Keratinocytes undergo apoptosis in a variety of physiological and pathological conditions. Galectin-3 is a member of a family of beta-galactoside-binding animal lectins expressed abundantly in keratinocytes and other epithelial cells. Here, we have studied the regulatory role of galectin-3 in keratinocyte apoptosis by using cells from gene-targeted galectin-3 null (gal3(-/-)) mice. We showed that galectin-3 mRNA was transiently upregulated in ultraviolet-B (UVB)-irradiated wild-type keratinocytes. We found that gal3(-/-) keratinocytes were significantly more sensitive to apoptosis induced by UVB as well as various other stimuli, both in vitro and in vivo, than wild-type cells. Moreover, we demonstrated that increased apoptosis in gal3(-/-) keratinocytes was attributable to higher extracellular signal-regulated kinase (ERK) activation and lower AKT activation after UVB irradiation. We conclude that endogenous galectin-3 is an anti-apoptotic molecule in keratinocytes functioning by suppressing ERK activation and enhancing AKT activation and may play a role in the development of apoptosis-related skin diseases.