Transient Receptor Potential (TRP) channels are evolutionarily conserved integral membrane proteins. The mammalian TRP superfamily of ion channels consists of 28 cation permeable channels that are grouped into six subfamilies based on sequence homology (Fig. 6.1). The canonical TRP (TRPC) subfamily is known for containing the founding member of mammalian TRP channels. The vanilloid TRP (TRPV) subfamily has been extensively studied due to the heat sensitivity of its founding member. The melastatin-related TRP (TRPM) subfamily includes some of the few known bi-functional ion channels, which contain functional enzymatic domains. The ankyrin TRP (TRPA) subfamily consists of a single chemo-nociceptor that has been proposed to be a target for analgesics. The mucolipin TRP (TRPML) subfamily channels are found primarily in intracellular compartments and were discovered based on their critical role in type IV mucolipidosis (ML-IV). The polycystic TRP (TRPP) subfamily is a diverse group of proteins implicated in autosomal dominant polycystic kidney disease (ADPKD). Overall, this superfamily of channels is involved in a vast array of physiological and pathophysiological processes making the study of these channels imperative to our understanding of subcellular biochemistry.

TRPV5 is a transient receptor potential channel involved in calcium reabsorption. Here we investigate the interaction of two endogenous modulators with TRPV5. Both phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and calmodulin (CaM) have been shown to directly bind to TRPV5 and activate or inactivate the channel, respectively. Using cryo-electron microscopy (cryo-EM), we determined TRPV5 structures in the presence of dioctanoyl PI(4,5)P2 and CaM. The PI(4,5)P2 structure reveals a binding site between the N-linker, S4-S5 linker and S6 helix of TRPV5. These interactions with PI(4,5)P2 induce conformational rearrangements in the lower gate, opening the channel. The CaM structure reveals two TRPV5 C-terminal peptides anchoring a single CaM molecule and that calcium inhibition is mediated through a cation-π interaction between Lys116 on the C-lobe of calcium-activated CaM and Trp583 at the intracellular gate of TRPV5. Overall, this investigation provides insight into the endogenous modulation of TRPV5, which has the potential to guide drug discovery.

Transient receptor potential (TRP) proteins form a superfamily Ca2+-permeable cation channels regulated by a range of chemical and physical stimuli. Structural analysis of a ‘minimal' TRP vanilloid subtype 1 (TRPV1) elucidated a mechanism of channel activation by agonists through changes in its outer pore region. Though homologous to TRPV1, other TRPV channels (TRPV2–6) are insensitive to TRPV1 activators including heat and vanilloids. To further understand the structural basis of TRPV channel function, we determined the structure of full-length TRPV2 at ∼5 Å resolution by cryo-electron microscopy. Like TRPV1, TRPV2 contains two constrictions, one each in the pore-forming upper and lower gates. The agonist-free full-length TRPV2 has wider upper and lower gates compared with closed and agonist-activated TRPV1. We propose these newly revealed TRPV2 structural features contribute to diversity of TRPV channels.

Summary Synaptic vesicle (SV) exo- and endocytosis are tightly coupled to sustain neurotransmission in presynaptic terminals, and both are regulated by Ca2+. Ca2+ influx triggered by voltage-gated Ca2+ channels is necessary for SV fusion. However, extracellular Ca2+ has also been shown to be required for endocytosis. The intracellular Ca2+ levels (< 1μM) that trigger endocytosis are typically much lower than those (> 10μM) needed to induce exocytosis, and endocytosis is inhibited when the Ca2+ level exceeds 1μM. Here we identify and characterize a transmembrane protein associated with SVs, which upon SV fusion, localizes at periactive zones. Loss of Flower results in impaired intracellular resting Ca2+ levels and impaired endocytosis. Flower multimerizes and is able to form a channel to control Ca2+ influx. We propose that Flower functions as a Ca2+ channel to regulate synaptic endocytosis and hence couples exo- with endocytosis.