Glioma results from unregulated expansion of a self-renewing glioma-initiating cell population. The regulatory pathways which are essential for sustaining the self-renewal of glioma-initiating cells remain largely unknown. Cell surface N-linked oligosaccharides play functional roles in determining cell fate and are associated with glioma malignancy. Previously, we have reported that beta 1,4-galactosyltransferase V (beta 1,4GalT V) effectively galactosylates the GlcNAc beta 1-->6Man arm of the highly branched N-glycans and positively regulates glioma cell growth. Here, we show that decreasing the expression of beta 1,4GalT V by RNA interference in glioma cells attenuated the formation of polylactosamine and inhibited the ability of tumor formation in vivo. Down-regulation of beta 1,4GalT V depleted CD133-positive cells in glioma xenograft, and inhibited the self-renewal capacity and the tumorigenic potential of glioma-initiating cells. These data reveal a critical role of beta 1,4GalT V in the self-renewal and tumorigenicity of glioma-initiating cells, and indicate that manipulating beta 1,4GalT V expression may have therapeutic potential for the treatment of malignant glioma. (C) 2010 Elsevier Inc. All rights reserved.