Testis expression of hormone-sensitive lipase is conferred by a specific promoter that contains four regions binding testicular nuclear proteins.
The testicular isoform of hormone-sensitive lipase (HSLtes) is encoded by a testis-specific exon and 9 exons common to the testis and adipocyte isoforms. In mouse, HSLtes mRNA appeared during spermiogenesis in round spermatids. Two constructs containing 1.4 and 0.5 kilobase pairs (kb) of the human HSLtes gene 5'-flanking region cloned upstream of the chloramphenicol acetyltransferase gene were microinjected into mouse oocytes. Analyses of enzyme activity in male and female transgenic mice showed that 0.5 kb of the HSLtes promoter was sufficient to direct expression only in testis. Cell transfection experiments showed that CREMtau, a testis-specific transcriptional activator, does not transactivate the HSLtes promoter. Using gel retardation assays, four testis-specific binding regions (TSBR) were identified using testis and liver nuclear extracts. The testis-specific protein binding on TSBR4 was selectively competed by a probe containing a SRY/Sox protein DNA recognition site. Sox5 and Sox6 which are expressed in post-meiotic germ cells bound TSBR4. Mutation of the AACAAAG motif in TSBR4 abolished the binding. Moreover, binding of the high mobility group domain of Sox5 induced a bend within TSBR4. Together, our results showed that 0.5 kb of the human HSLtes promoter bind Sox proteins and contain cis-acting elements essential for the testis specificity of HSL.
Blaise
R
R
INSERM Unit 317, Institut Louis Bugnard, Université Paul Sabatier, Hôpital Rangueil, F-31403 Toulouse Cedex 4, France.
Grober
J
J
Rouet
P
P
Tavernier
G
G
Daegelen
D
D
Langin
D
D
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D005260:Female; D006801:Humans; D008297:Male; D008540:Meiosis; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011494:Protein Kinases; D016914:Ribonuclease H / Q000378:metabolism; D002787:Sterol Esterase / Q000096:biosynthesis*; D013737:Testis / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection
D009687:Nuclear Proteins; D004247:DNA; D011494:Protein Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D002787:Sterol Esterase; D016914:Ribonuclease H