Extracellular pH Dynamically Controls Cell Surface Delivery of Functional TRPV5 Channels

Abstract

Extracellular pH has long been known to affect the rate and magnitude of ion transport processes among others via regulation of ion channel activity. The Ca 2؉ -selective transient receptor potential vanilloid 5 (TRPV5) channel constitutes the apical entry gate in Ca 2؉ -transporting cells, contributing significantly to the overall Ca 2؉ balance. Here, we demonstrate that extracellular pH determines the cell surface expression of TRPV5 via a unique mechanism. By a comprehensive approach using total internal reflection fluorescence microscopy, cell surface protein labeling, electrophysiology, 45 Ca 2؉ uptake assays, and functional channel recovery after chemobleaching, this study shows that upon extracellular alkalinization, a pool of TRPV5-containing vesicles is rapidly recruited to the cell surface without collapsing into the plasma membrane. These vesicles contain functional TRPV5 channels since extracellular alkalinization is accompanied by increased TRPV5 activity. Conversely, upon subsequent extracellular acidification, vesicles are retrieved from the plasma membrane, simultaneously resulting in decreased TRPV5 activity. Thus, TRPV5 accesses the extracellular compartment via transient openings of vesicles, suggesting that rapid responses of constitutive active TRP channels to physiological stimuli rely on vesicular "kiss and linger" interactions with the plasma membrane.The superfamily of transient receptor potential (TRP) channels is involved in diverse physiological processes, ranging from sensory activity to fertility and epithelial ion transport (15). The highly Ca 2ϩ -selective TRP vanilloid 5 (TRPV5) channel constitutes the apical entry gate in Ca 2ϩ -transporting cells and facilitates renal Ca 2ϩ influx from the prourine (10). Several lines of evidence indicate that TRPV5 activity is sensitive to pH. First, acid-based homeostasis is known to affect renal Ca 2ϩ handling as reflected by altered Ca 2ϩ excretion in kidneys during chronic acidosis or alkalosis, which is mediated at least in part by changes in TRPV5 gene expression (16). Second, in vitro studies indicated that intra-and extracellular pH directly regulate the activity of TRPV5. Acidification inhibited, whereas alkalinization stimulated, TRPV5 activity, likely mediated by conformational changes of the channel pore helix (24-26). An intrinsic physiological effect of extracellular pH is the regulation of trafficking processes like endo-and exocytosis and lysosomal trafficking (8,12,14). Since several TRP channels display constitutive activity, controlled recruitment of these channels towards the plasma membrane is important for the translation of physiological stimuli into increased ion permeability of the plasma membrane. For instance, an essential process during insulin-like growth factor-I stimulation of cell growth is TRPV2 recruitment facilitating Ca 2ϩ entry during progression through the cell cycle (11). In Drosophila photoreceptors, the TRP-like subunit is shuttled between the plasma membrane and an intracellular compartment by a light...

TRPV1 acts as a synaptic protein and regulates vesicle recycling

“…Also, TRPM7 and TRPV4 have been detected in small synaptic-like vesicles and in spines (Brauchi et al, 2008;Shibasaki et al, 2007). Our data, together with accumulating reports of TRPV5, indicate a more general role for TRPV channels in vesicle fusion (Lambers et al, 2007).…”

“…Also, TRPM7 and TRPV4 have been detected in small synaptic-like vesicles and in spines (Brauchi et al, 2008;Shibasaki et al, 2007). Our data, together with accumulating reports of TRPV5, indicate a more general role for TRPV channels in vesicle fusion (Lambers et al, 2007).But is TRPV1 also involved in the process of neuronal network formation? Our data and other reports suggest several cases where TRPV1 can be involved in network formation.…”

Role of the Transient Receptor Potential Vanilloid 5 (TRPV5) Protein N Terminus in Channel Activity, Tetramerization, and Trafficking

“…Therefore, other biochemical experiments like cell surface biotinylation were used to investigate the expression of TRPV5 at the plasma membrane (27). Because TRPV5-⌬1-38 was retained in the endoplasmic reticulum we investigated whether this mutant still reached the plasma membrane.…”

Active Ca2+ reabsorption in the connecting tubule

“…Recent advances in total reflection fluorescent microscopy analysis allowed the identification of the molecular basis underlying these in vivo observations. Exposure of TRPV5-expressing cells to an alkaline extracellular environment (pH 8.0) caused rapid recruitment of TRPV5-containing vesicles to the cell surface and a consequent increase in TRPV5 activity [40]. In the reciprocal experiment, acidic extracellular milieu (pH 6.5) induced the internalization of TRPV5-containing vesicles from the plasma membrane, resulting in a reduced channel activity [40].…”

“…Exposure of TRPV5-expressing cells to an alkaline extracellular environment (pH 8.0) caused rapid recruitment of TRPV5-containing vesicles to the cell surface and a consequent increase in TRPV5 activity [40]. In the reciprocal experiment, acidic extracellular milieu (pH 6.5) induced the internalization of TRPV5-containing vesicles from the plasma membrane, resulting in a reduced channel activity [40]. The extracellular acidity clearly affected the current kinetics resulting in diminished single channel conductance as shown by Yeh et al [77].…”