DAPK2 is a novel E2F1/KLF6 target gene involved in their proapoptotic function

Abstract

Death-associated protein kinase 2 (DAPK2) belongs to a family of proapoptotic Ca 2 þ /calmodulin-regulated serine/ threonine kinases. We recently identified DAPK2 as an enhancing factor during granulocytic differentiation. To identify transcriptional DAPK2 regulators, we cloned 2.7 kb of the 5 0 -flanking region of the DAPK2 gene. We found that E2F1 and Kru¨ppel-like factor 6 (KLF6) strongly activate the DAPK2 promoter. We mapped the E2F1 and KLF6 responsive elements to a GC-rich region 5 0 of exon 1 containing several binding sites for KLF6 and Sp1 but not for E2F. Moreover, we showed that transcriptional activation of DAPK2 by E2F1 and KLF6 is dependent on Sp1 using Sp1/KLF6-deficient insect cells, mithramycin A treatment to block Sp1-binding or Sp1 knockdown cells. Chromatin immunoprecipitation revealed recruitment of Sp1 and to lesser extent that of E2F1 and KLF6 to the DAPK2 promoter. Activation of E2F1 in osteosarcoma cells led to an increase of endogenous DAPK2 paralleled by cell death. Inhibition of DAPK2 expression resulted in significantly reduced cell death upon E2F1 activation. Similarly, KLF6 expression in H1299 cells increased DAPK2 levels accompanied by cell death that is markedly decreased upon DAPK2 knockdown. Moreover, E2F1 and KLF6 show cooperation in activating the DAPK2 promoter. In summary, our findings establish DAPK2 as a novel Sp1-dependent target gene for E2F1 and KLF6 in cell death response.

KLF6 Suppresses Metastasis of Clear Cell Renal Cell Carcinoma via Transcriptional Repression of E2F1

“…KLF6 upregulation abolished E2F1 expression, whereas KLF6 downregulation significantly increased E2F1 expression. The previous document demonstrated that E2F1 and KLF6 cooperated in activating the DAPK2 promoter (39). In our study, promoter luciferase reporter and ChIP assays illustrated that KLF6 directly binds to the human E2F1 gene promoter and inactivates it.…”

Death-associated protein kinase 2: Regulator of apoptosis, autophagy and inflammation

“…Hence, early reports -in accordance with the published DAPK1 phenotype -suggested an involvement of DAPK2 in apoptosis. Consistently, silencing of DAPK2 in cells exposed to intrinsic or extrinsic death triggers resulted in reduced apoptosis (Britschgi et al, 2008(Britschgi et al, , 2010.…”

“…Hence, low levels of DAPK2 in myeloid precursor cells could be increased by demethylating agents (Rizzi et al, 2007), and downregulation of DAPK2 expression in Hodgkin lymphoma was associated with DAPK2 promoter hypermethylation (Tur et al, 2009). The transcription factor SP1 was shown to be indispensable for basic DAPK2 promoter activity, with E2F1 and KLF6 transcription factors necessary for further DAPK2 regulation (Britschgi et al, 2008). Several other transcription factors have been implicated in the regulation of DAPK2 transcription.…”

Inhibition of GATE-16 attenuates ATRA-induced neutrophil differentiation of APL cells and interferes with autophagosome formation

“…Gene Expression Assays for GABARAPL1, GATE-16, MAP1LC3B and CSF3R used in a 96 well format on the ABI 7500 Sequence detection system were Hs00744468_s1, Hs00371854_m1, Hs00797944_s1 and Hs00167918_m1, respectively (Applied Biosystems, Rotkreuz, Switzerland). HMBS primer and probes have been described previously [28]. …”