PMID- 10499508
OWN - NLM
STAT- MEDLINE
DCOM- 19991012
LR  - 20131121
IS  - 0013-7227 (Print)
IS  - 0013-7227 (Linking)
VI  - 140
IP  - 10
DP  - 1999 Oct
TI  - Characterization of estrogen receptor-beta (ERbeta) messenger ribonucleic acid 
      and protein expression in rat granulosa cells.
PG  - 4530-41
AB  - We have examined estrogen-responsiveness of ovarian granulosa cells by focusing 
      on estrogen receptor (ER) expression. Estrogen responsiveness was determined by 
      examining the effect of 17beta-estradiol (1-10 nM) on luciferase reporter 
      activity in rat granulosa cells transfected with an ERE-luciferase construct. The 
      results demonstrate an estrogen-induced (approximately 3-fold) increase in 
      luciferase reporter activity, indicating that granulosa cells contain functional 
      estrogen response element (ERE)-binding transcriptional activators. Gel mobility 
      shift assays in combination with ER antibodies show that ERbeta is the 
      predominant ERE-binding protein in granulosa cells. Western blotting results show 
      that granulosa cells contain ERbeta-immunoreactive protein(s) migrating at a size 
      substantially larger than the recombinant protein generated from the originally 
      proposed 485 amino acid open-reading frame. This size discrepancy is not due to 
      granulosa cell expression of ERbeta isoforms with insertions within the coding 
      region because RT-PCR assays revealed products with sizes expected for ERbeta, 
      ERbetaB, and delta3 isoforms. This size discrepancy appears to be due to usage of 
      a well-conserved, upstream in-frame translation initiation codon (ATG436) leading 
      to a 530 amino acid open reading frame. ERbeta messenger RNA (mRNA) 
      characterization using 5'-rapid amplification of complementary DNA ends (5'-RACE) 
      show the presence of two different (P1- and P2-) 5'-ends of rat ERbeta mRNA 
      encoding the full-length ERbeta protein. The generation of the P2-specific exon 
      is likely due to initiation of transcription from an alternative promoter. Both 
      P1- and P2-specific exon-containing ERbeta mRNAs are expressed in granulosa 
      cells, and they are rapidly down-regulated by the cAMP-mediated intracellular 
      signaling pathway in cultured granulosa cells. Taken together, our results show 
      that rat granulosa cells produce two different 3',5'-cAMP-regulated ERbeta mRNA 
      species and that these mRNA species are capable of encoding the full-length 
      ERbeta protein.
FAU - O'Brien, M L
AU  - O'Brien ML
AD  - Graduate Center for Toxicology, University of Kentucky, Lexington 40536-0084, 
      USA.
FAU - Park, K
AU  - Park K
FAU - In, Y
AU  - In Y
FAU - Park-Sarge, O K
AU  - Park-Sarge OK
LA  - eng
GR  - ES08501/ES/NIEHS NIH HHS/United States
GR  - HD-30719/HD/NICHD NIH HHS/United States
GR  - HD-36879/HD/NICHD NIH HHS/United States
GR  - etc.
PT  - Journal Article
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - Endocrinology
JT  - Endocrinology
JID - 0375040
RN  - 0 (Estrogen Receptor beta)
RN  - 0 (RNA, Messenger)
RN  - 0 (Receptors, Estrogen)
RN  - 0 (Tissue Extracts)
RN  - 4TI98Z838E (Estradiol)
RN  - EC 1.13.12.- (Luciferases)
SB  - IM
MH  - Animals
MH  - Base Sequence/genetics
MH  - Cells, Cultured
MH  - Estradiol/pharmacology
MH  - Estrogen Receptor beta
MH  - Exons/physiology
MH  - Female
MH  - Granulosa Cells/drug effects/*metabolism
MH  - Luciferases/genetics/metabolism
MH  - Molecular Sequence Data
MH  - Ovary/chemistry
MH  - RNA, Messenger/*metabolism
MH  - Rats
MH  - Rats, Sprague-Dawley
MH  - Receptors, Estrogen/*genetics/*metabolism
MH  - Tissue Extracts/metabolism
MH  - Transcription, Genetic
MH  - Uterus/chemistry
EDAT- 1999/09/28 00:00
MHDA- 1999/09/28 00:01
CRDT- 1999/09/28 00:00
PHST- 1999/09/28 00:00 [pubmed]
PHST- 1999/09/28 00:01 [medline]
PHST- 1999/09/28 00:00 [entrez]
AID - 10.1210/endo.140.10.7032 [doi]
PST - ppublish
SO  - Endocrinology. 1999 Oct;140(10):4530-41. doi: 10.1210/endo.140.10.7032.