Nedd4-2-dependent Ubiquitination Potentiates the Inhibition of Human NHE3 by Cholera Toxin and Enteropathogenic Escherichia coli

• Type of Material: Article
• Authors: Kayte A Jenkin, Emory University; Yiran Han, Emory University; Songbai Lin, Emory University; Peijian He, Emory University; Chris C Yun, Emory University
• Language: English
• Date: 2022-01-01
• Publisher: ELSEVIER INC
• Publication Version: Final Published Version
• Copyright Statement: © 2021 The Authors
• License: Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1016/j.jcmgh.2021.11.006
• Title of Journal or Parent Work: CELLULAR AND MOLECULAR GASTROENTEROLOGY AND HEPATOLOGY
• Volume: 13
• Issue: 3
• Start Page: 695
• End Page: 716
• Grant/Funding Information: Supported by grants from the National Institutes of Health (R01DK107719) and the Veterans Administration Merit Award (I01BX004459). Confocal microscopic analyses were supported in part by the Integrated Cellular Imaging Shared Resources of Winship Cancer Institute of Emory University and NIH/NCI under award P30CA138292.
• Keywords: Sodium Transporter; EXPRESSION; LIGASE; NA+/H+ EXCHANGER NHE3; Phosphorylation; Gastroenterology & Hepatology; Ubiquitination; CHANNEL; Life Sciences & Biomedicine; Diarrhea; ENDOCYTOSIS; PROTEIN-KINASE-C; NEDD4-2; Science & Technology; PHOSPHORYLATION; ACTIVATION; INTESTINAL ION-TRANSPORT
• Research Categories: Health Sciences, Oncology

Abstract

Background & Aims: Diarrhea is one of the most common illnesses and is often caused by bacterial infection. Recently, we have shown that human Na+/H+ exchanger NHE3 (hNHE3), but not non-human NHE3s, interacts with the E3 ubiquitin ligase Nedd4-2. We hypothesize that this property of hNHE3 contributes to the increased severity of diarrhea in humans. Methods: We used humanized mice expressing hNHE3 in the intestine (hNHE3int) to compare the contribution of hNHE3 and mouse NHE3 to diarrhea induced by cholera toxin (CTX) and enteropathogenic Escherichia coli (EPEC). We measured Na+/H+ exchange activity and fluid absorption. The role of Nedd4-2 on hNHE3 activity and ubiquitination was determined by knockdown in Caco-2bbe cells. The effects of protein kinase A (PKA), the primary mediator of CTX-induced diarrhea, on Nedd4-2 and hNHE3 phosphorylation and their interaction were determined. Results: The effects of CTX and EPEC were greater in hNHE3int mice than in control wild-type (WT) mice, resulting in greater inhibition of NHE3 activity and increased fluid accumulation in the intestine, the hallmark of diarrhea. Activation of PKA increased ubiquitination of hNHE3 and enhanced interaction of Nedd4-2 with hNHE3 via phosphorylation of Nedd4-2 at S342. S342A mutation mitigated the Nedd4-2–hNHE3 interaction and blocked PKA-induced inhibition of hNHE3. Unlike non-human NHE3s, inhibition of hNHE3 by PKA is independent of NHE3 phosphorylation, suggesting a distinct mechanism of hNHE3 regulation. Conclusions: The effects of CTX and EPEC on hNHE3 are amplified, and the unique properties of hNHE3 may contribute to diarrheal symptoms occurring in humans.