Functional analysis of neural precursor cell expressed developmentally down regulated, Gene 4 isoforms that contain or lack a conserved region 2 of protein kinase C domain

A defining feature of NEDD4L protein isoforms is the presence or absence of an amino-terminal C2 domain, a class of subcellular, calcium-dependent targeting domains. We previously identified a common variant in human NEDD4L that generates isoforms that contain or lack a C2 domain. To address the potential functional significance of the NEDD4L common variant on NEDD4L subcellular localization, NEDD4L isoforms that either contained or lacked a C2 domain were tagged with enhanced green fluorescent protein, transfected into Xenopus laevis kidney epithelial cells, and imaged by performing confocal microscopy on live cells. We report that the presence or absence of this C2 domain exerts differential effects on the subcellular distribution of NEDD4L, the ability of C2 containing and lacking NEDD4L isoforms to mobilize in response to a calcium stimulus, and the intracellular transport of subunits of the NEDD4L substrate, ENaC. Furthermore, the ability of the C2-containing isoform to influence p-ENaC mobilization from intracellular pools involves the NEDD4L active site for ubiquitination. We propose a model to account for the potential impact of this common genetic variant on protein function at the cellular level. Additionally, to test in vivo the functional relevance of NEDD4L isoforms on sodium homeostasis and blood pressure we attempted to generate two gene targeted mice that would conditionally overexpress either NEDD4L isoform in the distal nephron. However, only the C2-lacking mouse was successfully generated so a comparative study between NEDD4L C2-containing and -lacking mice was not performed.