The function of Latexin (LXN) in inflammation has attracted attention. However, no data are available regarding its role in colitis. We report that LXN is a suppressor of colitis. LXN deficiency leads to the severity of colitis in DSS-induced mice, and LXN is required for the therapeutic effect of retinoic acid on colitis. Using a proteomics approach, we demonstrate that LXN interacts and forms a functional complex with HECTD1 (an E3 ubiquitin ligase) and ribosomal protein subunit3 (Rps3). IκBα is one of the substrates of HECTD1. Ectopic expression of LXN leads to IκBα accumulation in intestinal epithelial cells, however, LXN knockdown enhances the interaction of HECTD1 and Rps3, contributing to the ubiquitination degradation of IκBα, and subsequently enhances inflammatory response. Thus, our findings provided a novel mechanism underlying LXN modulates colitis via HECTD1/Rps3/NF-κB pathway and significant implications for the development of novel strategies for the treatment of colitis by targeting LXN.