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Sumoylation and ubiquitinylation reversibly regulate the activity of transcription factors through covalent attachment to lysine residues of target proteins. We examined whether the Ets-1 transcription factor is modified by sumoylation and/or ubiquitinylation. Among four potential SUMO motifs in Ets-1, we identified lysines 15 and 227 within the LK 15 YE and IK 227 QE motifs, as being the sumoylation acceptor sites. Using transfection of Ets-1 wildtype (WT) or its sumoylation deficient version (Ets-1 K15R/K227R), as well as WT or mutant proteins of the SUMO pathway, we further demonstrated that the E2 SUMO-conjugating enzyme Ubc9 and a E3 SUMO ligase, PIASy, can enhance Ets-1 sumoylation, while a SUMO protease, SENP1, can desumoylate Ets-1. We also found that Ets-1 is modified by K48-linked polyubiquitinylation independently of the sumoylation acceptor sites and is degraded through the 26S proteasome pathway, while sumoylation of Ets-1 does not affect its stability. Finally, sumoylation of Ets-1 leads to reduced transactivation and we demonstrated that previously identified critical lysine residues in Synergistic Control motifs are the sumoylation acceptor sites of Ets-1. These data show that Ets-1 can be modified by sumoylation and/or ubiquitinylation, with sumoylation repressing transcriptional activity of Ets-1 and having no clear antagonistic action on the ubiquitin-proteasome degradation pathway.

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Regulation of the Ets-1 transcription factor by sumoylation and ubiquitinylation

Degerny

Vintonenko

et al. 2006

“…Ets factors bind to the GGAA/T core motif (Ets-binding site, EBS) in the enhancer or promoter regions of their target genes (Sementchenko and Watson, 2000;Dittmer, 2003;Obika et al, 2003). ETS1 is involved in cell differentiation, proliferation, lymphoid cell development, transformation, apoptosis, angiogenesis, motility and invasion (Li et al, 1999;Kavurma et al, 2002;Watson et al, 2002;Xu et al, 2002).…”

Section: Introductionmentioning

confidence: 99%

SP100 expression modulates ETS1 transcriptional activity and inhibits cell invasion

Yordy

Sementchenko

et al. 2004

The ETS1 transcription factor is a member of the Ets family of conserved sequence-specific DNA-binding proteins. ETS1 has been shown to play important roles in various cellular processes such as proliferation, differentiation, lymphoid development, motility, invasion and angiogenesis. These diverse roles of ETS1 are likely to be dependent on specific protein interactions. To identify proteins that interact with ETS1, a yeast two-hybrid screen was conducted. Here, we describe the functional interaction between SP100 and ETS1. SP100 protein interacts with ETS1 both in vitro and in vivo. SP100 is localized to nuclear bodies and ETS1 expression alters the nuclear body morphology in living cells. SP100 negatively modulates ETS1 transcriptional activation of the MMP1 and uPA promoters in a dose-dependent manner, decreases the expression of these endogenous genes, and reduces ETS1 DNA binding. Expression of SP100 inhibits the invasion of breast cancer cells and is induced by Interferon-a, which has been shown to inhibit the invasion of cancer cells. These data demonstrate that SP100 modulates ETS1-dependent biological processes.

“…Ets proteins have been shown to be able to activate many genes (Sementchenko and Watson, 2000) and to be deregulated in various diseases, including cancer (Dittmer and Nordheim, 1998;Foos and Hanser, 2004;Hsu et al, 2004). Ets1 is overexpressed in a wide variety of invasive cancer cells and its expression in tumor cells or in tumor-associated vascular or avascular stroma often correlates with poor prognosis (Dittmer, 2003). How Ets1 is regulated in tumor cells is still not well understood.…”

Section: Introductionmentioning

confidence: 99%

Ets1 is an effector of protein kinase Cα in cancer cells

Vetter¹,

Blumenthal

Lindemann

et al. 2004

Self Cite

PKCa and Ets1 are both associated with breast cancer progression. Our previous studies suggested that these proteins are likely to functionally interact with one another. Here, we show that attenuation of endogenous PKCa expression (siPa) by RNA interference leads to reduced Ets1 protein expression in a variety of cancer cells. Pulse-chase experiments and treatment with proteasome inhibitor MG-132 revealed that siPa interferes with both Ets1 protein synthesis and stability. The effect of siPa on Ets1 expression could be partially prevented by KN-93, suggesting that calcium/calmodulin-dependent kinase II (CaMKII), a modulator of Ets1 activity, may play a role in PKCa-dependent Ets1 regulation. In contrast, Ets1-regulating kinases ERK1/ 2 were not found to be involved in this process. To assess the importance of the PKCa/Ets1 interaction, we compared the biological responses of MDA-MB-231 cells to PKCa-and Ets1-specific siRNAs (siE1). While only siPa induced changes in cellular morphology and anchorage-independent growth, both siRNAs similarly affected cellular responses to the antitumor drug mithramycin A and to UV light. Microarray analyses further showed that the expression of a certain set of genes was equally affected by siPa and siE1. The data suggest that Ets1 serves as an effector for PKCa to fulfil certain functions in cancer cells.