HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Jafri, F. Articles by Ergul, A. Search for Related Content PubMed PubMed Citation Articles by Jafri, F. Articles by Ergul, A.

---

BASIC BIOLOGY

Phosphorylation of Endothelin Converting Enzyme-1 Isoforms: Relevance to Subcellular Localization

Clinical and Experimental Therapeutics Program, University of Georgia College of Pharmacy and Vascular Biology Center, Medical College of Georgia, Augusta, Georgia 30912

To whom requests for reprints should be addressed at ¹ Medical College of Georgia, Clinical Pharmacy CJ-1020, Augusta, GA 30912. E-mail: aergul{at}mail.mcg.edu

Abstract

Endothelin-converting enzyme (ECE)-1 is a metalloenzyme with four subisoforms, which differ only in their amino-terminal domain. ECE-1a and c are the most common isoforms and are found at the plasma membrane and in the Golgi complex, whereas ECE-1b displays lysosomal localization. We have recently shown that ECE-1a but not ECE-1b also colocalizes with nuclear membrane markers, and that maintenance of cells in high glucose (25 mM) promotes relocalization of ECE-1a from the membrane to the intracellular compartment. To investigate the mechanisms involved in this process, we conducted a search for potential phosphorylation sites, which yielded a different number of putative sites for protein kinase (PK)-C and PKA in the amino-terminal region. Stimulation of Chinese hamster ovary (CHO) cells expressing a green fluorescent protein (GFP)-tagged human ECE-1a or ECE-1b with 100 nM phorbol myristate acetate (PMA) resulted in phosphorylation of ECE-1a, as determined by immunoprecipitation with an antibody to GFP followed by immunoblotting with an antibody to phosphoserine. Stimulation of cells with PMA also promoted intracellular relocalization, as seen in cells grown under high-glucose conditions. Incubation of cells grown in 25 mM glucose with the PKC inhibitor, calphostin C (100 nM), partially prevented the relocalization of ECE-1a from the plasma membrane to intracellular compartments. Stimulation of cells with 100 nM forskolin caused phosphorylation of ECE-1b and not ECE-1a, which is consistent with the lack of a putative PKA site in the ECE-1a amino-terminal sequence. Although phosphorylation is not required for ECE-1 enzymatic activity, these results suggest that ECE-1 isoforms are phosphorylated and that phosphorylation might play an important role in the regulation of intracellular trafficking of ECE-1 subisoforms.

Key Words: ECE • phosphorylation • localization

This article has been cited by other articles:

				B. E. Padilla, G. S. Cottrell, D. Roosterman, S. Pikios, L. Muller, M. Steinhoff, and N. W. Bunnett Endothelin-converting enzyme-1 regulates endosomal sorting of calcitonin receptor-like receptor and -arrestins J. Cell Biol., December 3, 2007; 179(5): 981 - 997. [Abstract] [Full Text] [PDF]

				D. Roosterman, G. S. Cottrell, B. E. Padilla, L. Muller, C. B. Eckman, N. W. Bunnett, and M. Steinhoff Endothelin-converting enzyme 1 degrades neuropeptides in endosomes to control receptor recycling PNAS, July 10, 2007; 104(28): 11838 - 11843. [Abstract] [Full Text] [PDF]