BubR1 mitotic checkpoint kinase monitors attachment of microtubules to kinetochores and links regulation of the chromosome-spindle attachment to mitotic checkpoint signaling. Defects in BubR1-mediated signaling severely perturb checkpoint control and are linked to diseases such as cancer. Studies using BubR1 mouse models suggest that BubR1 activities prevent premature aging and infertility. In this study, we show that BubR1 depletion in human adipose-derived mesenchymal stem cells (ASCs) precedes loss of the differentiation potential and induction of replicative senescence. These effects occur independently of p16(INK4A) expression and may involve DNA methylation. Our results reveal a new and unsuspected feature of BubR1 expression in regulation of adult stem cell differentiation.