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Synergistic role of specificity proteins and upstream stimulatory factor 1 in transactivation of the mouse carboxylesterase 2/microsomal acylcarnitine hydrolase gene promoter.

ABSTRACT: Mouse carboxylesterase 2 (mCES2), a microsomal acylcarnitine hydrolase, is thought to play some important roles in fatty acid (ester) metabolism, and it is therefore thought that the level of transcription of the mCES2 gene is under tight control. Examination of the tissue expression profiles revealed that mCES2 is expressed in the liver, kidney, small intestine, brain, thymus, lung, adipose tissue and testis. When the mCES2 promoter was cloned and characterized, it was revealed that Sp1 (specificity protein 1) and Sp3 could bind to a GC box, that USF (upstream stimulatory factor) 1 could bind to an E (enhancer) box, and that Sp1 could bind to an NFkappaB (nuclear factor kappaB) element in the mCES2 promoter. Co-transfection assays showed that all of these transcription factors contributed synergistically to transactivation of the mCES2 promoter. Taken together, our results indicate that Sp1, Sp3 and USF1 are indispensable factors for transactivation of the mCES2 gene promoter. To our knowledge, this is the first study in which transcription factors that interact with a CES2 family gene have been identified. The results of the present study have provided some clues for understanding the molecular mechanisms regulating mCES2 gene expression, and should be useful for studies aimed at elucidation of physiological functions of mCES2.

SUBMITTER: Furihata T

PROVIDER: S-EPMC1134093 | biostudies-literature | 2004 Nov

REPOSITORIES: biostudies-literature

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Synergistic role of specificity proteins and upstream stimulatory factor 1 in transactivation of the mouse carboxylesterase 2/microsomal acylcarnitine hydrolase gene promoter.

Furihata Tomomi T Hosokawa Masakiyo M Satoh Tetsuo T Chiba Kan K

The Biochemical journal 20041101 Pt 1

Mouse carboxylesterase 2 (mCES2), a microsomal acylcarnitine hydrolase, is thought to play some important roles in fatty acid (ester) metabolism, and it is therefore thought that the level of transcription of the mCES2 gene is under tight control. Examination of the tissue expression profiles revealed that mCES2 is expressed in the liver, kidney, small intestine, brain, thymus, lung, adipose tissue and testis. When the mCES2 promoter was cloned and characterized, it was revealed that Sp1 (specif ...[more]

PMID: 15283701

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Project description:BACKGROUND: Alternative splicing of the locus A beta H-J-J generates functionally distinct proteins: the enzyme aspartyl (asparaginyl) beta-hydroxylase (AAH), truncated homologs of AAH with a role in calcium homeostasis humbug and junctate and a structural protein of the sarcoplasmic reticulum membranes junctin. AAH and humbug are over expressed in a broad range of malignant neoplasms. We have previously reported that this locus contains two promoters, P1 and P2. While AAH and humbug are expressed in most tissues under the regulation of the P1 promoter, AAH, junctin and junctate are predominantly expressed in excitable tissues under the control of the P2 promoter. We previously demonstrated that Sp transcription factors positively regulate the P1 promoter. RESULTS: In the present study, we extended the functional characterization of the P1 promoter of the A beta H-J-J locus. We demonstrated by quantitative Real-time RT-PCR that mRNAs from the P1 promoter are actively transcribed in all the human cell lines analysed. To investigate the transcription mechanism we transiently transfected HeLa cells with sequentially deleted reporter constructs containing different regions of the -661/+81 P1 nucleotide sequence. Our results showed that (i) this promoter fragment is a powerful activator of the reporter gene in HeLa cell line, (ii) the region spanning 512 bp upstream of the transcription start site exhibits maximal level of transcriptional activity, (iii) progressive deletions from -512 gradually reduce reporter expression. The region responsible for maximal transcription contains an E-box site; we characterized the molecular interactions between USF1/2 with this E-box element by electrophoretic mobility shift assay and supershift analysis. In addition, our USF1 and USF2 chromatin immunoprecipitation results demonstrate that these transcription factors bind the P1 promoter in vivo. A functional role of USF1/USF2 in upregulating P1-directed transcription was demonstrated by analysis of the effects of (i) in vitro mutagenesis of the P1/E-box binding site, (ii) RNA interference targeting USF1 transcripts. CONCLUSION: Our results suggest that USF factors positively regulate the core of P1 promoter, and, together with our previously data, we can conclude that both Sp and USF DNA interaction and transcription activity are involved in the P1 promoter dependent expression of AAH and humbug.

Dataset Information

Synergistic role of specificity proteins and upstream stimulatory factor 1 in transactivation of the mouse carboxylesterase 2/microsomal acylcarnitine hydrolase gene promoter.

Publications

Synergistic role of specificity proteins and upstream stimulatory factor 1 in transactivation of the mouse carboxylesterase 2/microsomal acylcarnitine hydrolase gene promoter.

Similar Datasets

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