Z Gastroenterol 2006; 44 - P104
DOI: 10.1055/s-2006-950687

Differential effects of PDZ-Adapterprotein NHERF1, E3KARP and PDZK1 knockout on the regulation of NHE3 transport activity in native murine colonic epithelium

A Cinar 1, M Chen 1, B Riederer 1, B Hogema 2, H de Jonge 2, M Donowitz 3, EJ Weinman 4, O Kocher 5, U Seidler 1
  • 1Medizinische Hochschule Hannover, Hannover, Germany
  • 2Erasmus Universität, Rotterdam, Netherlands
  • 3John-Hopkins University, Baltimore, United States of America
  • 4University of Maryland, Baltimore, United States of America
  • 5Beth Israel Deaconess Medical Center, Boston, United States of America

Background & Aims: The PDZ-adapter proteins NHERF1, E3KARP and PDZK1 have all been shown to bind to the sodium hydrogen exchanger NHE3, and to be expressed in the brush border membrane (BBM) of murine intestine. In PS120 fibroblasts, NHERF1 or E3KARP coexpression with NHE3 is required for cAMP-mediated inhibition of NHE3 trans-port activity. We therefore investigated, whether the three NHE3 binding PDZ-proteins show functional redundancy, or have differential roles in the native intestinal epithelium.

Methods: NHERF1, E3KARP, and PDZK1deficient mice and their wild type NHE3 transport rates were assessed fluorometrically in the NHE3-expressing surface cell region of BCECF-loaded mur-ine colonic crypts isolated from (wt) littermates. NHE3 BBM abudance was measured by Western analysis of isolated BBM, NHE3 localization was assessed immunohistochemically, and NHE3 mRNA levels were determined by RT-PCR.

Results: In NHERF1-deficient sur-face enterocytes, acid-activated NHE3 transport rate was significantly reduced to 65% of the rate in wt enterocytes. Significant inhibition of NHE3 transport activity by forskolin was ob-served in NHERF1-deficient surface enterocytes, but this inhibition was significantly less pronouned than in wt enterocytes (56% vs. 75%, p<0.01).. In E3KARP-deficient colonic en-terocytes, acid-activated NHE3 transport rate, as well as inhibition by forskolin and 8-Br-cGMP was identical to wt enterocytes, but the inhibition by ionomycin was completely abol-ished. In PDZK1-deficient surface enterocytes, acid-activated NHE3 transport rate was strongly reduced to 35% of that in wt enterocytes, and no significant inhibition of transport rate was seen by any of the above second messengers. NHE3 BBM abundance was reduced in NHERF1 and PDZK1 -/- BBM, but not in NHERF2 BBM.

Conclusions: The results demon-strate that the PDZ-Proteins NHERF1, E3KARP and PDZK1 are important regulators of NHE3 transport activity in native murine intestinal epithelium and appear to serve different, and signal-specific functions.