FHL2, UBC9, and PIAS1 are novel estrogen receptor alpha-interacting proteins.
Estrogen plays important roles in the pathophysiology of atherosclerosis and cardiovascular diseases mediated by estrogen receptor alpha (ERalpha). To elucidate the molecular mechanisms, we screened ERalpha-interacting proteins from a human heart cDNA library using a yeast two-hybrid system, and identified the four and a half of LIM-only protein 2 (FHL2). FHL2 interacted with ERalpha in the presence of 17beta-estradiol, but not of tamoxifen or raloxifene in yeast. FHL2 mainly interacted with N-terminal A/B domain of ERalpha but not C-terminal ligand-binding domain. However, overexpression of full-length FHL2 did not affect ERalpha-dependent transcriptional activities of a reporter containing 3 copies of estrogen response element in COS-1 cells. Since tissue distribution of FHL2 was highly restricted to the heart, the function of FHL2 may be observed in a cell type- or promoter-specific manner. We have also detected strong interactions of ERalpha with Ubc9 and PIAS1 in yeast. Ubc9 and PIAS1, small ubiquitin-related modifier-1 (SUMO-1) conjugating enzyme and ligase, respectively, markedly interacted with ERalpha in a 17beta-estradiol-dependent manner. These proteins mainly interacted with the DNA-binding and ligand-binding domains of ERalpha. Overexpression of Ubc9 or PIAS1 potentiated ERalpha-mediated transcriptional activities in COS-1 cells in a dose-dependent manner, indicating that both Ubc9 and PIAS1 function as coactivators of ERalpha. In addition, the SUMOylation-defective mutant, Ubc9 (C93S) continued to enhance ERalpha-dependent transcriptional activities. These findings suggest that coactivator abilities and SUMOylation capacities of Ubc9 and PIAS1 are separable and distinct. The present studies indicate that ERalpha exhibit tissue-specific functions utilizing multiple tissue-restricted receptor-interacting proteins.
Kobayashi
Sakiko
S
Department of Internal Medicine, School of Medicine, Keio University, Tokyo, Japan.
Shibata
Hirotaka
H
Yokota
Kenichi
K
Suda
Noriko
N
Murai
Ayano
A
Kurihara
Isao
I
Saito
Ikuo
I
Saruta
Takao
T
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004958:Estradiol / Q000494:pharmacology; D047628:Estrogen Receptor alpha / Q000378:metabolism*; D015723:Gene Library; D018398:Homeodomain Proteins / Q000378:metabolism*; D006801:Humans; D060850:LIM-Homeodomain Proteins; D009124:Muscle Proteins / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D050828:Protein Inhibitors of Activated STAT; D025841:Small Ubiquitin-Related Modifier Proteins / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D020798:Two-Hybrid System Techniques; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism*; D015003:Yeasts; D000098843:Ubiquitin-Conjugating Enzyme UBC9
D047628:Estrogen Receptor alpha; C105987:FHL2 protein, human; D018398:Homeodomain Proteins; D060850:LIM-Homeodomain Proteins; D009124:Muscle Proteins; C106529:PIAS1 protein, human; D050828:Protein Inhibitors of Activated STAT; D025841:Small Ubiquitin-Related Modifier Proteins; D014157:Transcription Factors; D004958:Estradiol; D044763:Ubiquitin-Conjugating Enzymes; D000098843:Ubiquitin-Conjugating Enzyme UBC9