Adipose tissue is critical in the development of Metabolically unhealthy obesity (MUO). This study aimed to investigate the different responses and regulations of subcutaneous (SA) and visceral adipose tissue (VA) to MUO by integrative proteome and lysine acetylation (Kac) analyses. This study significantly expanded the known set of proteins and acetylated proteins by reporting the differential expression and Kac modifications in VA and SA in MUO for the first time, thereby establishing a foundation for exploring the pathogenesis and treatment of MUO.