Oligopeptide/Histidine Transporter PHT1 and PHT2 — Function, Regulation, and Pathophysiological Implications Specifically in Immunoregulation

Functional Characterization of the Lysosomal Peptide/Histidine Transporter PHT1 (SLC15A4) by Solid Supported Membrane Electrophysiology (SSME)

“…Previous studies have attempted to characterize the functional properties of PHT1 (SLC15A4) but the results are inconsistent [2,3,7,8,10,11]. One of the reasons for this variability is the intracellular location of PHT1, which has hindered the interpretation of the results obtained with standard cell- based transport assays, designed for transporters expressed in the plasma membranes [10,11].…”

“…To assess the substrate selectivity of PHT1, sensors loaded with PHT1-GFP OE or WT lysosomal membrane preparations were perfused with a series of AAs (L-histidine, L-alanine, L-leucine, Larginine, L-methionine and L-lysine), di-and tripeptides (Gly-Gly, Gly-Sar, Gly-Gly-Gly, His-leu and Leu-Leu) and peptidomimetic drugs (captopril and lisinopril), some of which have been previously proposed as PHT1 substrates [3,7,8], and Peakon currents were recorded with the SURFER 2 R N1 device. Interestingly, when comparing the PHT1-GFP OE and WT Peakon currents induced by AAs (Figure 2A), only the positively charged AAs (L-histidine, L-arginine, L-lysine) induced significant differences, while the uncharged AAs did not induce any current (L-alanine) or were the same for both lysosomal membrane preparations (L-leucine and L-methionine).…”

“…To date, five different membrane transporters encoded by the SLC15 family have been identified. The oligopeptide transporters PepT1 (SLC15A1) and PepT2 (SLC15A2) are mainly expressed in the apical membranes of the small intestine and kidney proximal tubules, respectively, and their physiological roles have been well characterized [3]. In contrast, peptide/histidine transporters PHT2 (SLC15A3) and PHT1 (SLC15A4) are expressed in endosomal and lysosomal membranes of immune cells, where they play essential roles in metabolic and inflammatory signaling events [4].…”

“…PHT1 (SLC15A4) was cloned from a rat brain cDNA library [7], while the human homologue was cloned from an intestinal cDNA library and Caco-2 cells [12]. The human SLC15A4 gene encodes a 577-amino acid membrane protein whose expression has been found primarily in brain, intestine, some tumor cells and a variety of immune cells, including monocytes/macrophages and dendritic cells [3]. PHT1 has been shown by immunofluorescence to be localized in lysosomes and late endosomes [13], where it plays a pivotal role in lysosomal activity by controlling the levels of certain amino acids (AAs) and oligopeptides and/or by scaffolding various signaling molecules involved in different metabolic and inflammatory events [4].…”

“…In this context, dysregulation of lysosomal pH due to PHT1 dysfunction has been associated with impaired Toll-like receptors (TLRs) mediated type I interferon (IFN-I) production [13,14], and also, with a failure of the IFN regulatory factor 7 (IRF7)-IFN-I regulatory circuit due to disruption of the mammalian target of rapamycin (mTOR) pathway [15]. In addition, PHT1 mediated transport of bacterial oligopeptides, such as MDP or Tri-DAP, triggers the activation of nuclear factor-kappa-B (NF-ΚƁ), which modulates the NOD-dependent immune response [3]. In terms of scaffolding activity, it has been shown that PHT1 interacts with Ragulator components such as LAMTOR1 and LAMTOR2, suggesting that it may be a component of the mTORC1 supercomplex [16].…”