Identification of the major diacylglycerol acyltransferase mRNA in mouse adipocytes and macrophages

Authors: Cao, Heping

Submitted to: BMC Biochemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/6/2018
Publication Date: 12/14/2018
Citation: Cao, H. 2018. Identification of the major diacylglycerol acyltransferase mRNA in mouse adipocytes and macrophages. BMC Biochemistry. 19(11):1-11. https://doi.org/10.1186/s12858-018-0103-y.
DOI: https://doi.org/10.1186/s12858-018-0103-y

Interpretive Summary: Triacylglycerols (TAGs) are the primary form of energy storage in eukaryotes. They also serve as a reservoir of fatty acids for membrane biogenesis of the cells and lead to obesity when excessively accumulated in adipose tissues. Diacylglycerol acyltransferases (DGATs) are responsible for the final and rate-limiting step of TAG biosynthesis in eukaryotic organisms. Understanding the roles of DGATs will help to create transgenic plants and microbes with value-added properties and provide information for therapeutic intervention for obesity and related diseases. It is generally accepted that DGATs are divided into DGAT1 and DGAT2 subfamilies in animals but it was not clear which form of DGAT was the major isoform expressed in mammalian cells. The objective of this study was to identify the major form of DGAT expressed in mammalian cells. TaqMan and SYBR Green quantitative real-time PCR assays were used to evaluate DGAT mRNA levels in mouse adipocytes and macrophages and tung tree seeds. Our results demonstrated that DGAT2 was the major form of DGATs expressed in eukaryotic cells. This conclusion was supported by results from three different cell types (mouse adipocytes, mouse macrophages, tung tree seeds), from different time-course ranging from 30 min to 24 h in animal cells and 1-11 weeks in developing seeds. Our results also showed that DGAT gene expression was much higher in lipid-synthesizing adipocytes than that in macrophages, which is in contrast to the expression patterns of TTP, an anti-inflammatory gene, in these two cell types. Our results have conclusively demonstrated that DGAT2 mRNA is the major form of DGATs expressed in both plant and animal cells. This finding should be useful for targeted approach to regulating lipid/oil accumulation in plant and animal tissues.

Technical Abstract: Triacylglycerols (TAGs) are the primary form of energy storage in eukaryotes. They also serve as a reservoir of fatty acids for membrane biogenesis of the cells and lead to obesity when excessively accumulated in adipose tissues. Diacylglycerol acyltransferases (DGATs) are responsible for the final and rate-limiting step of TAG biosynthesis in eukaryotic organisms. Mammalian DGATs are divided into DGAT1 and DGAT2 subfamilies. It was unclear which DGAT was the major isoform expressed in animals. The objective of this study was to identify the major form of DGATs expressed in animal cells and compared to those expressed in tung tree seeds. TaqMan and SYBR Green qPCR assays evaluated DGAT mRNA levels in mouse 3T3-L1 adipocytes and RAW264.7 macrophages and tung tree seeds. TaqMan qPCR showed that DGAT mRNA levels were stable during the time-course study, DGAT2 mRNA levels were 10-30 folds higher than DGAT1 in adipocytes and macrophages, and DGAT mRNA levels in adipocytes were 50-100 folds higher than those in macrophages. As a positive control, the anti-inflammatory tristetraprolin (TTP) mRNA levels were 2-4 folds higher in macrophages than those in adipocytes and similar to DGAT1 in adipocytes but 100-fold higher than DGAT1 in macrophages. SYBR Green qPCR analyses confirmed TaqMan qPCR results. DGAT2 mRNA as the major DGAT mRNA in mouse cells was similar to that in tung tree seeds where DGAT2 mRNA levels were 10-20-fold higher than DGAT1 or DGAT3. Our results conclusively demonstrated that DGAT2 mRNA was the major form of DGATs expressed in plant and animal cells.