Induction of apoptosis by Elk-1 and ΔElk-1 proteins

Abstract

Elk-1, an ets related gene codes for at least two splice variants Elk-1, which regulates c-fos transcription and DElk-1, both of which function as transcriptional activators. To investigate the role of Elk-1 and DElk-1 proteins in apoptosis; we have developed rat ®broblast cell lines and human breast cancer cell lines expressing Elk-1 and DElk-1. The expression of Elk-1 and DElk-1 proteins in the Elk-1/DElk-1 transfectants were analysed by immuno¯uorescence, immunohistochemistry, and Western blot analysis. The Elk-1 unlike DElk-1 transfectants showed a shortened and¯attened morphology compared to the parental cells. We have found that calcium ionophore treatment of Rat-1 Elk-1, MCF-7 Elk-1, Rat-1 DElk-1 and MCF-7 DElk-1 transfectants resulted in programmed cell death. These results indicate that constitutive expression of Elk-1 and DElk-1 proteins triggers apoptosis in Rat-1 ®broblasts and breast cancer cells when treated with calcium ionophore.Keywords: Elk-1; DElk-1; apoptosis; calcium ionophore; breast cancer; MCF-7; Rat-1The Elk-1 gene belongs to the ets family of ternary complex factors (TCFs), i.e. Elk-1, SAP1, and NET/ ERP/SAP2/Elk-3 (Rao et al., 1989;Hipskind et al., 1991;Giovane et al., 1994;Lopez et al., 1994;Dalton and Treisman, 1992;Price et al., 1995;Nozaki et al., 1996). The Elk-1 gene codes for at least two alternately spliced products Elk-1 (Rao et al., 1989) and DElk-1 (Rao and Reddy, 1993) which function as transcriptional activators (Rao and Reddy, 1992;Bhattacharya et al., 1993), are substrates for MAP kinases Marias et al., 1993;Hill et al., 1993) and JNK protein kinases (Gupta et al., 1996;Whitmarsh et al., 1995). As mentioned earlier, the Elk-1 protein is a TCF which in association with serum response factor (SRF) forms a ternary complex on the serum response element (SRE) of the c-fos promoter and regulates cfos transcription (Hipskind et al., 1991). The TCF's which includes Elk-1 have three domains with similar sequences and functions. The ets domain mediates DNA binding, the SRF interaction domain interacts with SRF to form a ternary complex with the c-fos SRE and the C-terminal domain activates transcription upon phosphorylation by MAP kinases (Rao et al., 1989;Rao and Reddy, 1992;Dalton and Treisman, 1992;Janknecht et al., 1993Janknecht et al., , 1994Marias et al., 1993;Giovane et al., 1994;Hipskind et al., 1994;Kortenjann et al., 1994;Lopez et al., 1994;Hill et al., 1995;Price et al., 1995;Whitmarsh et al., 1995) and JNK kinases (Gupta et al., 1996). Thus Elk-1 represents a key link between signal transduction and induction of gene transcription.The Gag-Myb-Ets fusion protein, identi®ed in the avian acute leukemia virus E26 was shown to inhibit apoptosis and induce erythroid di erentiation in hematopoietic cells (Athanasiou et al., 1996). Similarly the Ets-1 proto-oncogene was shown to be required for the normal survival and activation of B and T cells while an Ets-1 splice variant was shown to induce apoptosis in human colon cancer cells indicating a role in apoptosis (Bories et al....

Gene replacement with the human BRCA1 locus: tissue specific expression and rescue of embryonic lethality in mice

“…The failure of these promoter-BRCA1 constructs to support development indicates that overexpressed, or mis-expressed, BRCA1 blocks development. This result was anticipated, in part, because of studies which demonstrate very low levels of BRCA1 expression in normal cells, and other studies that show cellular growth-arrest or apoptosis in many cell lines which overexpress BRCA1 (Shao et al, 1996;Wilson et al, 1997;Zhang et al, 1998). When we transfected mammary cell lines with BRCA1 cDNAs linked to BRCA1 promoter sequences, we did not observe apoptosis or cellular arrest (T Lane and E Solomon, preliminary observations).…”

“…The process is complicated by cellular senescence or apoptosis in many cells engineered to overexpress the BRCA1 cDNA from heterologous promoters (Aprelikova et al, 1999;Shao et al, 1996;Wilson et al, 1997;Zhang et al, 1998). Since mouse embryos de®cient in Brca1 die early in development, a straightforward assay of function is to analyse human BRCA1 gene replacement vectors for their ability to rescue Brca1-de®cient mouse embryos.…”

c-Fos oncogene regulator Elk-1 interacts with BRCA1 splice variants BRCA1a/1b and enhances BRCA1a/1b-mediated growth suppression in breast cancer cells

“…Elk-1, SAP-1 and NET/ERP/SAP2/Elk-3 (Dalton and Treisman, 1992;Giovane et al, 1994;Hipskind et al, 1991;Lopez et al, 1994;Nozaki et al, 1996;Price et al, 1995;Rao et al, 1989) that associate with the serum response factor (SRF) to regulate the transcription of c-Fos protooncogene (Dalton and Treisman, 1992;Hipskind et al, 1991). The Elk-1 gene codes for two splice variants Elk-1 (Rao et al, 1989) and DElk-1 which function as sequence speci®c transcriptional activators Rao and Reddy, 1992), are substrates for MAP kinases (Hill et al, 1993;Marais et al, 1993; and JNK protein kinases (Gupta et al, 1996;Whitmarsh et al, 1995), and induce apoptosis (Shao et al, 1998). Elk-1 is an e cient substrate for all three classes of MAPK, unlike SAP-1 which is activated only by ERK and p38 MAPK, and Net is activated by expression of oncogenic Raf (Wasylyk et al, 1998).…”

Role of Erk1/2 activation in prion disease pathogenesis: Absence of CCR1 leads to increased Erk1/2 activation and accelerated disease progression

“…A member of the ets-family of transcription factors, Elk-1, can be phosphorylated by p-Erk1/2 and has been shown to induce apoptosis in cancer cells (Davis, 1993;McCarthy et al, 1997;Shao et al, 1998), has been implicated in pathways leading to neuronal death (Vanhoutte et al, 1999;Lee et al, 2003;Choe et al, 2004), as well as been shown to be activated in neuronal cultures exposed to PrPres (Marella et al, 2005). To determine whether Elk-1 may be activated during scrapie infection, we evaluated brain homogenates for the presence of phosphorylated Elk-1 (p-Elk-1) protein.…”