hPepT1 mediates bacterial tripeptide fMLP uptake in human monocytes

We examined hPepT1 expression in the monocytic cell line, KG‐1. Reverse transcription polymerase chain reaction (RT‐PCR) analysis revealed that hPepT1 is expressed in KG‐1 cells, while cDNA cloning and direct sequencing confirmed the sequence of KG‐1 hPept1. Immunoblotting of KG‐1 cell lysates revealed a ~100 kDa immunoreactive band mainly present in the membrane fraction. Uptake experiments showed that the transport of 20 μM radiolabeled Gly‐Sarcosine ([ 14 C]Gly‐Sar) in KG‐1 cells was Na + , Cl − dependent and disodium 4,4′‐diisothiocyanatostilbene‐2,2′‐disulfonate (DIDS)‐sensitive. In addition, hPepT1 activity was directly coupled to NHE3, as evidenced by the fact that [ 14 C]Gly‐Sar uptake was not affected by the absence of Na + when cells were incubated at low pH (5.2). Interestingly, hPepT1‐mediated di‐ and tripeptides were reduced in KG‐1 cells incubated at low pH, due to the non‐polar nature of KG‐1. Finally, we showed that hPepT1 is responsible for transporting fMLP into undifferentiated and differentiated (macrophage‐like) KG‐1 cells. Together, these results show that hPepT1 is expressed in non‐polarized immune cells such as macrophages, where the transporter functions best at the immune physiological level of pH 7.2. Furthermore, we provide evidence for hPepT1‐mediated fMLP transport, which might form a novel immune cell activation pathway during intestinal inflammation.