DcE2F, a Functional Plant E2F-like Transcriptional Activator from Daucus carota

Abstract

In animal cells the progression of the cell cycle through G 1 /S transition and S phase is under the control of the pRB/E2F regulatory pathway. The E2F transcription factors are key activators of genes coding for several regulatory proteins and for enzymes involved in nucleotide and DNA synthesis. In this report we have detected the presence of E2F-like DNA binding activities in carrot nuclear extracts, and we have isolated a carrot cDNA (DcE2F) encoding a plant E2F homologue. The DcE2F gene is expressed in proliferating cells and is induced during the G 1 /S transition of the cell cycle. Supershift experiments using anti-DcE2F antiserum have confirmed that the DcE2F protein is a component of the carrot E2F-like nuclear activities. DNA binding assays have demonstrated that the DcE2F protein can recognize a canonical E2F cis-element in association with a mammalian DP protein. Furthermore, transactivation assays have revealed that DcE2F is a functional transcription factor that can transactivate, together with a DP partner, an E2F-responsive reporter gene in both plant and mammalian cells.Cell division in plants is mainly restricted to meristems and is regulated both temporally and spatially in response to plant growth regulators and environmental signals. Meristematic cell division occurs during the entire plant life, and because plant cells cannot migrate, control of cell proliferation is responsible for the formation of plant organs and structures (1). Remarkably, however, plant cells can readily undergo multiple cycles of endoreduplication, and additionally most of them are totipotent and can reprogram cell division even after completing their differentiation. These unique features suggest a unique flexibility in the control of the cell cycle in plants.In animal cells a pivotal role in the progression of cell cycle is played by the E2F family of transcription factors, which are key components of a critical checkpoint regulating entry of cells into S phase (2, 3). Progression through the G 1 and S phase of the cell cycle ultimately depends on the activation of genes coding for regulatory proteins and for the enzymes involved in nucleotide and DNA synthesis. The expression of many of these genes is largely under the control of the E2F family of transcription factors, which appear to be activated by multiple mitogenic signaling pathways. The E2F proteins bind to specific DNA sequences through a winged helix motif, forming prevalently heterodimers with distantly related partners of the DP (DRTF1 polypeptide) family (4). In mammalian cells six distinct E2F proteins associate with two different DP members and bind to similar DNA elements, which are conserved in the promoters of several genes that are activated in late G 1 and near the boundary G 1 /S. E2F gene targets include cell growth regulators such as cyclin A, Cdc2, c-Myc, and proliferating cell nuclear antigen and enzymes such as dihydrofolate reductase, thymidine kinase, ribonucleotide reductase, and DNA polymerase ␣ (5, 6). The importance of the E2F factors...

Microarray analysis of E2Fa-DPa-overexpressing plants uncovers a cross-talking genetic network between DNA replication and nitrogen assimilation

“…Promoter analysis of E2Fa-DPa-regulated genes The DNA-binding domains of the E2F and DP proteins are highly conserved between plants and mammals and, correspondingly, plant E2F-DP proteins have been shown by the technique of electrophoresis mobility shift assay to bind to the same canonical DNA-binding site as their mammalian counterparts (Albani et al, 2000;Ramirez-Parra and Gutierrez, 2000;de Jager et al, 2001). Furthermore, these E2F-binding sites regulate the expression of several plant genes involved in DNA synthesis (Kosugi and Ohashi, 2002a;Chabouté et al, 2000;Castellano et al, 2001;Egelkrout et al, 2001;Stevens et al, 2002).…”

“…tobacco, carrot, Arabidopsis and rice (Ramírez-Parra et al, 1999;Sekine et al, 1999;Albani et al, 2000;Magyar et al, 2000;Ramirez-Parra and Gutierrez, 2000;Kosugi and Ohashi, 2002a). In the Arabidopsis genome there are three E2F (E2Fa, E2Fb and E2Fc) and two DP (DPa and DPb) genes (Vandepoele et al, 2002).…”

E2Ls, E2F-like Repressors of Arabidopsis That Bind to E2F Sites in a Monomeric Form

“…The plant E2Fs share the same set of conserved domains (the DNA binding domain, dimerization domain, marked box and Rb-binding domain) as the animal E2F proteins, but they have no distinguishable similarity with the individual animal proteins, although they slightly resemble E2F-4, E2F-5, or E2F-6 (30,36). Like the E2F family from animals, the plant E2F proteins can bind to the consensus binding sites of the animal E2F, and their DNA binding activities can be stimulated by the plant DP proteins and human DP-1 (30,(33)(34)(35)(36). It has been also shown that they can bind human Rb or Rb-like proteins from plants through the C-terminal region (31,32,36).…”

“…RNR and PCNA promoters from tobacco and rice contain E2F sites that indeed bind E2F and control the transcription (28 -30). A number of cDNAs encoding E2F or DP homologs have been isolated and characterized in plants (31)(32)(33)(34)(35)(36)(37). The plant E2Fs share the same set of conserved domains (the DNA binding domain, dimerization domain, marked box and Rb-binding domain) as the animal E2F proteins, but they have no distinguishable similarity with the individual animal proteins, although they slightly resemble E2F-4, E2F-5, or E2F-6 (30,36).…”

Utilização de Análise Bioinformática e Validação por PCR Quantitativa em Tempo Real na Identificação da Indução Transcricional dos Fatores de Transcrição E2F1 e E2F4 em Linhagens de Glioblastoma.

“…Assim, essas proteínas desempenham papel crucial no controle do ciclo celular; além disso, apresentam ação sobre proteínas supressoras de tumor. As proteínas E2F possuem múltiplos domínios conservados evolutivamente, os quais são encontrados na maioria dos membros dessa família, em diversos organismos, de plantas (Ramírez-Parra et al, 1999;Albani et al, 2000;Guo et al, 2007) a eucariotos superiores, como mamíferos (Reichel et al, 1987;Sardet et al, 1995;Cartwright et al, 1998). A família E2F é composta, basicamente, por dois tipos de proteínas: E2F típicos (E2F1-6) e E2F atípicos (E2F7,8).…”