Accessory Protein TM4SF4 Interacts with hTHTR-2 to Influence Thiamine Uptake

08/10/2021 - The human thiamine transporter-2 (hTHTR-2) plays a key role in the intestinal absorption of thiamine (vitamin B1). Recent studies with membrane transporters of other nutrients/substrates have shown that associated proteins affect different aspects of cell physiology and cell biology. 

Researchers currently have no good information on the protein(s) that interact with hTHTR-2 in intestinal epithelial cells, and how those proteins may influence cell function and/or cell biology.

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A team of researchers recently set out to identify protein partner(s) that interacts with hTHTR-2 in human intestinal cells and determine the physiological/biological consequence of that interaction. The research team included Veedamali S. Subramanian, Svetlana M. Nabokina, and Hamid M. Said. They are affiliated with the Departments of Medicine, Physiology and Biophysics, University of California in Irvine, CA; and the Department of Veterans Affairs Medical Center in Long Beach, CA.

Their team used the yeast split-ubiquitin two-hybrid approach to screen a human intestinal cDNA library. They followed with GST-pull-down and cellular co-localization approaches to confirm the interaction between hTHTR-2 and the associated protein(s). They used  3H-thiamine uptake assays to assess the effect of such an interaction on hTHTR-2 function.

Their assessment showed that the human TransMembrane 4 SuperFamily 4 (TM4SF4) is a potential inter-actor with hTHTR-2. To confirm the interaction, they used in vitro GST-pull-down assay, live-cell confocal imaging of HuTu-80 cells co-expressing hTHTR-2-GFP and mCherry-TM4SF4 (the latter of which showed a substantial overlap of these two proteins in intracellular vesicles and at the cell membrane). 

They found that co-expression of hTHTR-2 with TM4SF4 in HuTu-80 cells triggered a sharp rise in thiamine uptake. Conversely, silencing TM4SF4 with gene-specific siRNA led to a sharp decrease in thiamine uptake.

The team's results offer the first proof that the accessory protein TM4SF4 interacts with hTHTR-2 and influences the physiological function of the thiamine transporter.

This finding could help researchers to better understand how certain proteins influence cell function and/or cell biology.

Read more in Dig Dis Sci. 2014 Mar; 59(3): 583–590.Published online 2013 Nov 27.