Division of Gastroenterology and Liver Diseases, USC Research Center for Liver Diseases, Los Angeles, CA, USA.
Hepatology. 2013 Jun;57(6):2299-313. doi: 10.1002/hep.26258. Epub 2013 May 14.
Methionine adenosyltransferase 2B (MAT2B) encodes for two variant proteins (V1 and V2) that promote cell growth. Using in-solution proteomics, GIT1 (G Protein Coupled Receptor Kinase Interacting ArfGAP 1) was identified as a potential interacting partner of MAT2B. Here, we examined the functional significance of this interplay. Coimmunoprecipitation experiments examined protein interactions. Tissue expression levels of proteins were examined using immunohistochemistry and western blotting. Expression levels of proteins were varied using transient knockdown or overexpression to observe the effect of alterations in each protein on the entire complex. Direct interaction among individual proteins was further verified using in vitro translated and recombinant proteins. We found both MAT2B variants interact with GIT1. Overexpression of V1, V2, or GIT1 activated mitogen-activated protein kinase kinase 1 (MEK1) and extracellular signal-regulated kinase (ERK), raised cyclin D1 protein level, and increased growth, whereas the opposite occurred when V1, V2, or GIT1 was knocked down. MAT2B and GIT1 require each other to activate MEK1/ERK and increase growth. MAT2B directly interacts with MEK1, GIT1, and ERK2. Expression level of V1, V2, or GIT1 directly influenced recruitment of GIT1 or MAT2B and ERK2 to MEK1, respectively. In pull-down assays, MAT2B directly promoted binding of GIT1 and ERK2 to MEK1. MAT2B and GIT1 interact and are overexpressed in most human liver and colon cancer specimens. Increased expression of V1, V2, or GIT1 promoted growth in an orthotopic liver cancer model, whereas increased expression of either V1 or V2 with GIT1 further enhanced growth and lung metastasis.
MAT2B and GIT1 form a scaffold, which recruits and activates MEK and ERK to promote growth and tumorigenesis. This novel MAT2B/GIT1 complex may provide a potential therapeutic gateway in human liver and colon cancer. (HEPATOLOGY 2012).
甲硫氨酸腺苷基转移酶 2B（MAT2B）编码两种变体蛋白（V1 和 V2），可促进细胞生长。通过溶液蛋白质组学，GIT1（G 蛋白偶联受体激酶相互作用的 ArfGAP1）被鉴定为 MAT2B 的潜在相互作用伙伴。在这里，我们研究了这种相互作用的功能意义。免疫沉淀实验检测蛋白相互作用。使用免疫组织化学和蛋白质印迹法检测蛋白的组织表达水平。使用瞬时敲低或过表达来观察每个蛋白的变化对整个复合物的影响，从而改变蛋白的表达水平。使用体外翻译和重组蛋白进一步验证了单个蛋白之间的直接相互作用。我们发现两种 MAT2B 变体都与 GIT1 相互作用。V1、V2 或 GIT1 的过表达激活丝裂原活化蛋白激酶激酶 1（MEK1）和细胞外信号调节激酶（ERK），提高 cyclin D1 蛋白水平并促进生长，而当 V1、V2 或 GIT1 被敲低时则相反。MAT2B 和 GIT1 相互作用，需要彼此来激活 MEK1/ERK 并增加生长。MAT2B 直接与 MEK1、GIT1 和 ERK2 相互作用。V1、V2 或 GIT1 的表达水平直接影响 GIT1 或 MAT2B 和 ERK2 分别募集到 MEK1 的情况。在下拉实验中，MAT2B 直接促进 GIT1 和 ERK2 与 MEK1 的结合。MAT2B 和 GIT1 在大多数人类肝癌和结肠癌标本中相互作用和过表达。V1、V2 或 GIT1 的表达增加促进了原位肝癌模型中的生长，而 V1 或 V2 与 GIT1 一起表达增加进一步增强了生长和肺转移。
MAT2B 和 GIT1 形成一个支架，募集并激活 MEK 和 ERK 以促进生长和肿瘤发生。这种新型 MAT2B/GIT1 复合物可能为人类肝癌和结肠癌提供潜在的治疗途径。（HEPATOLOGY 2012）。
