RESUMEN
Hypoxia and the Net ternary complex factor (TCF) regulate similar processes (angiogenesis, wound healing, and cellular migration) and genes (PAI-1, c-fos, erg-1, NOS-2, HO-1, and vascular endothelial growth factor genes), suggesting that they are involved in related pathways. We show here that hypoxia regulates Net differently from the other TCFs and that Net plays a role in the hypoxic response in vivo in mice and in cells. Hypoxia induces Net depletion from target promoters, nuclear export, ubiquitylation, and proteasomal degradation. Key mediators of the hypoxic response, the prolyl-4-hydroxylases containing domain proteins (PHDs), regulate Net. PHD downregulation in normoxia leads to Net degradation, and PHD overexpression delays Net downregulation by hypoxia. Net inhibition by RNA interference or mutation leads to altered regulation by hypoxia of the Net targets PAI-1, c-fos, and egr-1. We propose that hypoxia stimulates transcription of target promoters through removal of the repressor function of Net. Interestingly, the hematocrit response to a chemical inducer of hypoxia-like responses (cobalt chloride) is strongly altered in Net mutant mice. Our results show that the Net TCF is part of the biological response to hypoxia, adding a new component to an important pathological and physiological process.
Asunto(s)
Regulación de la Expresión Génica , Hipoxia , Proteínas Proto-Oncogénicas c-ets/metabolismo , Factores Complejos Ternarios/metabolismo , Animales , Células Cultivadas , Regulación hacia Abajo , Humanos , Ratones , Mutación , Inhibidor 1 de Activador Plasminogénico/genética , Inhibidor 1 de Activador Plasminogénico/metabolismo , Regiones Promotoras Genéticas , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteínas Proto-Oncogénicas c-ets/genética , ARN Interferente Pequeño/metabolismo , Factores Complejos Ternarios/genética , Ubiquitina/metabolismoRESUMEN
The general transcription factor TFIID is a macromolecular complex comprising the TATA-binding protein (TBP) and a set of 13-14 TBP associated factors (TAFs). This review discusses biochemical, genetic and electron microscopic data acquired over the past years that provide a model for the composition, organisation and assembly of TFIID. We also revisit ideas on how TFIID is recruited to the promoters of active and possibly repressed genes. Recent observations show that recognition of acetylated and methylated histone residues by structural domains in several TAFs plays an important role. Finally, we highlight several genetic studies suggesting that TFIID is required for initiation of transcription, but not for maintaining transcription once a promoter is in an active state.
Asunto(s)
Factor de Transcripción TFIID/química , Factor de Transcripción TFIID/metabolismo , Animales , ADN/metabolismo , Histonas/metabolismo , Humanos , Modelos Moleculares , Complejos Multiproteicos/química , Complejos Multiproteicos/metabolismo , Regiones Promotoras Genéticas , Estructura Terciaria de Proteína , Factor de Transcripción TFIID/genética , Transcripción GenéticaRESUMEN
Collagen 6A3 (Col6a3), a component of extracellular matrix, is often up-regulated in tumours and is believed to play a pro-oncogenic role. However the mechanisms of its tumorigenic activity are poorly understood. We show here that Col6a3 is highly expressed in densely growing mouse embryonic fibroblasts (MEFs). In MEFs where the TAF4 subunit of general transcription factor IID (TFIID) has been inactivated, elevated Col6a3 expression prevents contact inhibition promoting their 3 dimensional growth as foci and fibrospheres. Analyses of gene expression in densely growing Taf4(-/-) MEFs revealed repression of the Hippo pathway and activation of Wnt signalling. The Hippo activator Kibra/Wwc1 is repressed under dense conditions in Taf4(-/-) MEFs, leading to nuclear accumulation of the proliferation factor YAP1 in the cells forming 3D foci. At the same time, Wnt9a is activated and the Sfrp2 antagonist of Wnt signalling is repressed. Surprisingly, treatment of Taf4(-/-) MEFs with all-trans retinoic acid (ATRA) restores contact inhibition suppressing 3D growth. ATRA represses Col6a3 expression independently of TAF4 expression and Col6a3 silencing is sufficient to restore contact inhibition in Taf4(-/-) MEFs and to suppress 3D growth by reactivating Kibra expression to induce Hippo signalling and by inducing Sfrp2 expression to antagonize Wnt signalling. All together, these results reveal a critical role for Col6a3 in regulating both Hippo and Wnt signalling to promote 3D growth, and show that the TFIID subunit TAF4 is essential to restrain the growth promoting properties of Col6a3. Our data provide new insight into the role of extra cellular matrix components in regulating cell growth.