In:
Physiology Journal, Hindawi Limited, Vol. 2013 ( 2013-04-16), p. 1-9
Abstract:
One response to hypertonic stress in the renal medulla and MDCK cells is the upregulation of betaine transporter (BGT1) synthesis, followed by trafficking to the plasma membrane (PM) and an increase in betaine transport. Upregulation of BGT1 was enhanced by inhibitors of phosphatases PP1 and PP2A and was attenuated by inhibitors of protein kinase C, suggesting an important role for phosphorylation reactions. This was tested using mutants of BGT1 tagged with EGFP. The PM trafficking motifs of BGT1 reside near the C terminus, and truncation at lysine 560 resulted in a protein that remained intracellular during hypertonic stress. This K560 Δ mutant colocalized with endoplasmic reticulum (ER). Substitution of alanine at Thr 40 , a putative phosphorylation site, also prevented trafficking to the PM during hypertonic stress. Live-cell imaging showed that T40A was not retained in the ER and colocalized with markers for Golgi and endosomes. In contrast, substitution of aspartate or glutamate at Thr 40 , to mimic phosphorylation, restored normal trafficking to the PM. HEK293 cells transfected with K560 Δ or T40A mutants had 10% of the GABA transport activity of native BGT1, but normal transport activity was restored in cells expressing T40E. Normal BGT1 trafficking likely requires phosphorylation at Thr 40 in addition to C-terminal motifs.
Type of Medium:
Online Resource
ISSN:
2314-4300
,
2314-4319
Language:
English
Publisher:
Hindawi Limited
Publication Date:
2013
detail.hit.zdb_id:
2710792-9
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