Department of Molecular & Cell Biology, University of California, Berkeley, Berkeley, CA, 94720, USA.
Helen Wills Neuroscience Institute, University of California, Berkeley, Berkeley, CA, 94720, USA.
Nat Commun. 2021 Nov 25;12(1):6913. doi: 10.1038/s41467-021-27283-8.
Tweety homologs (TTYHs) comprise a conserved family of transmembrane proteins found in eukaryotes with three members (TTYH1-3) in vertebrates. They are widely expressed in mammals including at high levels in the nervous system and have been implicated in cancers and other diseases including epilepsy, chronic pain, and viral infections. TTYHs have been reported to form Ca- and cell volume-regulated anion channels structurally distinct from any characterized protein family with potential roles in cell adhesion, migration, and developmental signaling. To provide insight into TTYH family structure and function, we determined cryo-EM structures of Mus musculus TTYH2 and TTYH3 in lipid nanodiscs. TTYH2 and TTYH3 adopt a previously unobserved fold which includes an extended extracellular domain with a partially solvent exposed pocket that may be an interaction site for hydrophobic molecules. In the presence of Ca, TTYH2 and TTYH3 form homomeric cis-dimers bridged by extracellularly coordinated Ca. Strikingly, in the absence of Ca, TTYH2 forms trans-dimers that span opposing membranes across a ~130 Å intermembrane space as well as a monomeric state. All TTYH structures lack ion conducting pathways and we do not observe TTYH2-dependent channel activity in cells. We conclude TTYHs are not pore forming subunits of anion channels and their function may involve Ca-dependent changes in quaternary structure, interactions with hydrophobic molecules near the extracellular membrane surface, and/or association with additional protein partners.
Tweety 同源物（TTYHs）是一种在真核生物中广泛存在的跨膜蛋白保守家族，脊椎动物中有三个成员（TTYH1-3）。它们在哺乳动物中广泛表达，包括在神经系统中高水平表达，并与癌症和其他疾病有关，包括癫痫、慢性疼痛和病毒感染。据报道，TTYHs 形成 Ca 和细胞体积调节的阴离子通道，在结构上与任何已鉴定的蛋白家族都不同，可能在细胞黏附、迁移和发育信号转导中发挥作用。为了深入了解 TTYH 家族的结构和功能，我们确定了 Mus musculus TTYH2 和 TTYH3 在脂质纳米盘中的冷冻电镜结构。TTYH2 和 TTYH3 采用了以前未观察到的折叠方式，包括一个延伸的细胞外结构域，其中部分溶剂暴露的口袋可能是与疏水分子相互作用的位点。在 Ca 的存在下，TTYH2 和 TTYH3 形成同源 cis-二聚体，通过细胞外配位的 Ca 桥接。引人注目的是，在没有 Ca 的情况下，TTYH2 形成跨二聚体，跨越约 130 Å 的膜间空间，以及单体状态。所有 TTYH 结构都缺乏离子传导途径，我们在细胞中也没有观察到 TTYH2 依赖性的通道活性。我们得出结论，TTYHs 不是阴离子通道的孔形成亚基，它们的功能可能涉及 Ca 依赖性的四级结构变化、与细胞外膜表面附近疏水分子的相互作用，以及/或与其他蛋白伴侣的结合。
