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Liprin Phosphorylation Regulates Binding to LAR: Evidence for Liprin Autophosphorylation

机译：Liprin磷酸化调节与LAR的结合：Liprin自身磷酸化的证据

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摘要

The LAR transmembrane tyrosine phosphatase associates with liprin-alpha proteins and colocalizes with liprin-alpha l at focal adhesions.LAR has been implicated in axon guidance,and liprins are involved in synapse formation and synapse protein trafficking.Several liprin mutants have weaker binding to LAR as assessed by yeast interaction trap assays,and the extents of in vitro and in vivo phosphorylation of these mutants were reduced relative to that of wild-type liprin-alpha l.Treatment of liprin-alpha l with calf intestinal phosphatase weakened its interaction with the recombinant GST-LAR protein.A liprin LH region mutant that inhibited liprin phosphorylation did not bind to LAR as assessed by coprecipitation studies.Endogenous LAR was shown to bind phosphorylated liprin-alpha l from MDA-486 cells labeled in vivo with [~(32)P]orthophosphate.In further characterizing the phosphorylation of liprin,we found immunoprecipitates of liprin-alpha l expressed in COS-7 cells to incorporate phosphate after washes of up to 4 M NaCl.Additionally,purified liprin-alpha l derived from Sf-9 insect cells retained the ability to incorporate phosphate in in vitro phosphorylation assays,and a liprin-alpha l truncation mutant incorporated phosphate after denaturation and/or renaturation in SDS gels.Finally,binding assays showed that liprin binds to ATP- agarose and that the interaction is challenged by free ATP,but not by free GTP.Moreover,liprin LH region mutations that inhibit liprin phosphorylation stabilized the association of liprin with ATP-agarose.Taken together,our results suggest that liprin autophosphorylation regulates its association with LAR.
机译：Liprin-α蛋白的大型跨膜酪氨酸磷酸酶与乳头蛋白-α1的局灶性粘连结合.LAR在轴突引导中涉及，脂利民涉及Synapse地层和Synapse蛋白贩运。脂蛋白突变体具有较弱的结合如酵母相互作用陷阱测定的评估，相对于野生型Liprin-α1的那种突变体的体外磷酸化和体内磷酸化的因子相对于脂素-α1进行减少，具有小牛肠道磷酸酶与其相互作用重组的GST-LAR蛋白。抑制脂素磷酸化的脂素LH区域突变体与共沉淀研究的评估没有结合大致结合。显示出从体内标记的MDA-486细胞中结合磷酸化的Liprin-α1与[〜（32 ）p]正向磷酸盐。进一步表征脂培素的磷酸化，我们发现在COS-7细胞中表达的Liprin-αL的免疫沉淀物掺入Phosp讨厌在洗涤后高达4米NaCl.additionally，衍生自SF-9昆虫细胞的纯化的Liprin-α1保留了将磷酸盐掺入体外磷酸化测定中的能力，并且在变性和/或者在SDS凝胶中复发。最后，结合测定表明，脂蛋白与ATP-琼脂糖结合，并且相互作用被游离ATP攻击，但不是通过游离的GTP。抑制脂素磷酸化的脂素LH区域突变稳定脂培养物atp-agarose.taken在一起，我们的结果表明Liprin自动磷酸化调节其与Lar的关系。

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  • 来源
    《Biochemistry》 |2005年第48期|共10页
  • 作者

    Carles Sena-Pages; Michel Streuli; Quintus G.Medley;

  • 作者单位

    Department of Cancer Immunology and AIDS Dana-Farber Cancer Institute Boston Massachusetts 02115 and Departments of Pathology and Medicine Harvard Medical School Boston Massachusetts 02115;

    Department of Cancer Immunology and AIDS Dana-Farber Cancer Institute Boston Massachusetts 02115 and Departments of Pathology and Medicine Harvard Medical School Boston Massachusetts 02115;

    Department of Cancer Immunology and AIDS Dana-Farber Cancer Institute Boston Massachusetts 02115 and Departments of Pathology and Medicine Harvard Medical School Boston Massachusetts 02115;

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  • 正文语种 eng
  • 中图分类 生物化学;
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