C/EBPα mediates the transcriptional suppression of human calreticulin gene expression by TNFα

Calreticulin (CRT), a 46 kDa endoplasmic reticulum chaperone, is critical in the folding and quality control of proteins. However, the mechanisms of its regulation are not fully understood. Our previous study had demonstrated that elevated TNF alpha levels that are hallmarks of diverse metabolic diseases negatively regulate cellular CRT levels. Here, we attempted to study the mode of this regulation of CRT by TNF alpha. Using luciferase reporter deletion constructs of the CRT promoter, we demonstrate that while the 2 kb and 1 kb promoter constructs depict comparable activities, the activity of the 0.5 kb region was greatly reduced suggesting the significance of the region between 1.0 kb and 0.5 kb during CRT promoter activity. Of the transcription factors that possess binding elements within this region, C/EBP alpha was prioritized since it was shown to be inhibited by TNF alpha in an earlier report from our laboratory. TNF alpha significantly inhibited the wild-type CRT promoter activity that was attenuated in a C/EBP alpha-deleted construct. C/EBP alpha mRNA levels and its nuclear content was also reduced in the presence of TNF alpha. This led to reduced C/EBP alpha occupancy on the CRT promoter and a decreased binding of the nuclear protein to the C/EBP alpha-consensus sequence. TNF alpha also reduced the nuclear content of C/EBP beta but it did not bind to the CRT promoter suggesting that it does not contribute to the inhibitory effect of TNF alpha. To conclude, our results suggest that C/EBP alpha is critical in mediating the inhibitory effect of TNF alpha on CRT expression that might be crucial in determining the deleterious cellular effects of TNF alpha. (C) 2011 Elsevier Ltd. All rights reserved.