Ets-2 and C/EBP-beta are important mediators of ovine trophoblast Kunitz domain protein-1 gene expression in trophoblast.
BACKGROUND
The trophoblast Kunitz domain proteins (TKDPs) constitute a highly expressed, placenta-specific, multigene family restricted to ruminant ungulates and characterized by a C-terminal "Kunitz" domain, preceded by one or more unique N-terminal domains. TKDP-1 shares an almost identical expression pattern with interferon-tau, the "maternal recognition of pregnancy protein" in ruminants. Our goal here has been to determine whether the ovine (ov) Tkdp-1 and IFNT genes possess a similar transcriptional code.


RESULTS
The ovTkdp-1 promoter has been cloned and characterized. As with the IFNT promoter, the Tkdp-1 promoter is responsive to Ets-2, and promoter-driven reporter activity can be increased over 700-fold in response to over-expression of Ets-2 and a constitutively active form of protein Kinase A (PKA). Unexpectedly, the promoter element of Tkdp-1 responsible for this up-regulation, unlike that of the IFNT, does not bind Ets-2. However, mutation of a CCAAT/enhancer binding element within this control region not only reduced basal transcriptional activity, but prevented Ets-2 as well as cyclic adenosine 5'-monophosphate (cAMP)/PKA and Ras/mitogen-activated protein kinase (MAPK) responsiveness. In vitro binding experiments and in vivo protein-protein interaction assays implicated CCAAT/enhancer binding protein-beta (C/EBP-beta) as involved in up-regulating the Tkdp-1 promoter activity. A combination of Ets-2 and C/EBP-beta can up-regulate expression of the minimal Tkdp-1 promoter as much as 930-fold in presence of a cAMP analog. An AP-1-like element adjacent to the CCAAT enhancer, which binds Jun family members, is required for basal and cAMP/ C/EBP-beta-dependent activation of the gene, but not for Ets-2-dependent activity.


CONCLUSIONS
This paper demonstrates how Ets-2, a key transcription factor for trophoblast differentiation and function, can control expression of two genes (Tkdp-1 and IFNT) having similar spatial and temporal expression patterns via very different mechanisms.
Chakrabarty
Anindita
A
Departments of Veterinary Pathobiology, University of Missouri-Columbia, Columbia, MO 65211, USA. anindita.chakrabarty@vanderbilt.edu <anindita.chakrabarty@vanderbilt.edu>
Roberts
Michael R
MR
D000818:Animals; D020029:Base Pairing; D001665:Binding Sites; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000235:genetics / Q000378:metabolism*; D002822:Choriocarcinoma / Q000201:enzymology / Q000235:genetics / Q000473:pathology; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D007370:Interferon Type I / Q000235:genetics / Q000378:metabolism; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D041681:NIH 3T3 Cells; D011247:Pregnancy; D011257:Pregnancy Proteins / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D050787:Proto-Oncogene Protein c-ets-2 / Q000235:genetics / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012756:Sheep / Q000235:genetics*; D018808:Transcription Factor AP-1 / Q000235:genetics; D014158:Transcription, Genetic; D014327:Trophoblasts / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000235:genetics* / Q000378:metabolism
D022782:CCAAT-Enhancer-Binding Protein-beta; D007370:Interferon Type I; D011257:Pregnancy Proteins; D050787:Proto-Oncogene Protein c-ets-2; D012333:RNA, Messenger; D018808:Transcription Factor AP-1; D014361:Trypsin Inhibitors; C020947:interferon tau; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D016283:Proto-Oncogene Proteins p21(ras)