Roles of Pak2 in Collecting Duct Cell Polarization and <i>Aqp2</i> Transcription

Vasopressin regulates Aqp2 transcription in the kidney collecting duct principal cells. Dysregulation in Aqp2 transcription is associated with many water balance disorders. Previously, we showed a positive correlation between the mRNA levels of Aqp2 and Pak2, P21 activated kinase 2, known to be involved in cell polarization, a condition required for cell responses to vasopressin with Aqp2 transcription. The Knepper’s lab showed increases in Pak2 protein levels in the nucleus and cytoplasm of the collecting duct principal cell model mpkCCD in response to vasopressin V2 receptor-specific analog dDAVP. The above observations are suggestive of Pak2’s roles in collecting duct cell polarization and Aqp2 transcription. Small-hairpin RNA-mediated Pak2 knockdown significantly reduced transepithelial electrical resistance of the mpkCCD cells grown on Transwell® plates. Pak2 knockdown cells failed to respond to dDAVP with Aqp2 mRNA and protein expression. Immunoblotting appeared to show lower levels of E-cadherin and ZO1 in the Pak2 knockdown cells. Mechanistically, Pak2 is known to phosphorylate LIM kinase and cofilin, an F-actin binding protein that promotes G-actin formation. G-actin binds myocardin-related transcription factor and inhibits transcription mediated by the serum-response transcription factor. Aqp2 has conserved serum-response factor binding elements in the 5’-flanking region. Consistent with this pathway, immunofluorescence confocal microscopy showed that Pak2 knockdown increased the amount of G-actin. Likewise, immunofluorescence confocal microscopy and biochemical measurements showed that cytochalasin D, an agent that inhibits actin polymerization, increased the amount of G-actin. RT-qPCR showed that cytochalasin D recapitulated the effects of Pak2 knockdown in Aqp2 transcription. Overall, our data are in line with roles of Pak2 in collecting duct cell polarization and Aqp2 transcription via G-actin. MOST 111-2320-B-002-084-MY3, National Science and Technology Council, Taipei, Taiwan. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.