Publications

• Human, rat and chicken small intestinal Na+ - Cl- -creatine transporter: functional, molecular characterization and localization.

  Peral M.J., Garcia-Delgado M., Calonge M.L., Duran J.M., De La Horra M.C., Wallimann T., Speer O., Ilundain A.

  In spite of all the fascinating properties of oral creatine supplementation, the mechanism(s) mediating its intestinal absorption has(have) not been investigated. The purpose of this study was to characterize intestinal creatine transport. [(14)C] creatine uptake was measured in chicken enterocyte ... >> More

  In spite of all the fascinating properties of oral creatine supplementation, the mechanism(s) mediating its intestinal absorption has(have) not been investigated. The purpose of this study was to characterize intestinal creatine transport. [(14)C] creatine uptake was measured in chicken enterocytes and rat ileum, and expression of the creatine transporter CRT was examined in human, rat and chicken small intestine by reverse transcription-polymerase chain reaction, Northern blot, in situ hybridization, immunoblotting and immunohistochemistry. Results show that enterocytes accumulate creatine against its concentration gradient. This accumulation was electrogenic, Na(+)- and Cl(-)-dependent, with a probable stoichiometry of 2 Na(+): 1 Cl(-): 1 creatine, and inhibited by ouabain and iodoacetic acid. The kinetic study revealed a K(m) for creatine of 29 microM. [(14)C] creatine uptake was efficiently antagonized by non-labelled creatine, guanidinopropionic acid and cyclocreatine. More distant structural analogues of creatine, such as GABA, choline, glycine, beta-alanine, taurine and betaine, had no effect on intestinal creatine uptake, indicating a high substrate specificity of the creatine transporter. Consistent with these functional data, messenger RNA for CRT was detected only in the cells lining the intestinal villus. The sequences of partial clones, and of the full-length cDNA clone, isolated from human and rat small intestine were identical to previously cloned CRT cDNAs. Immunological analysis revealed that CRT protein was mainly associated with the apical membrane of the enterocytes. This study reports for the first time that mammalian and avian enterocytes express CRT along the villus, where it mediates high-affinity, Na(+)- and Cl(-)-dependent, apical creatine uptake. << Less

  J. Physiol. (Lond.) 545:133-144(2002) [PubMed] [EuropePMC]

• Creatine transport and creatine kinase activity is required for CD8pisup>+pi/sup> T cell immunity.

  Samborska B., Roy D.G., Rahbani J.F., Hussain M.F., Ma E.H., Jones R.G., Kazak L.

  The factors that promote T cell expansion are not fully known. Creatine is an abundant circulating metabolite that has recently been implicated in T cell function; however, its cell-autonomous role in immune-cell function is unknown. Here, we show that creatine supports cell-intrinsic CD8<sup>+</s ... >> More

  The factors that promote T cell expansion are not fully known. Creatine is an abundant circulating metabolite that has recently been implicated in T cell function; however, its cell-autonomous role in immune-cell function is unknown. Here, we show that creatine supports cell-intrinsic CD8⁺ T cell homeostasis. We further identify creatine kinase B (CKB) as the creatine kinase isoenzyme that supports these T cell properties. Loss of the creatine transporter (Slc6a8) or Ckb results in compromised CD8⁺ T cell expansion in response to infection without influencing adenylate energy charge. Rather, loss of Slc6a8 or Ckb disrupts naive T cell homeostasis and weakens TCR-mediated activation of mechanistic target of rapamycin complex 1 (mTORC1) signaling required for CD8⁺ T cell expansion. These data demonstrate a cell-intrinsic role for creatine transport and creatine transphosphorylation, independent of their effects on global cellular energy charge, in supporting CD8⁺ T cell homeostasis and effector function. << Less

  Cell Rep 38:110446-110446(2022) [PubMed] [EuropePMC]

• Molecular characterization of the human CRT-1 creatine transporter expressed in Xenopus oocytes.

  Dai W., Vinnakota S., Qian X., Kunze D.L., Sarkar H.K.

  The protein sequence encoded by a creatine transporter cDNA cloned from a human heart library was identical to that cloned from a human kidney library (Nash et al., Receptors Channels 2, 165-174, 1994), except that at position 285 the former contained an Ala residue and the latter contained a Pro ... >> More

  The protein sequence encoded by a creatine transporter cDNA cloned from a human heart library was identical to that cloned from a human kidney library (Nash et al., Receptors Channels 2, 165-174, 1994), except that at position 285 the former contained an Ala residue and the latter contained a Pro residue. Expression of this human heart cDNA clone in Xenopus laevis oocytes induced a Na+- and Cl--dependent creatine uptake activity that saturated with a Km of approximately 20 microM for creatine. The induced uptake was inhibited by beta-guanidinopropionic acid (IC50 approximately 44.4 microM), 2-amino-1-imidazolidineacetic acid (cyclocreatine; IC50 approximately 369.8 microM), gamma-guanidinobutyric acid (IC50 approximately 697.9 microM), gamma-aminobutyric acid (IC50 approximately 6.47 mM), and amiloride (IC50 approximately 2.46 mM). The inhibitors beta-guanidinopropionic acid, cyclocreatine, and gamma-guanidinobutyric acid also inhibited the uptake activity of the Ala285 to Pro285 (A285P) mutant as effectively as that of the wild type. In contrast, guanidinoethane sulfonic acid, a potent inhibitor of taurine transport, inhibited the uptake activity of the A285P mutant approx. two times more effectively than that of the wild type. The protein kinase C activator phorbol 12-myristate 13-acetate (PMA), but not its inactive analog, 4alpha-phorbol 12, 13-didecanoate, inhibited the creatine uptake, and the inhibitory effect of PMA was both time and concentration dependent. The protein kinase A activator 8-bromo-cyclic AMP, however, had no effect on the creatine uptake. The rate of uptake increased hyperbolically with the increasing concentration of the external Cl- (equilibrium constant KCl-approximately 5 mM) and sigmoidally with the increasing concentration of the external Na+ (equilibrium constant KNa+ approximately 56 mM). Further analyses of the Na+ and Cl-concentration dependence data suggested that at least two Na+ and one Cl-were required to transport one creatine molecule via the creatine transporter. << Less

  Arch. Biochem. Biophys. 361:75-84(1999) [PubMed] [EuropePMC]

• Functional and electrophysiological characterization of four non-truncating mutations responsible for creatine transporter (SLC6A8) deficiency syndrome.

  Valayannopoulos V., Bakouh N., Mazzuca M., Nonnenmacher L., Hubert L., Makaci F.L., Chabli A., Salomons G.S., Mellot-Draznieks C., Brule E., de Lonlay P., Toulhoat H., Munnich A., Planelles G., de Keyzer Y.

  Intellectual disability coupled with epilepsy are clinical hallmarks of the creatine (Cr) transporter deficiency syndrome resulting from mutations in the SLC6A8 gene. So far characterization of pathogenic mutations of SLC6A8 has been limited to Cr uptake. The aim of our study was to characterize t ... >> More

  Intellectual disability coupled with epilepsy are clinical hallmarks of the creatine (Cr) transporter deficiency syndrome resulting from mutations in the SLC6A8 gene. So far characterization of pathogenic mutations of SLC6A8 has been limited to Cr uptake. The aim of our study was to characterize the electrogenic and pharmacological properties of non truncating SLC6A8 mutations identified in patients presenting variable clinical severity. Electrophysiological and pharmacological properties of four mutants (including two novel ones) were studied in X. laevis oocyte expression system. Creatine uptake was assessed with [(14)C]-Cr in X. laevis and patients' fibroblasts. Subcellular localization was determined by immunofluorescence and western blot. All mutants were properly targeted to the plasma membrane in both systems. Mutations led to the complete loss of both electrogenic and transport activities in X. laevis and Cr uptake in patients' fibroblasts. Among the Cr analogs tested, guanidinopropionate induced an electrogenic activity with the normal SLC6A8 transporter similar to creatine whereas a phosphocreatine derivative, PCr-Mg-CPLX, resulted in partial activity. SLC6A8 mutants displayed no electrogenic activity with all Cr analogs tested in X. laevis oocytes. Although the mutations altered various domains of SLC6A8 Cr uptake and electrogenic properties were completely inhibited and could not be dissociated. Besides the metabolic functions of Cr, the loss of SLC6A8 electrogenic activity, demonstrated here for the first time, may also play a role in the altered brain functions of the patients. << Less

  J. Inherit. Metab. Dis. 36:103-112(2013) [PubMed] [EuropePMC]