The apically localized riboflavin (RF) transporter-3 (RFVT-3) is involved in intestinal absorption of vitamin B2. Previous studies have characterized different physiological/biological aspects of the RFVT-3, but there is a lack of knowledge regarding possible existence of interacting partner(s) and consequence of interaction(s) on its function/cell biology. To address the latter, we performed yeast two-hybrid (Y2H) screening of a human colonic cDNA library and have identified transmembrane protein 237 (TMEM237) as a putative interactor with the human (h)RFVT-3; the interaction was further confirmed via “1-by-1” Y2H assay that involved appropriate positive and negative controls. TMEM237 was found to be highly expressed in human native intestine and in human intestinal epithelial cell lines; further, confocal images showed colocalization of the protein with hRFVT-3. The interaction between TMEM237 with hRFVT-3 in human intestinal epithelial HuTu-80 cells was established by coimmunoprecipitation. Expressing TMEM237 in HuTu-80 cells led to a significant induction in RF uptake, while its knockdown (with the use of gene-specific siRNA) led to a significant reduction in uptake. Transfecting TMEM237 into HuTu-80 cells also led to a marked enhancement in hRFVT-3 protein stability (reflected by an increase in the protein half-life). Interestingly, the level of expression of TMEM237 was found to be markedly reduced following treatment with TNF-α (a proinflammatory cytokine that inhibits intestinal RF uptake), while its expression was significantly upregulated following treatment with butyrate (an inducer of intestinal RF uptake). These findings identify TMEM237 as an interactor with the intestinal hRFVT-3 and show that the interaction has physiological/biological significance.