Regulation of Kruppel-like factor 4, 9, and 13 genes and the steroidogenic genes LDLR, StAR, and CYP11A in ovarian granulosa cells.
Kruppel-like factors (KLFs) are important Sp1-like eukaryotic transcriptional proteins. The LDLR, StAR, and CYP11A genes exhibit GC-rich Sp1-like sites, which have the potential to bind KLFs in multiprotein complexes. We now report that KLF4, KLF9, and KLF13 transcripts are expressed in and regulate ovarian cells. KLF4 and 13, but not KLF9, mRNA expression was induced and then repressed over time (P < 0.001). Combined LH and IGF-I stimulation increased KLF4 mRNA at 2 h (P < 0.01), whereas LH decreased KLF13 mRNA at 6 h (P < 0.05), and IGF-I reduced KLF13 at 24 h (P < 0.01) compared with untreated control. KLF9 was not regulated by either hormone. Transient transfection of KLF4, KLF9, and KLF13 suppressed LDLR/luc, StAR/luc, and CYP11A/luc by 80-90% (P < 0.001). Histone-deacetylase (HDAC) inhibitors stimulated LDLR/luc five- to sixfold and StAR/luc and CYP11A/luc activity twofold (P < 0.001) and partially reversed suppression by all three KLFs (P < 0.001). Deletion of the zinc finger domain of KLF13 abrogated repression of LDLR/luc. Lentiviral overexpression of the KLF13 gene suppressed LDLR mRNA (P < 0.001) and CYP11A mRNA (P = 0.003) but increased StAR mRNA (P = 0.007). Collectively, these data suggest that KLFs may recruit inhibitory complexes containing HDAC corepressors, thereby repressing LDLR and CYP11A transcription. Conversely, KLF13 may recruit unknown coactivators or stabilize StAR mRNA, thereby explaining enhancement of in situ StAR gene expression. These data introduce new potent gonadal transregulators of genes encoding proteins that mediate sterol uptake and steroid biosynthesis.
Endocrine Research Unit, Department of Internal Medicine, Mayo Clinic School of Medicine, Rochester, MN 55901, USA.
Sekar
S
Natesampillai
Jason
J
Kerkvliet
Peter C K
PC
Leung
Johannes D
JD
Veldhuis
D000818:Animals; D002478:Cells, Cultured; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000235:genetics*; D003001:Cloning, Molecular; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006107:Granulosa Cells / Q000378:metabolism*; D056572:Histone Deacetylase Inhibitors; D006801:Humans; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000096:biosynthesis / Q000235:genetics*; D016086:Lentivirus / Q000235:genetics; D010750:Phosphoproteins / Q000235:genetics*; D011973:Receptors, LDL / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012756:Sheep; D013256:Steroids / Q000096:biosynthesis*; D013268:Stimulation, Chemical; D013552:Swine; D014162:Transfection; D000098682:Steroidogenic Acute Regulatory Protein
D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; C000715467:KLF4 protein, human; C489676:KLF9 protein, human; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D010750:Phosphoproteins; D011973:Receptors, LDL; D013256:Steroids; D000098682:Steroidogenic Acute Regulatory Protein; D002786:Cholesterol Side-Chain Cleavage Enzyme