Regulation of transforming growth factor α and transferrin gene expression in human breast cancer cells /
17β-Estradiol (E2) induces transforming growth factor α (TGFα) gene expression in breast cancer cells and reporter gene activity in MCF-7 human breast cancer cells transfected with a construct containing a TGFα gene promoter insert (pTGFα-1145/-5). In MCF-7 cells transfected with pTGFα-1145/-377 co...
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| Format: | Thesis Book |
| Language: | English |
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[Place of publication not identified] :
[publisher not identified] ;
2001.
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| Subjects: | |
| Online Access: | http://proxy.library.tamu.edu/login?url=http://proquest.umi.com/pqdweb?did=728907911&sid=1&Fmt=2&clientId=2945&RQT=309&VName=PQD |
| Summary: | 17β-Estradiol (E2) induces transforming growth factor α (TGFα) gene expression in breast cancer cells and reporter gene activity in MCF-7 human breast cancer cells transfected with a construct containing a TGFα gene promoter insert (pTGFα-1145/-5). In MCF-7 cells transfected with pTGFα-1145/-377 containing -1145 to -377 from the TGFα gene promoter, E2 induced luciferase activity indicating additional E2-responsive elements upstream of two functional EREs. Deletion and mutation analysis identified a GC-rich site and an ERE half-site (Sp1(N)₃₀ERE1/2) which is E2-responsive and in electrophoretic mobility shift assays (EMSAs), incubation of [³²P]TGFα-625/-580 with nuclear extracts from MCF-7 cells gave a complex that could be decreased by coincubation with excess unlabeled ERE or Sp1 oligonucleotides. Furthermore, hERα and Sp1 antibodies supershifted this complex confirming that ERα and Sp1 bind this region of the TGFα gene promoter. The AhR ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) decreased TGFα mRNA levels induced by E2 and also decreased E2-induced luciferase activity in MCF-7 cells transfected with pTGFα-1145/-5 or pTGFα-1145/-377. Three iDREs were identified at -81, -464 and -485. TCDD did not inhibit luciferase activity of pTGFα-1145/-377-mDRE containing mutations of the iDREs at -464 and -485 or luciferase activity from B[a]P[] MCF-7 cells transfected with pTGFα-1145/-5. Cotreatment of MCF-7 cells with an AhR antagonist blocked inhibition by TCDD but not the antiestrogen ICI 182780. Footprinting with nuclear extracts from MCF-7 cells treated with E2 and TCDD resulted in protection of the iDREs indicating binding of nuclear proteins. E2 induces secretion of transferrin (Tf) and also induces reporter gene activity in MCF-7 cells transfected with pTf-3600/+39 containing 3.6 Kb of the Tf gene promoter. Deletion and mutation analysis has identified an E2-responsive element between -811 and -752 and EMSAs with [³²P]Tf-811/-752 and nuclear extracts from MCF-7 cells resulted in a complex that was not decreased by coincubation with unlabeled ERE, Sp1, NF1, or YY-1 oligonucleotides. However, EMSAs with [³²P]Tf-811/-752 and nuclear extracts from ER negative cells did not give a retarded complex and Tf-811/-752 was able to reduce binding of ER to [³²P]ERE. These results suggest that ER may bind Tf-811/-752 in a multiprotein complex to induce transcription of Tf. |
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| Item Description: | Vita. "Major Subject: Biochemistry". |
| Physical Description: | xii, 215 leaves : illustrations ; 28 cm. Issued also on microfiche from University Microfilm Inc. |
| Bibliography: | Includes bibliographical references (leaves 161-214). |