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Papers by Dariel Ashton-Beaucage

Research paper thumbnail of Identification et étude de mécanismes régulant l’expression de MAPK

Research paper thumbnail of MiSTIC, an integrated platform for the analysis of heterogeneity in large tumour transcriptome datasets

Nucleic acids research, Jan 4, 2017

Genome-wide transcriptome profiling has enabled non-supervised classification of tumours, reveali... more Genome-wide transcriptome profiling has enabled non-supervised classification of tumours, revealing different sub-groups characterized by specific gene expression features. However, the biological significance of these subtypes remains for the most part unclear. We describe herein an interactive platform, Minimum Spanning Trees Inferred Clustering (MiSTIC), that integrates the direct visualization and comparison of the gene correlation structure between datasets, the analysis of the molecular causes underlying co-variations in gene expression in cancer samples, and the clinical annotation of tumour sets defined by the combined expression of selected biomarkers. We have used MiSTIC to highlight the roles of specific transcription factors in breast cancer subtype specification, to compare the aspects of tumour heterogeneity targeted by different prognostic signatures, and to highlight biomarker interactions in AML. A version of MiSTIC preloaded with datasets described herein can be ac...

Research paper thumbnail of How Genetics Has Helped Piece Together the MAPK Signaling Pathway

Methods in Molecular Biology, 2016

Cells respond to changes in their environment, to developmental cues, and to pathogen aggression ... more Cells respond to changes in their environment, to developmental cues, and to pathogen aggression through the action of a complex network of proteins. These networks can be decomposed into a multitude of signaling pathways that relay signals from the microenvironment to the cellular components involved in eliciting a specific response. Perturbations in these signaling processes are at the root of multiple pathologies, the most notable of these being cancer. The study of receptor tyrosine kinase (RTK) signaling led to the first description of a mechanism whereby an extracellular signal is transmitted to the nucleus to induce a transcriptional response. Genetic studies conducted in drosophila and nematodes have provided key elements to this puzzle. Here, we briefly discuss the somewhat lesser known contribution of these multicellular organisms to our understanding of what has come to be known as the prototype of signaling pathways. We also discuss the ostensibly much larger network of regulators that has emerged from recent functional genomic investigations of RTK/RAS/ERK signaling.

Research paper thumbnail of Faculty of 1000 evaluation for Exon definition complexes contain the tri-snRNP and can be directly converted into B-like precatalytic splicing complexes

F1000 - Post-publication peer review of the biomedical literature, 2012

Research paper thumbnail of Faculty of 1000 evaluation for Application of an integrated physical and functional screening approach to identify inhibitors of the Wnt pathway

F1000 - Post-publication peer review of the biomedical literature, 2010

Research paper thumbnail of Faculty of 1000 evaluation for CWC22 Connects Pre-mRNA Splicing and Exon Junction Complex Assembly

F1000 - Post-publication peer review of the biomedical literature, 2012

Research paper thumbnail of Faculty of 1000 evaluation for Mapping of signaling networks through synthetic genetic interaction analysis by RNAi

F1000 - Post-publication peer review of the biomedical literature, 2011

Research paper thumbnail of Faculty of 1000 evaluation for Integrating protein-protein interaction networks with phenotypes reveals signs of interactions

F1000 - Post-publication peer review of the biomedical literature, 2013

Research paper thumbnail of The Deubiquitinase USP47 Stabilizes MAPK by Counteracting the Function of the N-end Rule ligase POE/UBR4 in Drosophila

RAS-induced MAPK signaling is a central driver of the cell proliferation apparatus. Disruption of... more RAS-induced MAPK signaling is a central driver of the cell proliferation apparatus. Disruption of this pathway is widely observed in cancer and other pathologies. Consequently, considerable effort has been devoted to understanding the mechanistic aspects of RAS-MAPK signal transmission and regulation. While much information has been garnered on the steps leading up to the activation and inactivation of core pathway components, comparatively little is known on the mechanisms controlling their expression and turnover. We recently identified several factors that dictate Drosophila MAPK levels. Here, we describe the function of one of these, the deubiquitinase (DUB) USP47. We found that USP47 acts post-translation-ally to counteract a proteasome-mediated event that reduces MAPK half-life and thereby dampens signaling output. Using an RNAi-based genetic interaction screening strategy, we identified UBC6, POE/UBR4, and UFD4, respectively, as E2 and E3 enzymes that oppose USP47 activity. Further characterization of POE-associated factors uncovered KCMF1 as another key component modulating MAPK levels. Together, these results identify a novel protein degradation module that governs MAPK levels. Given the role of UBR4 as an N-recognin ubiquitin ligase, our findings suggest that RAS-MAPK signaling in Drosophila is controlled by the N-end rule pathway and that USP47 counteracts its activity. The RAS-MAPK pathway plays a central role in the development of multicellular organisms , predominantly by regulating cell proliferation and differentiation. At its core, the pathway includes the RAS GTPase and three kinases (RAF, MEK, and MAPK) that transmit RAS signals through a phosphorylation cascade. Several factors have been discovered over the years that modulate signal transmission by altering the kinetics of phosphoryla-tion/dephosphorylation of the core pathway components. In contrast, scant information is available on the mechanisms governing their expression and steady-state levels. Here, we

Research paper thumbnail of Faculty of 1000 evaluation for Intronic motif pairs cooperate across exons to promote pre-mRNA splicing

F1000 - Post-publication peer review of the biomedical literature, 2011

Research paper thumbnail of Faculty of 1000 evaluation for A U1-U2 snRNP interaction network during intron definition

F1000 - Post-publication peer review of the biomedical literature, 2000

Research paper thumbnail of The exon junction complex controls the splicing of MAPK and other long intron-containing transcripts in Drosophila

Cell, Jan 15, 2010

Signaling pathways are controlled by a vast array of posttranslational mechanisms. By contrast, l... more Signaling pathways are controlled by a vast array of posttranslational mechanisms. By contrast, little is known regarding the mechanisms that regulate the expression of their core components. We conducted an RNAi screen in Drosophila for factors modulating RAS/MAPK signaling and identified the Exon Junction Complex (EJC) as a key element of this pathway. The EJC binds the exon-exon junctions of mRNAs and thus far, has been linked exclusively to postsplicing events. Here, we report that the EJC is required for proper splicing of mapk transcripts by a mechanism that apparently controls exon definition. Moreover, whole transcriptome and RT-PCR analyses of EJC-depleted cells revealed that the splicing of long intron-containing genes, which includes mapk, is sensitive to EJC activity. These results identify a role for the EJC in the splicing of a subset of transcripts and suggest that RAS/MAPK signaling depends on the regulation of MAPK levels by the EJC.

Research paper thumbnail of A Functional Screen Reveals an Extensive Layer of Transcriptional and Splicing Control Underlying RAS/MAPK Signaling in Drosophila

PLoS biology, 2014

The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-... more The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-MAPK module that lies downstream of RAS is one of the main conduits through which RAS transmits proliferative signals in normal and cancer cells. Genetic and biochemical studies conducted over the last two decades uncovered a small set of factors regulating RAS/MAPK signaling. Interestingly, most of these were found to control RAF activation, thus suggesting a central regulatory role for this event. Whether additional factors are required at this level or further downstream remains an open question. To obtain a comprehensive view of the elements functionally linked to the RAS/MAPK cascade, we used a quantitative assay in Drosophila S2 cells to conduct a genome-wide RNAi screen for factors impacting RAS-mediated MAPK activation. The screen led to the identification of 101 validated hits, including most of the previously known factors associated to this pathway. Epistasis experiments were then carried out on individual candidates to determine their position relative to core pathway components. While this revealed several new factors acting at different steps along the pathway--including a new protein complex modulating RAF activation--we found that most hits unexpectedly work downstream of MEK and specifically influence MAPK expression. These hits mainly consist of constitutive splicing factors and thereby suggest that splicing plays a specific role in establishing MAPK levels. We further characterized two representative members of this group and surprisingly found that they act by regulating mapk alternative splicing. This study provides an unprecedented assessment of the factors modulating RAS/MAPK signaling in Drosophila. In addition, it suggests that pathway output does not solely rely on classical signaling events, such as those controlling RAF activation, but also on the regulation of MAPK levels. Finally, it indicates that core splicing components can also specifically impact alternative splicing.

Research paper thumbnail of The PP2C Alphabet Is a Negative Regulator of Stress-Activated Protein Kinase Signaling in Drosophila

The Jun N-terminal kinase and p38 pathways, also known as stress-activated protein kinase (SAPK) ... more The Jun N-terminal kinase and p38 pathways, also known as stress-activated protein kinase (SAPK) pathways, are signaling conduits reiteratively used throughout the development and adult life of metazoans where they play central roles in the control of apoptosis, immune function, and environmental stress responses. We recently identified a Drosophila Ser/Thr phosphatase of the PP2C family, named Alphabet (Alph), which acts as a negative regulator of the Ras/ERK pathway. Here we show that Alph also plays an inhibitory role with respect to Drosophila SAPK signaling during development as well as under stress conditions such as oxidative or genotoxic stresses. Epistasis experiments suggest that Alph acts at a step upstream of the MAPKKs Hep and Lic. Consistent with this interpretation, biochemical experiments identify the upstream MAPKKKs Slpr, Tak1, and Wnd as putative substrates. Together with previous findings, this work identifies Alph as a general attenuator of MAPK signaling in Drosophila.

Research paper thumbnail of The Exon Junction Complex Controls the Splicing of mapk and Other Long Intron-Containing Transcripts in Drosophila

Cell, 2010

Signaling pathways are controlled by a vast array of posttranslational mechanisms. By contrast, l... more Signaling pathways are controlled by a vast array of posttranslational mechanisms. By contrast, little is known regarding the mechanisms that regulate the expression of their core components. We conducted an RNAi screen in Drosophila for factors modulating RAS/MAPK signaling and identified the Exon Junction Complex (EJC) as a key element of this pathway. The EJC binds the exon-exon junctions of mRNAs and thus far, has been linked exclusively to postsplicing events. Here, we report that the EJC is required for proper splicing of mapk transcripts by a mechanism that apparently controls exon definition. Moreover, whole transcriptome and RT-PCR analyses of EJC-depleted cells revealed that the splicing of long intron-containing genes, which includes mapk, is sensitive to EJC activity. These results identify a role for the EJC in the splicing of a subset of transcripts and suggest that RAS/MAPK signaling depends on the regulation of MAPK levels by the EJC.

Research paper thumbnail of The exon junction complex controls the splicing of MAPK and other long intron-containing transcripts in Drosophila.

Research paper thumbnail of The PP2C Alphabet Is a Negative Regulator of Stress-Activated Protein Kinase Signaling in Drosophila

Research paper thumbnail of The exon junction complex: A splicing factor for long intron containing transcripts?

Research paper thumbnail of A comprehensive map of the mTOR signaling network

Molecular Systems …, Jan 1, 2010

The mammalian target of rapamycin (mTOR) is a central regulator of cell growth and proliferation.... more The mammalian target of rapamycin (mTOR) is a central regulator of cell growth and proliferation. mTOR signaling is frequently dysregulated in oncogenic cells, and thus an attractive target for anticancer therapy. Using CellDesigner, a modeling support software for graphical notation, we present herein a comprehensive map of the mTOR signaling network, which includes 964 species connected by 777 reactions. The map complies with both the systems biology markup language (SBML) and graphical notation (SBGN) for computational analysis and graphical representation, respectively. As captured in the mTOR map, we review and discuss our current understanding of the mTOR signaling network and highlight the impact of mTOR feedback and crosstalk regulations on drug-based cancer therapy. This map is available on the Payao platform, a Web 2.0 based community-wide interactive process for creating more accurate and information-rich databases. Thus, this comprehensive map of the mTOR network will serve as a tool to facilitate systems-level study of up-to-date mTOR network components and signaling events toward the discovery of novel regulatory processes and therapeutic strategies for cancer.

Research paper thumbnail of La signalisation RTK/RAS/ERK élargie

médecine/sciences, Jan 1, 2010

Research paper thumbnail of Identification et étude de mécanismes régulant l’expression de MAPK

Research paper thumbnail of MiSTIC, an integrated platform for the analysis of heterogeneity in large tumour transcriptome datasets

Nucleic acids research, Jan 4, 2017

Genome-wide transcriptome profiling has enabled non-supervised classification of tumours, reveali... more Genome-wide transcriptome profiling has enabled non-supervised classification of tumours, revealing different sub-groups characterized by specific gene expression features. However, the biological significance of these subtypes remains for the most part unclear. We describe herein an interactive platform, Minimum Spanning Trees Inferred Clustering (MiSTIC), that integrates the direct visualization and comparison of the gene correlation structure between datasets, the analysis of the molecular causes underlying co-variations in gene expression in cancer samples, and the clinical annotation of tumour sets defined by the combined expression of selected biomarkers. We have used MiSTIC to highlight the roles of specific transcription factors in breast cancer subtype specification, to compare the aspects of tumour heterogeneity targeted by different prognostic signatures, and to highlight biomarker interactions in AML. A version of MiSTIC preloaded with datasets described herein can be ac...

Research paper thumbnail of How Genetics Has Helped Piece Together the MAPK Signaling Pathway

Methods in Molecular Biology, 2016

Cells respond to changes in their environment, to developmental cues, and to pathogen aggression ... more Cells respond to changes in their environment, to developmental cues, and to pathogen aggression through the action of a complex network of proteins. These networks can be decomposed into a multitude of signaling pathways that relay signals from the microenvironment to the cellular components involved in eliciting a specific response. Perturbations in these signaling processes are at the root of multiple pathologies, the most notable of these being cancer. The study of receptor tyrosine kinase (RTK) signaling led to the first description of a mechanism whereby an extracellular signal is transmitted to the nucleus to induce a transcriptional response. Genetic studies conducted in drosophila and nematodes have provided key elements to this puzzle. Here, we briefly discuss the somewhat lesser known contribution of these multicellular organisms to our understanding of what has come to be known as the prototype of signaling pathways. We also discuss the ostensibly much larger network of regulators that has emerged from recent functional genomic investigations of RTK/RAS/ERK signaling.

Research paper thumbnail of Faculty of 1000 evaluation for Exon definition complexes contain the tri-snRNP and can be directly converted into B-like precatalytic splicing complexes

F1000 - Post-publication peer review of the biomedical literature, 2012

Research paper thumbnail of Faculty of 1000 evaluation for Application of an integrated physical and functional screening approach to identify inhibitors of the Wnt pathway

F1000 - Post-publication peer review of the biomedical literature, 2010

Research paper thumbnail of Faculty of 1000 evaluation for CWC22 Connects Pre-mRNA Splicing and Exon Junction Complex Assembly

F1000 - Post-publication peer review of the biomedical literature, 2012

Research paper thumbnail of Faculty of 1000 evaluation for Mapping of signaling networks through synthetic genetic interaction analysis by RNAi

F1000 - Post-publication peer review of the biomedical literature, 2011

Research paper thumbnail of Faculty of 1000 evaluation for Integrating protein-protein interaction networks with phenotypes reveals signs of interactions

F1000 - Post-publication peer review of the biomedical literature, 2013

Research paper thumbnail of The Deubiquitinase USP47 Stabilizes MAPK by Counteracting the Function of the N-end Rule ligase POE/UBR4 in Drosophila

RAS-induced MAPK signaling is a central driver of the cell proliferation apparatus. Disruption of... more RAS-induced MAPK signaling is a central driver of the cell proliferation apparatus. Disruption of this pathway is widely observed in cancer and other pathologies. Consequently, considerable effort has been devoted to understanding the mechanistic aspects of RAS-MAPK signal transmission and regulation. While much information has been garnered on the steps leading up to the activation and inactivation of core pathway components, comparatively little is known on the mechanisms controlling their expression and turnover. We recently identified several factors that dictate Drosophila MAPK levels. Here, we describe the function of one of these, the deubiquitinase (DUB) USP47. We found that USP47 acts post-translation-ally to counteract a proteasome-mediated event that reduces MAPK half-life and thereby dampens signaling output. Using an RNAi-based genetic interaction screening strategy, we identified UBC6, POE/UBR4, and UFD4, respectively, as E2 and E3 enzymes that oppose USP47 activity. Further characterization of POE-associated factors uncovered KCMF1 as another key component modulating MAPK levels. Together, these results identify a novel protein degradation module that governs MAPK levels. Given the role of UBR4 as an N-recognin ubiquitin ligase, our findings suggest that RAS-MAPK signaling in Drosophila is controlled by the N-end rule pathway and that USP47 counteracts its activity. The RAS-MAPK pathway plays a central role in the development of multicellular organisms , predominantly by regulating cell proliferation and differentiation. At its core, the pathway includes the RAS GTPase and three kinases (RAF, MEK, and MAPK) that transmit RAS signals through a phosphorylation cascade. Several factors have been discovered over the years that modulate signal transmission by altering the kinetics of phosphoryla-tion/dephosphorylation of the core pathway components. In contrast, scant information is available on the mechanisms governing their expression and steady-state levels. Here, we

Research paper thumbnail of Faculty of 1000 evaluation for Intronic motif pairs cooperate across exons to promote pre-mRNA splicing

F1000 - Post-publication peer review of the biomedical literature, 2011

Research paper thumbnail of Faculty of 1000 evaluation for A U1-U2 snRNP interaction network during intron definition

F1000 - Post-publication peer review of the biomedical literature, 2000

Research paper thumbnail of The exon junction complex controls the splicing of MAPK and other long intron-containing transcripts in Drosophila

Cell, Jan 15, 2010

Signaling pathways are controlled by a vast array of posttranslational mechanisms. By contrast, l... more Signaling pathways are controlled by a vast array of posttranslational mechanisms. By contrast, little is known regarding the mechanisms that regulate the expression of their core components. We conducted an RNAi screen in Drosophila for factors modulating RAS/MAPK signaling and identified the Exon Junction Complex (EJC) as a key element of this pathway. The EJC binds the exon-exon junctions of mRNAs and thus far, has been linked exclusively to postsplicing events. Here, we report that the EJC is required for proper splicing of mapk transcripts by a mechanism that apparently controls exon definition. Moreover, whole transcriptome and RT-PCR analyses of EJC-depleted cells revealed that the splicing of long intron-containing genes, which includes mapk, is sensitive to EJC activity. These results identify a role for the EJC in the splicing of a subset of transcripts and suggest that RAS/MAPK signaling depends on the regulation of MAPK levels by the EJC.

Research paper thumbnail of A Functional Screen Reveals an Extensive Layer of Transcriptional and Splicing Control Underlying RAS/MAPK Signaling in Drosophila

PLoS biology, 2014

The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-... more The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-MAPK module that lies downstream of RAS is one of the main conduits through which RAS transmits proliferative signals in normal and cancer cells. Genetic and biochemical studies conducted over the last two decades uncovered a small set of factors regulating RAS/MAPK signaling. Interestingly, most of these were found to control RAF activation, thus suggesting a central regulatory role for this event. Whether additional factors are required at this level or further downstream remains an open question. To obtain a comprehensive view of the elements functionally linked to the RAS/MAPK cascade, we used a quantitative assay in Drosophila S2 cells to conduct a genome-wide RNAi screen for factors impacting RAS-mediated MAPK activation. The screen led to the identification of 101 validated hits, including most of the previously known factors associated to this pathway. Epistasis experiments were then carried out on individual candidates to determine their position relative to core pathway components. While this revealed several new factors acting at different steps along the pathway--including a new protein complex modulating RAF activation--we found that most hits unexpectedly work downstream of MEK and specifically influence MAPK expression. These hits mainly consist of constitutive splicing factors and thereby suggest that splicing plays a specific role in establishing MAPK levels. We further characterized two representative members of this group and surprisingly found that they act by regulating mapk alternative splicing. This study provides an unprecedented assessment of the factors modulating RAS/MAPK signaling in Drosophila. In addition, it suggests that pathway output does not solely rely on classical signaling events, such as those controlling RAF activation, but also on the regulation of MAPK levels. Finally, it indicates that core splicing components can also specifically impact alternative splicing.

Research paper thumbnail of The PP2C Alphabet Is a Negative Regulator of Stress-Activated Protein Kinase Signaling in Drosophila

The Jun N-terminal kinase and p38 pathways, also known as stress-activated protein kinase (SAPK) ... more The Jun N-terminal kinase and p38 pathways, also known as stress-activated protein kinase (SAPK) pathways, are signaling conduits reiteratively used throughout the development and adult life of metazoans where they play central roles in the control of apoptosis, immune function, and environmental stress responses. We recently identified a Drosophila Ser/Thr phosphatase of the PP2C family, named Alphabet (Alph), which acts as a negative regulator of the Ras/ERK pathway. Here we show that Alph also plays an inhibitory role with respect to Drosophila SAPK signaling during development as well as under stress conditions such as oxidative or genotoxic stresses. Epistasis experiments suggest that Alph acts at a step upstream of the MAPKKs Hep and Lic. Consistent with this interpretation, biochemical experiments identify the upstream MAPKKKs Slpr, Tak1, and Wnd as putative substrates. Together with previous findings, this work identifies Alph as a general attenuator of MAPK signaling in Drosophila.

Research paper thumbnail of The Exon Junction Complex Controls the Splicing of mapk and Other Long Intron-Containing Transcripts in Drosophila

Cell, 2010

Signaling pathways are controlled by a vast array of posttranslational mechanisms. By contrast, l... more Signaling pathways are controlled by a vast array of posttranslational mechanisms. By contrast, little is known regarding the mechanisms that regulate the expression of their core components. We conducted an RNAi screen in Drosophila for factors modulating RAS/MAPK signaling and identified the Exon Junction Complex (EJC) as a key element of this pathway. The EJC binds the exon-exon junctions of mRNAs and thus far, has been linked exclusively to postsplicing events. Here, we report that the EJC is required for proper splicing of mapk transcripts by a mechanism that apparently controls exon definition. Moreover, whole transcriptome and RT-PCR analyses of EJC-depleted cells revealed that the splicing of long intron-containing genes, which includes mapk, is sensitive to EJC activity. These results identify a role for the EJC in the splicing of a subset of transcripts and suggest that RAS/MAPK signaling depends on the regulation of MAPK levels by the EJC.

Research paper thumbnail of The exon junction complex controls the splicing of MAPK and other long intron-containing transcripts in Drosophila.

Research paper thumbnail of The PP2C Alphabet Is a Negative Regulator of Stress-Activated Protein Kinase Signaling in Drosophila

Research paper thumbnail of The exon junction complex: A splicing factor for long intron containing transcripts?

Research paper thumbnail of A comprehensive map of the mTOR signaling network

Molecular Systems …, Jan 1, 2010

The mammalian target of rapamycin (mTOR) is a central regulator of cell growth and proliferation.... more The mammalian target of rapamycin (mTOR) is a central regulator of cell growth and proliferation. mTOR signaling is frequently dysregulated in oncogenic cells, and thus an attractive target for anticancer therapy. Using CellDesigner, a modeling support software for graphical notation, we present herein a comprehensive map of the mTOR signaling network, which includes 964 species connected by 777 reactions. The map complies with both the systems biology markup language (SBML) and graphical notation (SBGN) for computational analysis and graphical representation, respectively. As captured in the mTOR map, we review and discuss our current understanding of the mTOR signaling network and highlight the impact of mTOR feedback and crosstalk regulations on drug-based cancer therapy. This map is available on the Payao platform, a Web 2.0 based community-wide interactive process for creating more accurate and information-rich databases. Thus, this comprehensive map of the mTOR network will serve as a tool to facilitate systems-level study of up-to-date mTOR network components and signaling events toward the discovery of novel regulatory processes and therapeutic strategies for cancer.

Research paper thumbnail of La signalisation RTK/RAS/ERK élargie

médecine/sciences, Jan 1, 2010