修饰酶AARS1, the writer of lactylation

已有 1479 次阅读 2024-4-23 16:03 |系统分类:论文交流

     蛋白质乳酰化修饰(lactylation)自2019年被首次报道以来受到了广泛关注,作为一种新型蛋白质翻译后修饰(PTM)参与调控了多种生理和病理过程,为乳酸的非代谢功能研究提供了新的视角。p300等酰基转移酶被认为可以催化底物蛋白的赖氨酸残基发生乳酰化修饰;在该体系中,p300利用乳酰辅酶A(Lactyl-CoA)作为乳酰基团供体。然而,目前哺乳动物细胞中从乳酸到乳酰辅酶A的转化过程仍未明确,且肿瘤细胞中乳酰辅酶A浓度比乙酰辅酶A(Acetyl-CoA)低1000倍以上,比乳酸低100000倍以上,这可能大大限制了细胞中p300的乳酰基转移酶活性(即使有的话),也提出了细胞中是否存在直接利用乳酸作为乳酰基供体的乳酰化修饰酶这一科学问题。因此,明确哺乳动物细胞中的乳酰基转移酶及催化反应过程具有重要意义。

    2024年4月22日,苏州大学生物医学研究院周芳芳教授团队在Cell 在线发表题为“Alanyl-tRNA synthetase, AARS1, is a lactate sensor and lactyltransferase that lactylates p53 and contributes to tumorigenesis”的研究论文,发现AARS1作为乳酸传感器介导肿瘤细胞中的全局赖氨酸乳酰化。AARS1与乳酸结合并催化形成乳酸腺苷酸(lactate-AMP,Lac-AMP),随后乳酸转移到裂解酶受体残基。

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    蛋白质组学研究揭示了大量的AARS1靶点,包括p53,其中DNA结合域的赖氨酸120和赖氨酸139被乳酸化。携带本构乳酸化赖氨酸残基的p53变异体的产生和利用表明,p53的AARS1乳酸化阻碍了其液-液相分离、DNA结合和转录激活。在携带野生型p53的癌症患者中,AARS1表达和p53酰化与预后不良相关。在动物模型中,β-丙氨酸破坏乳酸与AARS1的结合,降低p53的乳酸化,并减轻肿瘤的发生。该研究认为AARS1通过将肿瘤细胞代谢与蛋白质组改变相结合而参与肿瘤发生。

    代谢异常是癌症的重要标志。即使有氧气存在,大多数癌细胞也对有氧糖酵解上瘾这种代谢改变,也被称为“Warburg效应”,启动微环境并导致糖酵解中间体(如乳酸)的升高,以支持细胞增殖。有证据表明,乳酸参与调节多种生物过程,如免疫细胞分化、肿瘤免疫监视、纤维化、缺血性损伤,并通过一种被称为乳酸化的表观遗传修饰调节基因表达。后者表明乳酸可以共价修饰蛋白质,但细胞内乳酸被检测并转化为乳酸化的机制尚不清楚。

    肿瘤抑制因子p53介导多种功能;作为一种转录因子,它通过激活许多控制DNA修复、细胞周期进程和应激下存活的靶标来保护细胞免受恶性转化。p53的活性在肿瘤发生过程中经常被抑制,超过一半的人类癌症含有p53突变,损害p53肿瘤抑制活性。p53的活性也受翻译后修饰的调控,如磷酸化、乙酰化和泛素化了解p53调控的机制可能会拓宽癌症患者的治疗策略,以提高p53的活性。

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Lac-AMP的形成AARS1能够响应细胞内的乳酸水平,并通过ATP依赖的方式激活乳酸,形成Lac-AMP。这一步骤类似于氨基酸tRNA合成酶在氨基酸激活过程中形成氨基酸腺苷酸(aminoacyl-AMP)。

    该研究进行了CRISPR筛选,发现AARS1是细胞内乳酸感应器,直接结合乳酸并催化全局赖氨酸乳酸化,包括p53。通过ATP依赖性乳酸- AMP的形成,AARS1将乳酸转移到赖氨酸残基共价结合,并介导蛋白质组范围赖氨酸乳酸基的形成。该研究发现大肠杆菌AlaRS (EcAlaRS)也能催化真核蛋白质组和p53的乳酸化,这表明AARS1的乳酸转移酶活性是一种古老而保守的功能。AARS1介导其DNA结合域(DBD)的位点特异性p53乳酸化,阻止p53与含有p53应答元件(p53RE-DNA)的DNA结合,从而抑制p53液-液相分离(LLPS)和转录。这项工作还提供了一种利用β-丙氨酸阻止p53乳酸化的方法,β-丙氨酸与乳酸竞争结合AARS1,从而加强癌症化疗。总之,这些研究拓展了蛋白质乳酸化急需的基础知识,揭示了代谢物乳酸与肿瘤发生和发展之间的直接相互作用。

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 Highlights

•Tumor-derived lactate is a natural inhibitor of the tumor suppressor p53

•AARS1 is an intracellular lactate sensor that mediates global lysine lactylation

•AARS1 lactylation of p53 at the K120 and K139 residues impairs p53 LLPS and DNA binding

•β-alanine inhibits lactate binding to AARS1, global lacylation, and tumorigenesis

摘要:Summary    Lysine lactylation is a post-translational modification that links cellular metabolism to protein function. Here, we find that AARS1 functions as a lactate sensor that mediates global lysine lacylation in tumor cells. AARS1 binds to lactate and catalyzes the formation of lactate-AMP, followed by transfer of lactate to the lysince acceptor residue. Proteomics studies reveal a large number of AARS1 targets, including p53 where lysine 120 and lysine 139 in the DNA binding domain are lactylated. Generation and utilization of p53 variants carrying constitutively lactylated lysine residues revealed that AARS1 lactylation of p53 hinders its liquid-liquid phase separation, DNA binding, and transcriptional activation. AARS1 expression and p53 lacylation correlate with poor prognosis among cancer patients carrying wild type p53. β-alanine disrupts lactate binding to AARS1, reduces p53 lacylation, and mitigates tumorigenesis in animal models. We propose that AARS1 contributes to tumorigenesis by coupling tumor cell metabolism to proteome alteration.

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    近日,复旦大学生命科学学院周兆才、焦石赵世民教授合作在国际学术期刊Journal of Clinical Investigation发表题为"The alanyl-tRNA synthetase AARS1 moonlights as a lacty-transferase to promote YAP signaling in gastric cancer"的文章,也发现AARS1在其经典的丙氨酰tRNA合成酶功能之外,还可以兼职作为一种乳酰基转移酶,直接利用乳酸作为乳酰基供体,催化Hippo信号通路关键组分YAP和TEAD发生乳酰化修饰,激活下游基因转录表达,促进胃癌细胞恶性增殖。 

    由于乳酸丙氨酸的三维结构高度相似,研究者推测乳酸可以被丙氨酰tRNA合成酶识别并转移到蛋白质赖氨酸残基上。随后,研究者利用异源表达纯化的蛋白构建了纯体外乳酰化修饰体系,并结合一系列体内实验,证实了AARS1的催化中心能够直接识别结合乳酸,在ATP存在的条件下将乳酸转化为高能中间分子(Lac-AMP),从而催化底物蛋白发生乳酰化修饰。

JCI1.jpg

    为了进一步研究AARS1作为乳酰基转移酶的功能,研究者通过乳酰化修饰组学在胃癌细胞中鉴定到1182个蛋白的2789个乳酰化修饰位点。其中,Hippo信号通路核心组分YAP和TEAD存在保守的乳酰化修饰位点。同时,研究者发现AARS1能够响应细胞内乳酸水平升高而进入细胞核,从而催化YAP和TEAD发生乳酰化修饰,增强其转录活性,促进胃癌细胞增殖。此外,AARS1还被发现是Hippo信号通路下游的一个直接靶基因,从而与YAP-TEAD乳酰化修饰形成一个正反馈调控环路。

JCI2.jpg

    总之,乳酸辅酶A(lactyl-CoA)的浓度相对于乳酸(lactate)和乙酰辅酶A(acetyl-CoA)是非常低的,这可能限制了它在细胞内作为乳酸化反应中供体的可行性。相反,AARS1能够直接利用细胞内更丰富的乳酸和ATP来进行蛋白质乳酸化。丙氨酰tRNA合成酶(AARS1感应细胞内乳酸水平,作为乳酰基转移酶的非经典功能;其完全不需要乳酰辅酶A,而是直接利用乳酸作为乳酰基供体,催化YAP-TEAD等蛋白质发生乳酰化修饰的功能和机制。AARS1这一非经典功能将肿瘤细胞内的高水平乳酸与YAP驱动的恶性增殖信号联系起来,为深入理解肿瘤细胞的Warburg效应提供了新的视角。

JCI888.jpg

摘要:Lactylation has been recently identified as a new type of posttranslational modification widely occurring on lysine residues of both histone and non-histone proteins. The acetyl transferase p300 is thought to mediate protein lactylation, yet the cellular concentration of the proposed lactyl-donor, lactyl-coenzyme A is about 1,000 times lower than that of acetyl-CoA, raising the question whether p300 is a genuine lactyl-transferase. Here, we report the Alanyl-tRNA synthetase 1 (AARS1) moonlights as a bona fide lactyl-transferase that directly uses lactate and ATP to catalyze protein lactylation. Among the candidate substrates, we focused on the Hippo pathway that has a well-established role in tumorigenesis. Specifically, AARS1 was found to sense intracellular lactate and translocate into the nucleus to lactylate and activate YAP-TEAD complex; and AARS1 itself was identified as a Hippo target gene that forms a positive feedback loop with YAP-TEAD to promote gastric cancer (GC) cell proliferation. Consistently, the expression of AARS1 was found to be upregulated in GC, and elevated AARS1 expression was found to be associated with poor prognosis for GC patients. Collectively, this work discovered AARS1 with lactyl-transferase activity in vitro and in vivo and revealed how the metabolite lactate is translated into a signal of cell proliferation.

参考文献:

1. Zhang, D. et al. Metabolic regulation of gene expression by histone lactylation. Nature  (2019) 574, 575-580, doi:10.1038/s41586-019-1678-1.

2. Zhi Zong et al. Alanyl-tRNA synthetase, AARS1, is a lactate sensor and lactyltransferase that lactylates p53 and contributes to tumorigenesis. Cell (2024) DOI: https://doi.org/10.1016/j.cell.2024.04.002

3.  TJunyi Ju et al. he alanyl-tRNA synthetase AARS1 moonlights as a lacty-transferase to promote YAP signaling in gastric cancer. JCI (2024) https://www.jci.org/articles/view/174587 



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