Abstract

Phosphorylation and lysine (K)‐acetylation are dynamic post‐translational modifications of proteins. K‐acetylation of histone proteins is classically regarded as a critical regulatory pathway for “opening” chromatin structure and promoting transcription. Previous proteomic studies have identified over 170,000 phosphorylation sites, and greater than 15,000 K‐acetylation sites of mammalian proteins. We recently reported that the inner medullary collecting duct (IMCD), which functions in the regulation of vasopressin‐dependent water‐reabsorption, expresses many of the enzymes (acetyltransferases and deacetylases) that can modulate K‐acetylation. The purpose of this study was to determine the K‐acetylated or phosphorylated proteins expressed in IMCD cells. Second, we asked whether vasopressin V2 receptor activation significantly affects the IMCD acetylome or phosphoproteome? K‐acetylated or serine‐, threonine‐, or tyrosine‐phosphorylated peptides were identified from native rat IMCDs using proteomic analysis with four different enzymes (trypsin, chymotrypsin, ASP‐N, or Glu‐C) to generate a high‐resolution proteome. We identified 709 K‐acetylated sites that mapped to 431 unique proteins. Of these K‐acetylated sites 583 (64%) were novel. The K‐acetylated proteins were expressed in all compartments of the cell, and were enriched in pathways including glycolysis (12 proteins), the TCA cycle (8 proteins), and vasopressin‐regulated water reabsorption (8 proteins). In these same IMCD cells, we detected 312 phosphorylated sites that mapped to 215 proteins, including AQP2 peptides that were phosphorylated at 4 serines: 256, 261, 264, and 269. Moreover, we identified 155 novel phosphorylation sites. Acute dDAVP did not significantly affect the IMCD acetylome; however it did significantly affect previously known vasopressin‐regulated phosphorylation sites. In conclusion, presence of K‐acetylated proteins involved in metabolism, ion and water transport in the IMCD points to multiple roles of K‐acetylation beyond its canonical role in transcriptional regulation.Support or Funding InformationK01‐DK‐105038 to K. A. H., projects ZIA‐HL‐001285 and ZIA‐HL‐006129, M. A. K.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

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