Translin, a highly conserved DNA/RNA binding protein that forms a hetero-octamer together with Translin-associated factor X (TRAX), possesses a broad variety of functions, including RNA processing and DNA repair. Recent studies have reported that Translin is involved in mesenchymal cell physiology. Thus, here we analyzed the intrinsic role of Translin in mesenchymal cell proliferation and differentiation. Translin-deficient E11.5 mouse embryonic fibroblasts showed enhanced growth. Translin-deficient bone marrow-derived mesenchymal stem cells showed substantial expansion in vivo and enhanced proliferation in vitro. These cells also showed enhanced osteogenic and adipocytic differentiation. Histological analyses showed adipocytic hypertrophy in various adipose tissues. Translin knockout did not affect the growth of subcutaneous white adipose tissue-derived stem cells, but enhanced adipocytic differentiation was observed in vitro. Contrary to previous reports, in vitro-fertilized Translin-null mice were not runted and exhibited normal metabolic homeostasis, indicating the fragility of these mice to environmental conditions. Together, these data suggest that Translin plays an intrinsic role in restricting mesenchymal cell proliferation and differentiation.