The peptide transporter PEPT2 is targeted by the protein kinase SGK1 and the scaffold protein NHERF2.
PEPT1 and PEPT2 are members of the family of proton-dependent oligopeptide transporters that mediate electrogenic uphill transport of small peptides and peptidomimetics into a variety of cells. Kinetic properties and substrate recognition sites of those transporters have been well defined previously. Little is known, however, about regulation of those transporters. Both PEPT isoforms contain putative phosphorylation sites for the serum and glucocorticoid inducible kinase SGK1 and a C-terminal PDZ binding motif that might be recognized by PDZ domains of the Na(+)/H(+) exchanger regulatory factors NHERF1 and NHERF2. Thus, the present study attempted to clarify the role of SGK1 and NHERFs in the modulation of PEPT isoforms. Expression studies in Xenopus oocytes with subsequent electrophysiology and immunoassays revealed that SGK1 and NHERF2, but not the NHERF1 isoform specifically enhance PEPT2 function and surface abundance. The kinase is effective through phosphorylation of (185)Ser within the SGK1 consensus site, since disruption of this site prevented transporter modulation by the kinase. NHERF2 failed to regulate the C-terminal deletion mutant (PEPT2DeltaC) indicating that the C-terminal PDZ-binding motif in PEPT2 governs transport modulation by NHERF2. Coexpression of NHE3 stimulates PEPT2 activity to a similar extent as coexpression of NHERF2. Dynasore experiments demonstrated that SGK1 and NHERF2 activate PEPT2 by stabilizing the transporter at the cell surface. In conclusion, the present results reveal two novel PEPT2 posttranslational modulators, SGK1 and NHERF2, which might regulate transport of oligopeptides and peptidomimetic drugs.
Boehmer
Christoph
C
Department of Physiology I, University of Tübingen, Tübingen, Germany.
Palmada
Monica
M
Klaus
Fabian
F
Jeyaraj
Sankarganesh
S
Lindner
Ricco
R
Laufer
Jorg
J
Daniel
Hannelore
H
Lang
Florian
F
D020816:Amino Acid Motifs; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D004705:Endocytosis; D006801:Humans; D017874:Immediate-Early Proteins / Q000378:metabolism*; D015640:Ion Channel Gating; D007700:Kinetics; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D017923:Sodium-Hydrogen Exchangers / Q000737:chemistry / Q000378:metabolism*; D027981:Symporters / Q000737:chemistry / Q000378:metabolism*; D014981:Xenopus; D000099184:Serum-Glucocorticoid Regulated Kinases
D017874:Immediate-Early Proteins; D010750:Phosphoproteins; D017346:Protein Serine-Threonine Kinases; D017923:Sodium-Hydrogen Exchangers; D027981:Symporters; D000099184:Serum-Glucocorticoid Regulated Kinases; C093694:hydrogen-coupled oligopeptide transporter PepT2; C093376:sodium-hydrogen exchanger regulatory factor