Department of Molecular Medicine, Cornell University, Ithaca, NY 14853.
Field of Biochemistry, Molecular, and Cell Biology, Cornell University, Ithaca, NY 14853.
Mol Biol Cell. 2024 Oct 1;35(10):ar127. doi: 10.1091/mbc.E24-05-0218. Epub 2024 Aug 7.
Genetic, colocalization, and biochemical studies suggest that the ankyrin repeat-containing proteins Inversin (INVS) and ANKS6 function with the NEK8 kinase to control tissue patterning and maintain organ physiology. It is unknown whether these three proteins assemble into a static "Inversin complex" or one that adopts multiple bioactive forms. Through the characterization of hyperactive alleles in , we discovered that the Inversin complex is activated by dimerization. Genome engineering of an RFP tag onto the nematode homologues of INVS (MLT-4) and NEK8 (NEKL-2) induced a gain-of-function, cyst-like phenotype that was suppressed by monomerization of the fluorescent tag. Stimulated dimerization of MLT-4 or NEKL-2 using optogenetics was sufficient to recapitulate the phenotype of a constitutively active Inversin complex. Further, dimerization of NEKL-2 bypassed a lethal MLT-4 mutant, demonstrating that the dimeric form is required for function. We propose that dynamic switching between at least two functionally distinct states - an active dimer and an inactive monomer - gates the output of the Inversin complex.
遗传、共定位和生化研究表明，含有锚蛋白重复序列的蛋白 Inversin（INVS）和 ANKS6 与 NEK8 激酶一起作用，控制组织模式形成并维持器官生理学。目前尚不清楚这三种蛋白是否组装成一个静态的“Inversin 复合物”，还是形成多种具有生物活性的形式。通过对 的超活性等位基因的特征分析，我们发现 Inversin 复合物通过二聚化激活。通过基因组工程将 RFP 标签标记到线虫同源物 INVS（MLT-4）和 NEK8（NEKL-2）上，诱导出一种获得功能的、类似囊肿的表型，该表型被荧光标签的单体化抑制。使用光遗传学刺激 MLT-4 或 NEKL-2 的二聚化足以重现组成性激活的 Inversin 复合物的表型。此外，NEKL-2 的二聚化绕过了致死性的 MLT-4 突变体，表明二聚体形式是必需的。我们提出，至少两种功能不同的状态——活性二聚体和非活性单体之间的动态转换——控制着 Inversin 复合物的输出。
