LIGHT is a member of tumor necrosis factor (TNF) superfamily, and its function is mediated through lymphotoxin-β receptor (LTβR), which is known to play important roles in inflammatory and immune responses through activation of NF-κB signaling pathways. However, molecular mechanism of LTβR ligation-induced NF-κB signaling remains incompletely understood. In this report we demonstrate that a novel zinc-finger protein 91 (ZFP91) is a critical regulator in LIGHT-induced activation of non-canonical NF-κB pathway. ZFP91 appears to be required for NF-κB2 (p100) processing to p52, nuclear translocation of p52 and RelB, and DNA-binding activity of NF-κB in LIGHT-induced activation of non-canonical NF-κB pathway. Furthermore, ZFP91 knock-down by RNA interference blocks the LIGHT-induced accumulation of NIK and p100 processing, as well as the expression of non-canonical NF-κB target genes. These data clearly indicate that ZFP91 is a key regulator in LIGHT-induced activation of non-canonical NF-κB pathway in LTβR signaling.