In vitro interactions between nuclear proteins and uncoupling protein gene promoter reveal several putative transactivating factors including Ets1, retinoid X receptor, thyroid hormone receptor, and a CACCC box-binding protein.
Previous studies of rat ucp (uncoupling protein) gene organization carried out in this laboratory identified regulatory sequences located in the 5'-flanking region. In this work, DNase I footprint analysis of the enhancer revealed two domains at base pairs (bp) -2444 to -2423 and bp -2352 to -2319. The former domain can bind in vitro, in a cooperative manner, factors related to nuclear factor 1 and Ets1; the latter domain contains a type 3 directly repeated sequence that was shown to be able to bind the retinoid X and triiodothyronine receptors. Moreover, a positive effect of retinoic acid on ucp mRNA levels in immortalized brown adipocytes was observed. DNase I footprint analysis identified two hypersensitive regions, A and B, at bp -509 to -472 and bp -403 to -350, respectively; region A contains a repeated CACCC box, and region B can bind protein related to Ets1. The A box differentially binds liver and brown adipose tissue nuclear proteins and could be involved in uncoupling protein induction. Further analysis showed three foot-printed boxes, C-E, at bp -182 to -159, -147 to -120, and -111 to -85, able to bind in vitro proteins related to nuclear factor 1, cAMP response element-binding protein, and Sp1, respectively.
Cassard-Doulcier
A M
AM
Centre de Recherches sur l'Endocrinologie Moléculaire et le Développement, CNRS, Meudon, France.
Larose
M
M
Matamala
J C
JC
Champigny
O
O
Bouillaud
F
F
Ricquier
D
D
D002001:Adipose Tissue, Brown / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D002478:Cells, Cultured; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism; D007473:Ion Channels; D008099:Liver / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008928:Mitochondria / Q000378:metabolism; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D015513:Oncogene Proteins; D011401:Promoter Regions, Genetic; D050783:Proto-Oncogene Proteins c-ets; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D018168:Receptors, Retinoic Acid; D011988:Receptors, Thyroid Hormone / Q000378:metabolism*; D047488:Retinoid X Receptors; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation; D000071256:Uncoupling Protein 1
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C086924:Elk3 protein, mouse; D007473:Ion Channels; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D015513:Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D018160:Receptors, Cytoplasmic and Nuclear; D018168:Receptors, Retinoic Acid; D011988:Receptors, Thyroid Hormone; D047488:Retinoid X Receptors; D014157:Transcription Factors; D000071256:Uncoupling Protein 1; D003850:Deoxyribonuclease I