DHFR2 is part of the large Dihydrofolate reductase gene family, in which DHFR is the mainly characterised member. For a long time, DHFR was considered the only gene of the family able to make a functional protein, with the remaining members catalogued as pseudogenes. DHFR has been thoroughly studied, considering its crucial role in facilitating the entry of folic acid in the One-Carbon Metabolism pathway, and its implications in neural tube defects and cancer. In the past decade, DHFRP4 was renamed DHFR2 and classified as a protein-coding gene since it harbours an intact open reading frame that can result in a protein, in vitro. However, no information was available in the literature about the endogenous DHFR2 protein, its enzymatic activity, or cellular localisation.