Understanding the role of E2F7 in the regulation of cellular proliferation and DNA damage responses.

188 p.E2F transcription factors control diverse biological processes through regulation of target gene expression. The identification of a large set of genes regulated by each individual E2F, including those coding for microRNAs, has led to a better understanding of the functions performed by the different members of the family. Many studies have detailed the role of classical E2Fs in cell cycle control and DNA damage response. By contrast, the contribution of the atypical members of the family, E2F7 and E2F8, to these processes has not been clearly defined. A recent study from our group identified a set of novel microRNAs and protein-coding genes regulated by E2F7. These genes are involved in processes such as cell cycle regulation or DNA damage response. In this work, we have examined the role that E2F7 plays in the regulation of these processes through the transcriptional regulation of its target genes. We have identified E2F7 as a transcription factor required for the repression of a set of microRNAs that promote cellular proliferation. We show that miR-25, miR-92 and miR-7 expression is controlled at the transcriptional level by the antagonistic activity of E2F7 and E2F1-3. Interestingly, we find that several E2F7-repressed microRNAs downregulate the expression of cell cycle progression inhibitors and promote cellular proliferation, suggesting that E2F7 restrains cell cycle progression through repression of proliferation-promoting microRNAs.Importantly, we show that E2F7 plays a key role in the maintenance of genomic stability. We present evidence of E2F7-dependent transcriptional and non-transcriptional mechanisms for modulating cellular responses to genotoxic exposure. We identify an E2F7-dependent transcriptional regulation program that restricts homologous recombination-mediated DNA repair and cellular recovery upon induction of DNA lesions that interfere with replication fork progression (DNA interstrand cross-links and PARP1 inhibition).Additionally, we present evidence of a non-transcriptional mechanism by which E2F7 modulates cellular responses to alkylating DNA damage, possibly involving interaction with the repair protein XRRC1. Loss of E2F7 confers an increased resistance to chemotherapy in homologous recombination-deficient cells, a potentially harmful outcome for cancer treatment. Altogether, results in this work reveal a key role for E2F7 in limiting cellular proliferation and promoting genomic stability by ensuring the timely expression of protein-coding and microRNA genes that are required for cell cycle progression and DNA damage repair

E2F7 regulates transcription and maturation of multiple microRNAs to restrain cell proliferation

This work was supported by the Spanish Ministry [SAF2012-33551, co-funded by the European RegionalDevelopment fund to A.M.Z., SAF2012-38215 to M.M.,SAF2014-57791-REDC to A.M.Z. and to M.M.]; BasqueGovernment [IT634-13 to A.M.Z.]; University of theBasque Country UPV/EHU [UFI1120 to A.M.Z.]; Excellence Network CellSYS [BFU2014-52125-REDT to M.M.];Comunidad de Madrid [S2010/BMD-2470 to M.M.];Basque Government Fellowship for graduate studies (to J.M.). Funding for open access charge: Basque Government [IT634-13]. Conflict of interest statement. None declared.E2F transcription factors (E2F1-8) are known to coordinately regulate the expression of a plethora of target genes, including those coding for microRNAs (miRNAs), to control cell cycle progression. Recent work has described the atypical E2F factor E2F7 as a transcriptional repressor of cell cycle-related protein-coding genes. However, the contribution of E2F7 to miRNA gene expression during the cell cycle has not been defined. We have performed a genome-wide RNA sequencing analysis to identify E2F7-regulated miRNAs and show that E2F7 plays as a major role in the negative regulation of a set of miRNAs that promote cellular proliferation. We provide mechanistic evidence for an interplay between E2F7 and the canonical E2F factors E2F1-3 in the regulation of multiple miRNAs. We show that miR-25, -26a, -27b, -92a and -7 expression is controlled at the transcriptional level by the antagonistic activity of E2F7 and E2F1-3. By contrast, let-7 miRNA expression is controlled indirectly through a novel E2F/c-MYC/LIN28B axis, whereby E2F7 and E2F1-3 modulate c-MYC and LIN28B levels to impact let-7 miRNA processing and maturation. Taken together, our data uncover a new regulatory network involving transcriptional and post-transcriptional mechanisms controlled by E2F7 to restrain cell cycle progression through repression of proliferation-promoting miRNAs.S

Sustained CHK2 activity, but not ATM activity, is critical to maintain a G1 arrest after DNA damage in untransformed cells

BACKGROUND: The G1 checkpoint is a critical regulator of genomic stability in untransformed cells, preventing cell cycle progression after DNA damage. DNA double-strand breaks (DSBs) recruit and activate ATM, a kinase which in turn activates the CHK2 kinase to establish G1 arrest. While the onset of G1 arrest is well understood, the specific role that ATM and CHK2 play in regulating G1 checkpoint maintenance remains poorly characterized. RESULTS: Here we examine the impact of ATM and CHK2 activities on G1 checkpoint maintenance in untransformed cells after DNA damage caused by DSBs. We show that ATM becomes dispensable for G1 checkpoint maintenance as early as 1h after DSB induction. In contrast, CHK2 kinase activity is necessary to maintain the G1 arrest, independently of ATM, ATR, and DNA-PKcs, implying that the G1 arrest is maintained in a lesion-independent manner. Sustained CHK2 activity is achieved through auto-activation and its acute inhibition enables cells to abrogate the G1-checkpoint and enter into S-phase. Accordingly, we show that CHK2 activity is lost in cells that recover from the G1 arrest, pointing to the involvement of a phosphatase with fast turnover. CONCLUSION: Our data indicate that G1 checkpoint maintenance relies on CHK2 and that its negative regulation is crucial for G1 checkpoint recovery after DSB induction.This research was funded by grants from MCIU/AEI/FEDER, UE (SAF2015-67562-R and RTI2018-097497-B-100) and Basque Government, Department of Education (IT1257-19) to A.M.Z., and Cancer Genomics Center Gravity Program (CGC.nl), Oncode Institute, Dutch Cancer Society (NKI 2014-6787) grants to R.H.M. I.G.-S. was supported with a postdoctoral fellowship from the Basque Country Government (Spain). J.V.-R. was supported by a postdoctoral fellowship from the University of the Basque Country (UPV/EHU)

Golgi Oncoprotein GOLPH3 Gene Expression is Regulated by Functional E2F and CREB/ATF Promoter Elements

The Golgi organelle duplicates its protein and lipid content to segregate evenly between two daughter cells after mitosis. However, how Golgi biogenesis is regulated during interphase remains largely unknown. Here we show that messenger RNA (mRNA) expression of GOLPH3 and GOLGA2, two genes encoding Golgi proteins, is induced specifically in G1 phase, suggesting a link between cell cycle regulation and Golgi growth. We have examined the role of E2F transcription factors, critical regulators of G1 to S progression of the cell cycle, in the expression of Golgi proteins during interphase. We show that promoter activity for GOLPH3, a Golgi protein that is also oncogenic, is induced by E2F1-3 and repressed by E2F7. Mutation of the E2F motifs present in the GOLPH3 promoter region abrogates E2F1-mediated induction of a GOLPH3 luciferase reporter construct. Furthermore, we identify a critical CREB/ATF element in the GOLPH3 promoter that is required for its steady state and ATF2-induced expression. Interestingly, depletion of GOLPH3 with small interfering RNA (siRNA) delays the G1 to S transition in synchronized U2OS cells. Taken together, our results reveal a link between cell cycle regulation and Golgi function, and suggest that E2F-mediated regulation of Golgi genes is required for the timely progression of the cell cycle.This work was supported by grants from the Spanish Ministry (SAF2015-67562-R, co-financed by Feder funds, and SAF2014-57791-REDC) and the Basque Government (IT634-13) to AMZ. B.P.-G. is recipient of a Spanish Ministry FPI fellowship for graduate studies; J.V.R. was recipient of a UPV/EHU fellowship for graduate studies; G.M. was recipient of a Spanish Ministry FPU fellowship for graduate studies

Golgi Oncoprotein GOLPH3 Gene Expression is Regulated by Functional E2F and CREB/ATF Promoter Elements

The Golgi organelle duplicates its protein and lipid content to segregate evenly between two daughter cells after mitosis. However, how Golgi biogenesis is regulated during interphase remains largely unknown. Here we show that messenger RNA (mRNA) expression of GOLPH3 and GOLGA2, two genes encoding Golgi proteins, is induced specifically in G1 phase, suggesting a link between cell cycle regulation and Golgi growth. We have examined the role of E2F transcription factors, critical regulators of G1 to S progression of the cell cycle, in the expression of Golgi proteins during interphase. We show that promoter activity for GOLPH3, a Golgi protein that is also oncogenic, is induced by E2F1-3 and repressed by E2F7. Mutation of the E2F motifs present in the GOLPH3 promoter region abrogates E2F1-mediated induction of a GOLPH3 luciferase reporter construct. Furthermore, we identify a critical CREB/ATF element in the GOLPH3 promoter that is required for its steady state and ATF2-induced expression. Interestingly, depletion of GOLPH3 with small interfering RNA (siRNA) delays the G1 to S transition in synchronized U2OS cells. Taken together, our results reveal a link between cell cycle regulation and Golgi function, and suggest that E2F-mediated regulation of Golgi genes is required for the timely progression of the cell cycle.This work was supported by grants from the Spanish Ministry (SAF2015-67562-R, co-financed by Feder funds, and SAF2014-57791-REDC) and the Basque Government (IT634-13) to AMZ. B.P.-G. is recipient of a Spanish Ministry FPI fellowship for graduate studies; J.V.R. was recipient of a UPV/EHU fellowship for graduate studies; G.M. was recipient of a Spanish Ministry FPU fellowship for graduate studies

An E2F7-Dependent Transcriptional Program Modulates DNA Damage Repair And Genomic Stability

Corrigendum published on 03 July 2019 Nucleic Acids Research 47 (14) : 7716–7717 (2019) https://doi.org/10.1093/nar/gkz587The cellular response to DNA damage is essential for maintaining the integrity of the genome. Recent evidence has identified E2F7 as a key player in DNA damage-dependent transcriptional regulation of cell-cycle genes. However, the contribution of E2F7 to cellular responses upon genotoxic damage is still poorly defined. Here we show that E2F7 represses the expression of genes involved in the maintenance of genomic stability, both throughout the cell cycle and upon induction of DNA lesions that interfere with replication fork progression. Knockdown of E2F7 leads to a reduction in 53BP1 and FANCD2 foci and to fewer chromosomal aberrations following treatment with agents that cause interstrand crosslink (ICL) lesions but not upon ionizing radiation. Accordingly, E2F7-depleted cells exhibit enhanced cell-cycle re-entry and clonogenic survival after exposure to ICL-inducing agents. We further report that expression and functional activity of E2F7 are p53-independent in this context. Using a cell-based assay, we show that E2F7 restricts homologous recombination through the transcriptional repression of RAD51. Finally, we present evidence that downregulation of E2F7 confers an increased resistance to chemotherapy in recombination-deficient cells. Taken together, our results reveal an E2F7-dependent transcriptional program that contributes to the regulation of DNA repair and genomic integrity.This work was supported by grants from the Spanish Ministry [SAF2012-33551 and SAF2015-67562-R, co-financed by FEDER funds, and SAF2014-57791-REDC], the Basque Government [IT634-13 and KK-2015/89], and the University of the Basque Country UPV/EHU [UFI11/20] to AMZ; and grants from the Spanish Ministry [SAF2015-69920-R], and Worldwide Cancer Research [15-0278] to MM. JM was recipient of a Basque Government fellowship for graduate studies and JVR is recipient of a UPV/EHU fellowship for graduate studies. M.A.F. was supported by a young investigator grant from MINECO [SAF2014-60442-JIN; co-financed by FEDER funds]. Funding for open access charge: Spanish Ministry [SAF2015-67562-R, co-financed by FEDER funds]; Basque Government [IT634-13]

Impact of COVID-19 on cardiovascular testing in the United States versus the rest of the world

Objectives: This study sought to quantify and compare the decline in volumes of cardiovascular procedures between the United States and non-US institutions during the early phase of the coronavirus disease-2019 (COVID-19) pandemic. Background: The COVID-19 pandemic has disrupted the care of many non-COVID-19 illnesses. Reductions in diagnostic cardiovascular testing around the world have led to concerns over the implications of reduced testing for cardiovascular disease (CVD) morbidity and mortality. Methods: Data were submitted to the INCAPS-COVID (International Atomic Energy Agency Non-Invasive Cardiology Protocols Study of COVID-19), a multinational registry comprising 909 institutions in 108 countries (including 155 facilities in 40 U.S. states), assessing the impact of the COVID-19 pandemic on volumes of diagnostic cardiovascular procedures. Data were obtained for April 2020 and compared with volumes of baseline procedures from March 2019. We compared laboratory characteristics, practices, and procedure volumes between U.S. and non-U.S. facilities and between U.S. geographic regions and identified factors associated with volume reduction in the United States. Results: Reductions in the volumes of procedures in the United States were similar to those in non-U.S. facilities (68% vs. 63%, respectively; p = 0.237), although U.S. facilities reported greater reductions in invasive coronary angiography (69% vs. 53%, respectively; p < 0.001). Significantly more U.S. facilities reported increased use of telehealth and patient screening measures than non-U.S. facilities, such as temperature checks, symptom screenings, and COVID-19 testing. Reductions in volumes of procedures differed between U.S. regions, with larger declines observed in the Northeast (76%) and Midwest (74%) than in the South (62%) and West (44%). Prevalence of COVID-19, staff redeployments, outpatient centers, and urban centers were associated with greater reductions in volume in U.S. facilities in a multivariable analysis. Conclusions: We observed marked reductions in U.S. cardiovascular testing in the early phase of the pandemic and significant variability between U.S. regions. The association between reductions of volumes and COVID-19 prevalence in the United States highlighted the need for proactive efforts to maintain access to cardiovascular testing in areas most affected by outbreaks of COVID-19 infection

Golgi Oncoprotein <i>GOLPH3</i> Gene Expression Is Regulated by Functional E2F and CREB/ATF Promoter Elements

The Golgi organelle duplicates its protein and lipid content to segregate evenly between two daughter cells after mitosis. However, how Golgi biogenesis is regulated during interphase remains largely unknown. Here we show that messenger RNA (mRNA) expression of GOLPH3 and GOLGA2, two genes encoding Golgi proteins, is induced specifically in G1 phase, suggesting a link between cell cycle regulation and Golgi growth. We have examined the role of E2F transcription factors, critical regulators of G1 to S progression of the cell cycle, in the expression of Golgi proteins during interphase. We show that promoter activity for GOLPH3, a Golgi protein that is also oncogenic, is induced by E2F1-3 and repressed by E2F7. Mutation of the E2F motifs present in the GOLPH3 promoter region abrogates E2F1-mediated induction of a GOLPH3 luciferase reporter construct. Furthermore, we identify a critical CREB/ATF element in the GOLPH3 promoter that is required for its steady state and ATF2-induced expression. Interestingly, depletion of GOLPH3 with small interfering RNA (siRNA) delays the G1 to S transition in synchronized U2OS cells. Taken together, our results reveal a link between cell cycle regulation and Golgi function, and suggest that E2F-mediated regulation of Golgi genes is required for the timely progression of the cell cycle