van der Wijst, J. | Search bioRxiv

bioRxiv2021

Functional basis for calmodulation of the TRPV5 calcium channel

Within the transient receptor potential (TRP) superfamily of ion channels, TRPV5 is a highly Ca2+-selective channel important for active reabsorption of Ca2+ in the kidney. Its channel activity is controlled by a negative feedback mechanism involving calmodulin (CaM) binding. Combining advanced microscopy techniques and biochemical assays, this study characterized the dynamic bilobal CaM regulation and binding stoichiometry. We demonstrate for the first time that functional (full-length) TRPV5 interacts with CaM in the absence of Ca2+, and this interaction is intensified at increasing Ca2+ concentrations sensed by the CaM C-lobe that achieves channel pore blocking. Channel inactivation occurs without CaM N-lobe calcification. Moreover, we reveal a 1:2 stoichiometry of TRPV5:CaM binding by implementing single molecule photobleaching counting, a technique with great potential for studying TRP channel regulation. In conclusion, our study proposes a new model for CaM- dependent regulation - calmodulation - of the Ca2+-selective TRPV5 that involves apoCaM interaction and lobe-specific actions.

cell biology↗

bioRxiv2018

Structural insight into TRPV5 channel function and modulation

TRPV5 (transient receptor potential vanilloid) is a unique calcium-selective TRP channel that is essential for calcium homeostasis. TRPV5 and its close homologue TRPV6 do not exhibit thermosensitivity or ligand-dependent activation, unlike other TRPV channels, but are constitutively opened at physiological membrane potentials. Here, we report high resolution electron cryo-microscopy (cryo-EM) structures of truncated and full length TRPV5 in lipid nanodisc, as well as a TRPV5 W583A mutant structure and a complex structure of TRPV5 with calmodulin (CaM). These structures highlight and explain functional differences between the thermosensitive and calcium-selective TRPV channels. An extended S1-S2 linker folds on top of the channel that might shield it from modulation by extracellular factors. Resident lipid densities in the homologous vanilloid pocket are different from those previously found in TRPV1, supporting a comparatively more rigid architecture of TRPV5. A ring of tryptophan residues (W583) at the bottom of the pore coordinates a density and mutation of W583 resultes in opening of the lower gate. Moreover, we provide structural insight into the calcium-dependent channel inactivation and propose a flexible stoichiometry for TRPV5 and CaM binding.

biophysics↗