Disruption of transport activity in a D93H mutant thiamine transporter 1, from a Rogers Syndrome family

Abstract

Rogers syndrome is an autosomal recessive disorder resulting in megaloblastic anemia, diabetes mellitus, and sensorineural deafness. The gene associated with this disease encodes for thiamine transporter 1 (THTR1), a member of the SLC19 solute carrier family including THTR2 and the reduced folate carrier (RFC). Using transient transfections into NIH3T3 cells of a D93H mutant THTR1 derived from a Rogers syndrome family, we determined the expression, post-translational modification, plasma membrane targeting and thiamine transport activity. We also explored the impact on methotrexate (MTX) transport activity of a homologous missense D88H mutation in the human RFC, a close homologue of THTR1. Western blot analysis revealed that the D93H mutant THTR1 was normally expressed and underwent a complete N-glycosylation. However, while this mutant THTR1 was targeted to the plasma membrane, it was completely devoid of thiamine transport activity. Consistently, introduction into MTX transport null cells of a homologous D88H mutation in the hRFC did not result in restoration of MTX transport activity, thereby suggesting that D88 is an essential residue for MTX transport activity. These results suggest that the D93H mutation does not interfere with transporter expression, glycosylation and plasma membrane targeting. However, the substitution of this negatively charged amino acid (Asp93) by a positively charged residue (His) in an extremely conserved region (the border of transmembrane domain 2/intracellular loop 2) in the SLC19 family, presumably inflicts deleterious structural alterations that abolish thiamine binding and/or translocation. Hence, this functional characterization of the D93H mutation provides a molecular basis for Rogers syndrome.

An exogenous retrovirus isolated from koalas with malignant neoplasias in a US zoo

“…To investigate whether THTR1 requires retention of its ability to transport thiamine to facilitate KoRV-B entry, we used a point mutant of THTR1 (D93H) that lacks the thiamine transporter function but traffics to the cell membrane like the native protein (12). This mutant transporter retains KoRV-B receptor properties, albeit at a reduced efficiency, when expressed in KoRV-B-resistant murine cells (Table 2).…”

The Impact of Thiamine Treatment in the Diabetes Mellitus

“…Thiamine-responsive megaloblastic anemia (TRMA) is a rare autosomal recessive condition, characterized by megaloblastic anemia, non-autoimmune DM, and sensory-neural loss [12-13]. TRMA fibroblasts displayed only 5-10% of thiamine uptake when compared with healthy individuals [14].…”