Test system for studying biotin transport upon <i>SLC5A6</i> gene inactivation

This paper introduces a test system for the investigation of biotin transport following inactivation of the <i>SLC5A6</i> gene, which encodes the sodium-dependent multivitamin transporter SLC5A6. The aim was to develop a method for assessing the efficiency of biotin penetration across the cell membrane following inactivation of the <i>SLC5A6</i> gene and to explore the feasibility of delivering biotin derivatives into cells independent of SLC5A6. The test system is built upon modified HEK293 cell lines with overexpression of the BirA* biotin ligase, with the first line comprising a functional <i>SLC5A6</i> gene and the second one involving an inactivated version of this gene mimicking impaired biotin transport. This test system was used to investigate the transport of biotin and its two derivatives, namely the biotin conjugate with <i>p</i>-aminophenylalanine (Bio-1) and biotin methyl ester (Bio-2), through the cell membrane. It has been determined that biotin and its methyl ester (Bio-2) can enter cells independently of the SLC5A6 transporter, which points to the presence of alternative transport pathways. The biotin derivative Bio-1, which contains <i>p</i>-aminophenylalanine, is internalized into cells solely through the hSMVT transporter. The novel test system will serve as a tool for investigating the pathways involved in vitamin entry into cells and for developing therapeutic strategies for individuals with mutations in the <i>SLC5A6</i> gene, as well as other transport-related genes.