Department of Biochemistry, University of Toronto, Toronto, ON M5G 1M1, Canada.
Department of Biochemistry, University of Toronto, Toronto, ON M5G 1M1, Canada; Department of Chemistry and Biochemistry, University of Regina, Regina, SK S4S 0A2, Canada.
Cell Rep. 2024 Feb 27;43(2):113713. doi: 10.1016/j.celrep.2024.113713. Epub 2024 Feb 1.
R2TP is a chaperone complex consisting of the AAA+ ATPases RUVBL1 and RUVBL2, as well as RPAP3 and PIH1D1 proteins. R2TP is responsible for the assembly of macromolecular complexes mainly acting through different adaptors. Using proximity-labeling mass spectrometry, we identified deleted in primary ciliary dyskinesia (DPCD) as an adaptor of R2TP. Here, we demonstrate that R2TP-DPCD influences ciliogenesis initiation through a unique mechanism by interaction with Akt kinase to regulate its phosphorylation levels rather than its stability. We further show that DPCD is a heart-shaped monomeric protein with two domains. A highly conserved region in the cysteine- and histidine-rich domains-containing proteins and SGT1 (CS) domain of DPCD interacts with the RUVBL2 DII domain with high affinity to form a stable R2TP-DPCD complex both in cellulo and in vitro. Considering that DPCD is one among several CS-domain-containing proteins found to associate with RUVBL1/2, we propose that RUVBL1/2 are CS-domain-binding proteins that regulate complex assembly and downstream signaling.
R2TP 是一个由 AAA+ ATPases RUVBL1 和 RUVBL2 以及 RPAP3 和 PIH1D1 蛋白组成的伴侣复合物。R2TP 负责组装主要通过不同衔接蛋白起作用的大分子复合物。通过邻近标记质谱法，我们鉴定出原发性纤毛运动障碍（DPCD）缺失蛋白是 R2TP 的衔接蛋白。在这里，我们证明 R2TP-DPCD 通过与 Akt 激酶相互作用来调节其磷酸化水平而不是稳定性，从而通过独特的机制影响纤毛发生起始。我们进一步表明，DPCD 是一种具有两个结构域的心脏形单体蛋白。DPCD 中富含半胱氨酸和组氨酸的结构域以及 SGT1（CS）结构域中的高度保守区域与 RUVBL2 DII 结构域以高亲和力相互作用，在细胞内和体外形成稳定的 R2TP-DPCD 复合物。考虑到 DPCD 是与 RUVBL1/2 相关的几个 CS 结构域蛋白之一，我们提出 RUVBL1/2 是 CS 结构域结合蛋白，可调节复合物组装和下游信号转导。
