One of two Ets-binding sites in the cytokeratin EndoA enhancer is essential for enhancer activity and binds to Ets-2 related proteins.

Expression of the mouse cytokeratin EndoA gene is restricted in endodermal and epithelial cells, and is regulated by an enhancer that is located 1 kilobase (kb) 3' downstream from the gene. The enhancer consists of six direct repeats, of which each contains two predicted Ets binding sites (EBS1 and EBS2) containing GGAA as a core. Mutation analysis showed that EBS1 is essential for the enhancer activity and additional effects of EBS2, suggesting that some Ets-related proteins bind and activate the enhancer through EBS1. We also showed that Ets-2 mRNA is expressed in PYS-2 cells and that Ets-2 protein produced by E. coli interacts with EBS1 but not with EBS2. Using co-transfection assays, we showed that Ets-2 can trans-activate the enhancer in PYS-2 cells. Mutations that impair Ets-2 binding abolished the activity of the EndoA enhancer. The results obtained from the binding competition assay using an Ets-2 specific antibody, however, suggest that EBS1 binds to an Ets protein which is distinct from Ets-2. These data show that Ets-2 related protein binds and activates the EndoA enhancer in a sequence-specific fashion

Phytochrome Signaling Mechanisms

Phytochromes are red (R)/far-red (FR) light photoreceptors that play fundamental roles in photoperception of the light environment and the subsequent adaptation of plant growth and development. There are five distinct phytochromes in Arabidopsis thaliana, designated phytochrome A (phyA) to phyE. phyA is light-labile and is the primary photoreceptor responsible for mediating photomorphogenic responses in FR light, whereas phyB-phyE are light stable, and phyB is the predominant phytochrome regulating de-etiolation responses in R light. Phytochromes are synthesized in the cytosol in their inactive Pr form. Upon light irradiation, phytochromes are converted to the biologically active Pfr form, and translocate into the nucleus. phyB can enter the nucleus by itself in response to R light, whereas phyA nuclear import depends on two small plant-specific proteins FAR-RED ELONGATED HYPOCOTYL 1 (FHY1) and FHY1-LIKE (FHL). Phytochromes may function as light-regulated serine/threonine kinases, and can phosphorylate several substrates, including themselves in vitro. Phytochromes are phosphoproteins, and can be dephosphorylated by a few protein phosphatases. Photoactivated phytochromes rapidly change the expression of light-responsive genes by repressing the activity of CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1), an E3 ubiquitin ligase targeting several photomorphogenesis-promoting transcription factors for degradation, and by inducing rapid phosphorylation and degradation of Phytochrome-Interacting Factors (PIFs), a group of bHLH transcription factors repressing photomorphogenesis. Phytochromes are targeted by COP1 for degradation via the ubiquitin/26S proteasome pathway