Expression and protein kinase C-dependent regulation of peptide/H+ co-transport system in the Caco-2 human colon carcinoma cell line

Brandsch, Matthias; Miyamoto, Yusei; Ganapathy, Vadivel; Leibach, Frederick H.

doi:10.1042/bj2990253

Cited by 130 publications

(98 citation statements)

References 27 publications

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“…To our knowledge, this Km is among the lowest Km s reported for hPepT1. For example Gly-Sar, which is the most commonly used PepT1 substrate, has a Km ≥ 1 mM in Caco2-BBE cells (33). Similar results were found in Jurkat cells.…”

Section: Discussionsupporting

confidence: 72%

PepT1-Mediated Tripeptide KPV Uptake Reduces Intestinal Inflammation

Dalmasso

Charrier-Hisamuddin²,

Nguyen

et al. 2008

Gastroenterology

102

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Background & Aims-KPV is a tripeptide (Lys-Pro-Val) which possesses anti-inflammatory properties however its mechanisms of action still remain unknown. PepT1 is a di/tripeptide transporter normally expressed in the small intestine and induced in colon during inflammatory bowel disease (IBD). The aim of this study was to 1) investigate whether KPV anti-inflammatory effect is PepT1-mediated in intestinal epithelial and immune cells, and 2) examine KPV anti-inflammatory effect in two models of mice colitis.

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Section: Discussionsupporting

confidence: 72%

PepT1-Mediated Tripeptide KPV Uptake Reduces Intestinal Inflammation

Dalmasso

Charrier-Hisamuddin²,

Nguyen

et al. 2008

Gastroenterology

102

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“…16,20,21) It is noted that the activity of hPEPT1 in this cells showed about 35-fold larger than that in Caco-2 cells. [20][21][22] The CHO/hPEPT1 is a powerful tool to study the transport mechanism of hPEPT1. hPEPT1 is an electrogenic H ϩ /dipeptide co-transporter and the substrate of hPEPT1 should cause intracellular acidification.…”

Section: Discussionmentioning

confidence: 99%

“…First, we established stably transfected CHO cells which show high transport activities for various substrates with almost the same K m values as those obtained from those of a model human intestine epithelial cell line, Caco-2. [20][21][22] Thus, this CHO cell line over-expressing hPEPT1 (designated as CHO/hPEPT1) is useful and convenient for transport studies of hPEPT1. In the present study, we then examined the pH-dependency of transport activity by hPEPT1 using CHO/hPEPT1 cells.…”

Section: )mentioning

confidence: 99%

“…The K m value was consistent with those reported for Caco-2 cells, and V max is about 35-times larger than those reported. [20][21][22] Since the gene of hPEPT1 is stably retained in the genome of CHO cells and the transporter express functionally in the membranes, CHO/hPEPT1 cells are very useful and convenient for transport experiments of hPEPT1.…”

Section: Establishment Of a Cell Line Which Stably Overexpresses Hpept1mentioning

confidence: 99%

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The Extracellular pH Dependency of Transport Activity by Human Oligopeptide Transporter 1 (hPEPT1) Expressed Stably in Chinese Hamster Ovary (CHO) Cells: A Reason for the Bell-Shaped Activity versus pH

Fujisawa

Tateoka

Nara

et al. 2006

Biological & Pharmaceutical Bulletin

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It has been estimated that more than 80% of amino acids found in the intestinal lumen after a protein meal are in the form of small peptides rather than free amino acids.1) Therefore, a majority of protein digestion products are obligatorily absorbed via an intestine oligo-peptide transporter called PEPT1. Of interest, PEPT1 also mediates the absorption of a huge number of pharmacologically important compounds including peptidomimetic drugs, 2,3) such as oral b-lactam antibiotics, 4-6) ACE-inhibitors like captopril, 7) or the peptidase inhibitor bestatine 8) and even compounds without an obvious peptide bond or equivalent, 9) such as d-amino levulinic acid.10) This surprisingly high tolerance for structural diversity of the substrate by PEPT1 has been expected to be used as a channel to increase the intestinal absorption of poorly absorbable drugs. 11)The transport of substrates via PEPT1 is coupled to the downward movement of a proton in accordance with its electrochemical gradient. 2,3,12) It has been demonstrated in intestinal membrane vesicles, 4,13,14) Xenopus oocytes 15) and culture cells 16) that the transport activity of PEPT1 shows a bellshaped pH dependence with an optimal pH 5.5 to 6.0. This optimal pH is physiologically collaborated with an acidic unstirred water layer (pH 5.5-6.0) in the intestinal lumen. 1,17) Hence, PEPT1 can function most efficiently in epithelial cells of the small intestine.As for the bell-shaped activity of PEPT1 vs. pH, the reason why the transport activity decreases with an increase in pH is obviously the decrease in the driving force of the proton electrochemical potential gradient. On the other hand, why does the activity decrease in a more acidic region although the driving force should increase? To answer this question, we will try to obtain a relationship between the transport activity and extracellular pH. However, there is a problem to be solved: The driving force changes along with the change in extracellular pH. The pH-dependent properties of PEPT1 should be determined without changing the driving force. Here, we employ the experimental condition that the proton concentration gradient across the membrane is dissipated by the addition of monensin and nigericin 18,19) ; then the driving force is solely the membrane potential. To perform this experiment, a cell line is necessary which shows high transport activity. First, we established stably transfected CHO cells which show high transport activities for various substrates with almost the same K m values as those obtained from those of a model human intestine epithelial cell line, Caco-2. [20][21][22] Thus, this CHO cell line over-expressing hPEPT1 (designated as CHO/hPEPT1) is useful and convenient for transport studies of hPEPT1. In the present study, we then examined the pH-dependency of transport activity by hPEPT1 using CHO/hPEPT1 cells. Human oligopeptide transporter (hPEPT1) translocates di/tri-peptide by coupling to movement of proton down the electrochemical gradient. This transporter has the characteristics ...

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“…On the other hand, either exposure to two immunosuppressive agents, tacrolimus and cyclosporin A (44), or exposure to thyroid hormone (5) or long-term basolateral stimulation with EGF inhibited the uptake of dipeptides via PEPT1 in Caco-2 cells (44). Moreover, it has been demonstrated that activation of signaling pathways that involve protein kinases C and A changes the kinetic properties of PEPT1 and PEPT2 in intestinal and renal cell lines (13,73). Whether these in vitro observations are of physiological importance in vivo in renal tissues remains to be determined.…”

Section: Regulation Of Expression Level and Transport Activity Of Pepmentioning

confidence: 99%

An update on renal peptide transporters

Daniel

Rubio‐Aliaga

2003

American Journal of Physiology-Renal Physiology

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Daniel, Hannelore, and Isabel Rubio-Aliaga. An update on renal peptide transporters. Am J Physiol Renal Physiol 284: F885-F892, 2003; 10.1152/ajprenal.00123.2002The brush-border membrane of renal epithelial cells contains PEPT1 and PEPT2 proteins that are rheogenic carriers for short-chain peptides. The carrier proteins display a distinct surface expression pattern along the proximal tubule, suggesting that initially di-and tripeptides, either filtered or released by surface-bound hydrolases from larger oligopeptides, are taken up by the low-affinity but high-capacity PEPT1 transporter and then by PEPT2, which possesses a higher affinity but lower transport capacity. Both carriers transport essentially all possible di-and tripeptides and numerous structurally related drugs. A unique feature of the mammalian peptide transporters is the capability of proton-dependent electrogenic cotransport of all substrates, regardless of their charge, that is achieved by variable coupling in proton movement along with the substrate down the transmembrane potential difference. This review focuses on the postcloning research efforts to understand the molecular physiology of peptide transport processes in renal tubules and summarizes available data on the underlying genes, protein structures, and transporter function as derived from studies in heterologous expression systems. PEPT1; PEPT2; renal physiology; localization; functional analysis RENAL TUBULAR PEPTIDE TRANSPORT activity was originally discovered after intravenous infusion of the resistant dipeptide Gly-Sar in rats that resulted in a high accumulation of the intact dipeptide in renal tissue (1, 3). Studies with renal brush-border membrane vesicles established that renal apical peptide uptake was an electrogenic, proton-dependent uphill transport process for di-and tripeptides and related drugs mediated by two kinetically different systems (for a review, see Ref. 38). The underlying proteins, differing in transport characteristics, now designated as PEPT1 (SLC15A1) and PEPT2 (SLC15A2), have been identified, and the corresponding genes have been cloned from a variety of species (10,24,25,39,41,(52)(53)(54). The transporters have been expressed in various cellular models, and information on structure, transport function, and regulation has been gathered by flux studies and electrophysiological techniques. Expression analysis of transporter mRNA and protein by in situ hybridization and immunohistochemistry has extended our knowledge of tissue distribution, particularly the renal zonation of PEPT1 and PEPT2 surface expression. THE MOLECULAR ENTITIES OF APICAL PEPTIDE TRANSPORTPEPT1 and PEPT2 are polytopic integral membrane proteins with 12 predicted membrane-spanning domains and NH 2 -and COOH-terminal ends facing the cytosol. Transporter architecture and membrane topology have not been studied systematically, and therefore structural information is still mainly based on hydropathic analysis of amino acid sequences. The mammalian PEPT1 proteins comprise 701-710 amino acids, ...

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Expression and protein kinase C-dependent regulation of peptide/H+ co-transport system in the Caco-2 human colon carcinoma cell line

Cited by 130 publications

References 27 publications

PepT1-Mediated Tripeptide KPV Uptake Reduces Intestinal Inflammation

PepT1-Mediated Tripeptide KPV Uptake Reduces Intestinal Inflammation

The Extracellular pH Dependency of Transport Activity by Human Oligopeptide Transporter 1 (hPEPT1) Expressed Stably in Chinese Hamster Ovary (CHO) Cells: A Reason for the Bell-Shaped Activity versus pH

An update on renal peptide transporters

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