SPAK dependent regulation of peptide transporters PEPT1 and PEPT2.
OBJECTIVE
SPAK (STE20-related proline/alanine-rich kinase) is a powerful regulator of renal tubular ion transport and blood pressure. Moreover, SPAK contributes to the regulation of cell volume. Little is known, however, about a role of SPAK in the regulation or organic solutes. The present study thus addressed the influence of SPAK on the peptide transporters PEPT1 and PEPT2.


METHODS
To this end, cRNA encoding PEPT1 or PEPT2 were injected into Xenopus laevis oocytes without or with additional injection of cRNA encoding wild-type, SPAK, WNK1 insensitive inactive (T233A)SPAK, constitutively active (T233E)SPAK, and catalytically inactive (D212A)SPAK. Electrogenic peptide (glycine-glycine) transport was determined by dual electrode voltage clamp and PEPT2 protein abundance in the cell membrane by chemiluminescence. Intestinal electrogenic peptide transport was estimated from peptide induced current in Ussing chamber experiments of jejunal segments isolated from gene targeted mice expressing SPAK resistant to WNK-dependent activation (spak(tg/tg)) and respective wild-type mice (spak(+/+)).


RESULTS
In PEPT1 and in PEPT2 expressing oocytes, but not in oocytes injected with water, the dipeptide gly-gly (2 mM) generated an inward current, which was significantly decreased following coexpression of SPAK. The effect of SPAK on PEPT1 was mimicked by (T233E)SPAK, but not by (D212A)SPAK or (T233A)SPAK. SPAK decreased maximal peptide induced current of PEPT1. Moreover, SPAK decreased carrier protein abundance in the cell membrane of PEPT2 expressing oocytes. In intestinal segments gly-gly generated a current, which was significantly higher in spak(tg/tg) than in spak(+/+) mice.


CONCLUSIONS
SPAK is a powerful regulator of peptide transporters PEPT1 and PEPT2.
Department of Physiology I, University of Tübingen, Gmelinstr. 5, 70276 Tübingen, Germany.
Jamshed
J
Warsi
Luo
L
Dong
Bernat
B
Elvira
Madhuri S
MS
Salker
Ekaterina
E
Shumilina
Zohreh
Z
Hosseinzadeh
Florian
F
Lang
D000818:Animals; D004151:Dipeptides; D007413:Intestinal Mucosa / Q000378:metabolism; D049449:Luminescence; D051379:Mice; D008822:Mice, Transgenic; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D000074011:Peptide Transporter 1; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000502:physiology*; D011817:Rabbits; D027981:Symporters / Q000235:genetics / Q000502:physiology*; D014982:Xenopus laevis
D004151:Dipeptides; D000074011:Peptide Transporter 1; D027981:Symporters; C093694:hydrogen-coupled oligopeptide transporter PepT2; C116037:PAS domain kinases; D017346:Protein Serine-Threonine Kinases