Endogenous Rab29 does not impact basal or stimulated LRRK2 pathway activity.
Endogenous Rab29 does not impact basal or stimulated LRRK2 pathway activity.
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内源性Rab29不影响基础或刺激的LRRK2途径活性。
DOI：
10.1042/bcj20200458
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发表时间：
2020-11-27
期刊：
影响因子：
--
通讯作者：
Alessi DR
中科院分区：
文献类型：
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作者：
Kalogeropulou AF;Freemantle JB;Lis P;Vides EG;Polinski NK;Alessi DR
增强LRRK 2蛋白激酶活性的突变导致遗传性帕金森病。LRRK 2磷酸化效应子结合开关-II基序内的一组Rab GT3蛋白，包括Rab 10和Rab 12。先前的工作表明，PARK 16基因座，其中含有Rab 29的基因编码，参与帕金森氏症，Rab 29在一个共同的途径与LRRK 2。Rab 29和LRRK 2的共表达通过将LRRK 2募集到反式高尔基体网络的表面来刺激LRRK 2活性。在这里，我们报告说，敲除Rab 29不影响内源性LRRK 2活性，基于Rab 10和Rab 12磷酸化的评估，在野生型LRRK 2，LRRK 2 [R1441 C]或VPS 35 [D 620 N]敲入小鼠组织和原代细胞系，包括脑提取物和胚胎成纤维细胞。我们发现，在脑提取物中，Rab 12磷酸化比Rab 10磷酸化更受LRRK 2抑制剂和致病突变的影响。小鼠模型中Rab 29的转基因过表达也不足以刺激基础LRRK 2活性。我们观察到，刺激Rab 10和Rab 12磷酸化诱导的代理人，强调内溶酶体系统（尼日利亚菌素，莫能菌素，氯喹和LLOMe）被LRRK 2抑制剂抑制，但不阻断Rab 29缺陷细胞。在测试的药物中，尼日利亚菌素诱导Rab 10和Rab 12磷酸化的增加最大（5至9倍）。我们的研究结果表明，基础的，致病的，以及尼日利亚菌素和莫能菌素刺激LRRK 2途径的活性是不控制Rab 29。需要进一步的工作来确定LRRK 2活性是如何调节的，以及其他Rab蛋白是否可以通过将其靶向不同的膜来控制LRRK 2。
Mutations that enhance LRRK2 protein kinase activity cause inherited Parkinson's disease. LRRK2 phosphorylates a group of Rab GTPase proteins, including Rab10 and Rab12, within the effector-binding switch-II motif. Previous work has indicated that the PARK16 locus, which harbors the gene encoding for Rab29, is involved in Parkinson's, and that Rab29 operates in a common pathway with LRRK2. Co-expression of Rab29 and LRRK2 stimulates LRRK2 activity by recruiting LRRK2 to the surface of the trans Golgi network. Here, we report that knock-out of Rab29 does not influence endogenous LRRK2 activity, based on the assessment of Rab10 and Rab12 phosphorylation, in wild-type LRRK2, LRRK2[R1441C] or VPS35[D620N] knock-in mouse tissues and primary cell lines, including brain extracts and embryonic fibroblasts. We find that in brain extracts, Rab12 phosphorylation is more robustly impacted by LRRK2 inhibitors and pathogenic mutations than Rab10 phosphorylation. Transgenic overexpression of Rab29 in a mouse model was also insufficient to stimulate basal LRRK2 activity. We observed that stimulation of Rab10 and Rab12 phosphorylation induced by agents that stress the endolysosomal system (nigericin, monensin, chloroquine and LLOMe) is suppressed by LRRK2 inhibitors but not blocked in Rab29 deficient cells. From the agents tested, nigericin induced the greatest increase in Rab10 and Rab12 phosphorylation (5 to 9-fold). Our findings indicate that basal, pathogenic, as well as nigericin and monensin stimulated LRRK2 pathway activity is not controlled by Rab29. Further work is required to establish how LRRK2 activity is regulated, and whether other Rab proteins can control LRRK2 by targeting it to diverse membranes.