Cryo-EM Structure of Native Human Uromodulin, a Zona Pellucida Module Polymer

Abstract

SUMMARYAssembly of extracellular filaments and matrices mediating fundamental biological processes such as morphogenesis, hearing, fertilization and antibacterial defense is driven by a ubiquitous polymerization module known as zona pellucida (ZP) “domain”. Despite the conservation of this element from hydra to human, no information is available on the filamentous conformation of any ZP module protein. Here we report the cryo-electron microscopy structure of uromodulin (UMOD)/Tamm-Horsfall protein, the most abundant protein in human urine and an archetypal ZP module-containing molecule, in its mature homopolymeric state. UMOD forms a one-start helix with an unprecedented 180-degree twist between subunits enfolded by interdomain linkers that have completely reorganized as a result of propeptide dissociation. Lateral interaction between filaments in the urine generates sheets exposing a checkerboard of binding sites to capture uropathogenic bacteria, and UMOD-based models of mammalian and avian heteromeric egg coat filaments identify a common sperm-binding region at the interface between subunits.

Cryo‐EM structure of native human uromodulin, a zona pellucida module polymer

“…Notably, our data also do not support the UMOD architecture suggested by a recent tomographic analysis, where polymers are made up of ZP modules that simply stack in a zig‐zag manner, without any conformational change of the interdomain linker and associated separation of ZP‐N and ZP‐C (Weiss et al , 2020). On the other hand, our results are in agreement with a study of the UMOD filament core (Stanisich et al , 2020) that appeared while this work was under revision and publicly available as a preprint version (preprint: Stsiapanava et al , 2020).…”

The cryo-EM structure of the human uromodulin filament core reveals a unique assembly mechanism

“…This UMOD assembly model is analogous to the recently proposed assembly mechanism of filamentous type V pili, in which assembly is linked with proteolytic release of pilus subunits from the outer bacterial membrane ( Shibata et al, 2020 ). During the preparation of this manuscript, a related preprint article on the cryo-EM structure of the UMOD filament core was published ( Stsiapanava et al, 2020 ). In said study, an alternative model of UMOD assembly was proposed, based on the assumption that assembly starts from membrane-bound pro-UMOD homodimers.…”

A C. elegans Zona Pellucida domain protein functions via its ZPc domain

“…In proteins like uromodulin that have polymerizing ZPn domains, ZPc domains are thought to inhibit the ZPn domain [30]. While our study was under review, two groups reported cryo-EM structures for full-length uromodulin that suggest filament assembly involves interactions of both ZPn and ZPc domains with linker regions of other monomers, which would suggest that both domains are functionally important [69,70]. In the TGFβ receptors Betaglycan and Endoglin, ZPc domains dimerize and can bind ligands, but have not been reported to form higher-order structures [28,29,71].…”