【學術前沿】史岸冰/林瓏合作揭示LET-502/ROCK通過促進分選內吞體...

小GTP酶 (small GTPase)Rab5/RAB-5定位於早期內吞體，募集效應因子以調控包括內吞循環運輸在內的多類型生物學過程【1-3】，例如Rab5效應蛋白Rabenosyn-5與Rab4、膜塑性蛋白EHD1相互作用並促進循環運輸【1,4】。Rab5/RAB-5活性受到精密時空調控，對RAB-5活性的動態管理對於從早期內吞體至循環途徑的貨物輸送至關重要。在秀麗線蟲腸上皮細胞中，組成型活性RAB-5(Q78L)的過表達顯著損害循環運輸，而CED-10/Rac1通過募集TBC-2下調RAB-5活性【5】。後續研究還發現，RAB-10和BAR結構域蛋白AMPH-1/Amphiphysin也能夠募集TBC-2至內吞體，RAB-10或AMPH-1缺失導致內吞體膜上活性RAB-5增加，循環貨物滯留在早期內吞體【6】。

Rab活性受鳥嘌呤核苷酸交換因子(GEFs)和Rab GTPase激活蛋白(GAPs)調節【7】。結合GTP的活性態Rabs通常與不同內膜結構結合，與膜上效應蛋白相互作用以實現功能，而結合GDP的無活性態Rabs大多位於細胞質中【8】。作為重要分子開關，Rab5/RAB-5活性需要由相應的GEF和GAP嚴格管理，以保障正確的貨物分選運輸。TBC-2是人類TBC1D2A和TBC1D2B的線蟲同源物【9】。TBC1D2A具有Rab7-GAP活性，與LC3和LRRK1相互作用以調節自噬【10,11】，TBC1D2B與Rab22之間存在相互作用，但其細胞功能有待進一步闡明【12】。與哺乳動物同源物不同，TBC-2在內吞體上促進RAB-5失活，促進RAB-5到RAB-7的轉化【9】。秀麗線蟲基因組編碼三種潛在RAB-5-GEF蛋白：RIN-1，RABX-5和RME-6 【13-15】。RME-6駐留在網格蛋白包被囊泡中促進內吞【14】，而RABX-5定位於早期內吞體，與RAB-5-to-RAB-7轉化有關【13】。

ROCK最初被表徵為Rho效應因子，其actin相關功能在高爾基體結構維持、腎纖毛髮生和內皮完整性方面被廣泛報導【16-19】。LET-502是ROCK的線蟲同源物【20】，作為RHO-1/Rho效應因子參與P細胞遷移和胚胎形態發生【21-23】。2020年9月22日，華中科技大學基礎醫學院林瓏和史岸冰合作在Cell Reports在線發表了題為LET-502/ROCK regulatesendocytic recycling by promoting activation of RAB-5 in a distinctsubpopulation of sorting endosomes (LET-502/ROCK通過促進分選內吞體亞群上RAB-5活化調節內吞循環運輸) 的研究論文。

通過全基因組RNAi篩選，研究人員發現LET-502是一種新型循環調節因子。在腸上皮細胞中，LET-502和RAB-5在內吞體上共定位，LET-502缺失導致內吞體上的RAB-5標記水平下降。值得注意的是，LET-502僅與結合GDP的無活性態RAB-5相互作用，且LET-502促進RABX-5與無活性態RAB-5(S33N)間的相互作用，促進RABX-5的RAB-5-GEF活性，提示LET-502和RABX-5互作可促進RABX-5-VPS9結構域從其自抑制構象中釋放。研究人員進一步對TBC-2/RAB-5-GAP、CED-10/Rac1與LET-502之間的遺傳學關係進行了解析，發現功能衝突的LET-502和TBC-2同時缺失會導致RAB-5在內吞體上過度積累，暗示CED-10可能在時空水平調節RAB-5定位：在CED-10的總控下，TBC-2負調節RAB-5活性，然後RAB-5被LET-502-RABX-5模塊重新激活。受以上結果的啟發，研究人員分析了LET-502-RABX-5模塊和RME-6的功能位置，嘗試勾勒RAB-5活性管理的空間特徵。他們發現在缺失RABX-5/RAB-5-GEF或其輔助因子LET-502的細胞中，RAB-5主要殘留於細胞外圍的內吞體，而RME-6/RAB-5-GEF缺失導致細胞質深處RAB-5陽性結構的殘留，TBC-2過表達則廣泛減少RAB-5的內吞體定位，沒有空間差異性。這些結果表明RABX-5和RME-6的功能位置不同，分別影響RAB-5在兩個空間分布不同的內吞體亞群上的駐留。TBC-2介導細胞外圍分選內吞體上的RAB-5失活，此後LET-502在細胞質深處內吞體亞群上協助再次激活RAB-5，以維持正常的循環運輸進程。

基礎醫學院博士研究生張文娟為該論文的第一作者，林瓏副教授和史岸冰教授為共同通訊作者。

原文連結：

https://doi.org/10.1016/j.celrep.2020.108173

參考文獻

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原標題：《【學術前沿】史岸冰/林瓏合作揭示LET-502/ROCK通過促進分選內吞體亞群上RAB-5活化調節內吞循環運輸》

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