This Article Full Text Full Text (PDF) Submit a related Letter to the Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Brothers, S. P. Articles by Conn, P. M. Search for Related Content PubMed PubMed Citation Articles by Brothers, S. P. Articles by Conn, P. M.

Unexpected Effects of Epitope and Chimeric Tags on Gonadotropin-Releasing Hormone Receptors: Implications for Understanding the Molecular Etiology of Hypogonadotropic Hypogonadism

Division of Neuroscience, Oregon National Primate Research Center and Departments of Physiology and Pharmacology and Cell and Developmental Biology, Oregon Health and Science University, Beaverton, Oregon 97006

Address all correspondence and requests for reprints to: P. Michael Conn, Oregon National Primate Research Center/Oregon Health and Science University, 505 NW 185th Avenue, Beaverton, Oregon 97006. E-mail: connm{at}ohsu.edu.

In the case of human GnRH receptor (GnRHR) mutants associated with hypogonadotropic hypogonadism, a view emerged that these mutants are correctly routed to the plasma membrane. This view, supported almost entirely by studies using the HA-tag (hemagglutinin influenza virus epitope tag) and other epitope and chimeric tags, obscured recognition that GnRHR mutants frequently become misrouted proteins. The underlying assumption in epitope and chimeric tagging studies is that the cell does not distinguish tagged from unmodified proteins. It should not have been surprising, in retrospect, to find that even a single amino acid mutation dramatically alters protein function or routing because increased plasma membrane expression is associated with deletion of a single amino acid in the human GnRHR (K191), and point mutations have been shown to block plasma membrane routing of many receptors, including most of those responsible for the hypogonadotropic hypogonadism phenotype. Our present observations suggest that epitope and chimeric tags do have a significant effect on protein localization and function. Although rarely provided, control experiments addressing the effects of epitope or chimeric tagging are an essential part of any study relying on these proteomic tools.

This work was supported by National Institutes of Health Grants HD-19899, RR-00163, and HD-18185.

Abbreviations: Ctail, Terminal 51 amino acid cytoplasmic carboxyl terminus of the African catfish GnRHR; GFP, green fluorescent protein; GnRHR, GnRH receptor; HA, hemagglutinin influenza virus epitope; HH, hypogonadotropic hypogonadism; IP, inositol phosphates; N/C, average numbers of expressed receptors per cell; WT, wild type.

This article has been cited by other articles:

				J. H. Robben, M. Sze, N. V. A. M. Knoers, and P. M. T. Deen Functional rescue of vasopressin V2 receptor mutants in MDCK cells by pharmacochaperones: relevance to therapy of nephrogenic diabetes insipidus Am J Physiol Renal Physiol, January 1, 2007; 292(1): F253 - F260. [Abstract] [Full Text] [PDF]

				S. P Brothers, J. A. Janovick, and P M. Conn Calnexin regulated gonadotropin-releasing hormone receptor plasma membrane expression J. Mol. Endocrinol., December 1, 2006; 37(3): 479 - 488. [Abstract] [Full Text] [PDF]

				J. A. Janovick, P. E. Knollman, S. P. Brothers, R. Ayala-Yanez, A. S. Aziz, and P. M. Conn Regulation of G Protein-coupled Receptor Trafficking by Inefficient Plasma Membrane Expression: MOLECULAR BASIS OF AN EVOLVED STRATEGY J. Biol. Chem., March 31, 2006; 281(13): 8417 - 8425. [Abstract] [Full Text] [PDF]

				P. E. Knollman, J. A. Janovick, S. P. Brothers, and P. M. Conn Parallel Regulation of Membrane Trafficking and Dominant-negative Effects by Misrouted Gonadotropin-releasing Hormone Receptor Mutants J. Biol. Chem., July 1, 2005; 280(26): 24506 - 24514. [Abstract] [Full Text] [PDF]

				C. Castro-Fernandez, G. Maya-Nunez, and P. M. Conn Beyond the Signal Sequence: Protein Routing in Health and Disease Endocr. Rev., June 1, 2005; 26(4): 479 - 503. [Abstract] [Full Text] [PDF]

				C. K. Cheng and P. C. K. Leung Molecular Biology of Gonadotropin-Releasing Hormone (GnRH)-I, GnRH-II, and Their Receptors in Humans Endocr. Rev., April 1, 2005; 26(2): 283 - 306. [Abstract] [Full Text] [PDF]

				A. Ulloa-Aguirre, J. A. Janovick, A. Leanos-Miranda, and P. M. Conn Misrouted cell surface GnRH receptors as a disease aetiology for congenital isolated hypogonadotrophic hypogonadism Hum. Reprod. Update, March 1, 2004; 10(2): 177 - 192. [Abstract] [Full Text] [PDF]