Skip to main content
Article thumbnail
Location of Repository

The role of Arg445 and Asp498 in the human thyroid hormone transporter MCT8

By S. (Stefan) Groeneweg, E.C.H. (Edith) Friesema, S. (Simone) Kersseboom, W. (Willem) Klootwijk, W.E. (Wil Edward) Visser, R.P. (Robin) Peeters and T.J. (Theo) Visser

Abstract

Monocarboxylate transporter 8 (MCT8) facilitates cellular influx and efflux of the thyroid hormones (THs) T4 and T3. Mutations in MCT8 lead to severe psychomotor retardation. Here, we studied the importance of 2 highly conserved residues (Arg445 in transmembrane domain 8 and Asp498 in transmembrane domain 10) for substrate recognition and helix interactions. We introduced single and double mutations (R445A, R445C, R445D, R445K, D498A, D498E, D498N, D498R, R445A+D498A, R445D+D498R, and R445K+D498E) in human MCT8 cDNA and studied the effects on MCT8-mediated TH uptake and metabolism in transfected cells. The impact of these mutations on MCT8 protein expression, dimerization capacity, and subcellular localization was studied by Western blottingandconfocal microscopy.Wefound that mutations in Arg445 or Asp498 that alter the local charge resulted in a near-complete loss of TH uptake capacity, whereas the expression, stability, and subcellular localization of these mutant proteins was similar to those for wild-type MCT8. Given the impaired TH uptake, TH efflux could not be adequately studied. The importance of opposite charges at Arg445 and Asp498 was studied by exchanging these residues (R445D+D498R). In particular, T4 uptake was less severely reduced by the exchange mutation than by the single mutations. Mutations of Arg445 and Asp498 to equally charged residues (R445K and/or D498E) resulted in TH uptake levels similar to wild-type MCT8. The presence of 2 oppositely charged residues at positions Arg445 and Asp498 that are predicted in close structural proximity is crucial for efficient TH uptake, which may indicate the presence of an, at least transient, charge pair between these residues. Copyrigh

Year: 2014
DOI identifier: 10.1210/en.2013-1521
OAI identifier: oai:repub.eur.nl:69260
Download PDF:
Sorry, we are unable to provide the full text but you may find it at the following location(s):
  • http://repub.eur.nl/pub/69260 (external link)
  • Suggested articles


    To submit an update or takedown request for this paper, please submit an Update/Correction/Removal Request.