Adipose tissue affects metabolic-related diseases as it consists of various types of cells that are involved in fat metabolism and adipokine release. CXC ligand 5 (CXCL5) is a member of the CXC chemokine family and is highly expressed by macrophages in white adipose tissue (WAT). In this study, we generated and investigated the function of Cxcl5 in knockout (KO) mouse. Cxcl5 KO mouse was generated using CRISPR/Cas9. The KO male mice didn’t show significant phenotype difference in normal condition. However, proteomic analysis revealed that many proteins are enriched in the process of fatty acid beta-oxidation and mitochondrial localization in iWAT of Cxcl5 KO mice. Cxcl5 KO mouse showed rather decreased proteins and transcripts expression associated with thermogenesis including uncoupled protein 1 (UCP1), a well-known thermogenic gene, and increased expressions associated with inflammation. The increase of Ucp1 expression in cold conditions was significantly retarded in Cxcl5 KO mice. The expression of transcription factors involved in thermogenesis including peroxisome proliferator-activated receptor gamma coactivator 1-alpha (Pgc1α) was not properly increased in Cxcl5 KO under cold challenge and its mimic conditions. CXCL5 treatment increased the expression of transcription factors for Ucp1 and Ucp1 itself. Collectively, our data shows that Ucp1 expression is induced in adipocyte by CXCL5 secreted by β-Adrenergic stimulation in M1 macrophages. Our data indicates that CXCL5 has a crucial role in regulating energy metabolism, particularly in cold exposure. These results strongly suggest that targeting CXCL5 could be a therapeutic aim for people with a disorder of energy metabolism.