Brown adipocytes oxidize fatty acids to produce heat in response to cold or 
caloric overfeeding. The motivation and function of the development of brown fat 
may thus counteract obesity, though this remains uncertain. We investigated the 
brown adipocyte proteome by two-dimensional gel electrophoresis followed by mass 
spectrometry. Comparative analyses of proteins focused on total protein spots to 
filter differentially expressed proteins during the differentiation of mouse 
primary brown preadipocytes. A Western blot analysis was performed to verify the 
target proteins. The results indicated that 10 protein spots were differentially 
expressed with significant changes, including the three up-regulated proteins of 
prohibitin, hypoxanthine-guanine phosphoribosyltransferase, and enoyl-CoA 
hydratase protein; the 5 down-regulated proteins of triosephosphate isomerase, 
elongation factor 2, α-tropomyosin slow, endophilin-B1, and cofilin-1 (CFL1); 
and the two unequivocally expressed proteins of peroxiredoxin-1 and collagen 
α-1(i) chain precursor. We found that during brown adipogenesis, CFL1 has an 
inhibitory effect on brown adipocyte differentiation. The overexpression of CFL1 
inhibited the brown fat deposition and repressed the brown marker genes UCP1, 
PRDM16, PGC-1α and PPARγ via actin dynamics and polymerization. These 
observations may be novel findings that bring new insight into the detailed 
mechanisms of brown adipogenesis and identify possible therapeutic targets for 
anti-obesity.
BIOLOGICAL SIGNIFICANCE: We use 2-DE to compare the proteomes of adipocytes 
during the brown adipogenesis of primary mouse preadipocytes. We identified 10 
proteins that are differentially expressed. Among these, we found that the actin 
binding protein CFL1 inhibits the differentiation of brown preadipocytes. CFL1 
overexpressing cells showed lower deposition of brown fat droplets, and the 
brown marker genes of UCP1, PRDM16, PGC-1α and PPARγ were decreased through 
actin dynamics and polymerization.