6 research outputs found
GC content shapes mRNA storage and decay in human cells.
mRNA translation and decay appear often intimately linked although the rules of this interplay are poorly understood. In this study, we combined our recent P-body transcriptome with transcriptomes obtained following silencing of broadly acting mRNA decay and repression factors, and with available CLIP and related data. This revealed the central role of GC content in mRNA fate, in terms of P-body localization, mRNA translation and mRNA stability: P-bodies contain mostly AU-rich mRNAs, which have a particular codon usage associated with a low protein yield; AU-rich and GC-rich transcripts tend to follow distinct decay pathways; and the targets of sequence-specific RBPs and miRNAs are also biased in terms of GC content. Altogether, these results suggest an integrated view of post-transcriptional control in human cells where most translation regulation is dedicated to inefficiently translated AU-rich mRNAs, whereas control at the level of 5' decay applies to optimally translated GC-rich mRNAs
Molecular and cellular mechanisms of formation and dissolution of "processing body" under physiological and physiopathological conditions
Les « processing bodies » ou p-bodies sont des structures sans membranes (MLO : Membraneless Organelles) formĂ©es par un processus connu sous le nom de sĂ©paration de phase liquide-liquide (LLPS). Les p-bodies sont des agrĂ©gats de protĂ©ines et dâARNs localisĂ©s dans le cytoplasme des cellules. Ils ont Ă©tĂ© caractĂ©risĂ©s comme Ă©tant un lieu de stockage des ARNs rĂ©primĂ©s, puisque les ARNs qui y sont accumulĂ©s, sont stables et non traduits. Pour autant, lâabsence de membrane rend leur contenu rapidement mobilisable par les cellules, suggĂ©rant un rĂŽle important dans les rĂ©ponses immĂ©diates au stress. Au cours de ma thĂšse, je me suis intĂ©ressĂ©e, Ă la fois au mĂ©canisme de formation et de dissolution des p-bodies. Tout dâabord, jâai mis en Ă©vidence lâeffet de la dissolution des p-bodies sur les ARNm. Lâinduction de la dissolution des p-bodies conduit Ă une augmentation de la traduction des ARNm qui y sont stockĂ©s et rĂ©primĂ©s tels que les oncogĂšnes KRAS et NRAS. Il semble alors, que ce rĂ©servoir dâARNm constitue un mĂ©canisme dynamique dâadaptation au stress cellulaire, et en particulier pour le dĂ©veloppement de cellules rĂ©sistantes aux MEKi. Ensuite, afin de mieux comprendre la formation des p-bodies, jâai Ă©tudiĂ© la localisation de lâexoribonuclĂ©ase 1 (XRN1). Cette enzyme comme son nom lâindique est impliquĂ©e dans la dĂ©gradation des ARNs, mais est retrouvĂ©e accumulĂ©e dans les p-bodies sous forme inactive. Dans ce contexte, afin de dĂ©terminer le rĂŽle de la protĂ©ine XRN1 dans les p-bodies, je me suis intĂ©ressĂ©e aux rĂ©gions importantes pour son adressage dans les p-bodies. Ainsi, jâai pu montrer que les zones permettant lâaccumulation de XRN1 dans les p-bodies se trouvent dans la partie C-terminale de la protĂ©ine au niveau des acides aminĂ©s 1406-1468 et 1607-1706. Ces rĂ©sultats indiquent donc que ces deux rĂ©gions conservĂ©es chez les vertĂ©brĂ©s sont indispensables Ă la localisation de la protĂ©ine de façon indĂ©pendante de la partie catalytique situĂ©e en N-terminal. En conclusion, jâai, au cours de ma thĂšse, travaillĂ© sur deux aspects des p-bodies : la sĂ©questration protĂ©ique et la sĂ©questration des ARNs. Ces travaux permettent de mieux comprendre le rĂŽle jouĂ© par p-bodies dans des processus physiologiques ou physiopathologiques tels que le cancer.« Processing bodies » or p-bodies are Membraneless Organelles (MLO) formed by a processus called liquid liquid phase separation (LLPS). P-bodies are aggregates of proteins and RNAs localized in cell cytoplasm. They were characterized as a storage place of repressed RNAs, without degradation nor decay. However, membraneless properties makes their content quickly mobilizable by cells, suggesting an important role for immediate responses to stress. During my thesis, I was interested both in mechanisms of p-bodies formation and dissolution. First, I highlighted the impact of p-bodies dissolution on mRNAs translation. Indeed, P-bodies dissolution induction leads to increased translation of these repressed and stored mRNAs, like KRAS and NRAS oncogenes. It seems that this mRNA pool constitutes a dynamic adaptation mechanism to cellular stress, in particular for MEKi drug tolerant cancer cell onset. In a second part of my thesis, in order to understand p-bodies formation, I studied exoribonuclease 1 (XRN1) localization. This crucial enzyme for RNAs decay is also found accumulated in p-bodies in an inactive form. In this context, to determine XRN1 role in p-bodies, I have been interested in p-bodies localization signal regions. Thus, I showed that region leading to XRN1 accumulation are found in C-terminal part of the protein at amino acid level: 1406-1468 et 1607-1706. These results indicate that these two conserved regions in vertebrates are indispensable for protein localization independently of the catalytic part localized in N-terminal part of XRN1 protein. To conclude, during my thesis I have worked on two p-bodies aspects: protein and RNA sequestration. These data will contribute to better understand p-bodies role in physiological and physiopathological diseases like cancer
Mécanismes moléculaires et cellulaires de formation et de dissolution des « processing bodies » en conditions physiologiques et physiopathologiques
« Processing bodies » or p-bodies are Membraneless Organelles (MLO) formed by a processus called liquid liquid phase separation (LLPS). P-bodies are aggregates of proteins and RNAs localized in cell cytoplasm. They were characterized as a storage place of repressed RNAs, without degradation nor decay. However, membraneless properties makes their content quickly mobilizable by cells, suggesting an important role for immediate responses to stress. During my thesis, I was interested both in mechanisms of p-bodies formation and dissolution. First, I highlighted the impact of p-bodies dissolution on mRNAs translation. Indeed, P-bodies dissolution induction leads to increased translation of these repressed and stored mRNAs, like KRAS and NRAS oncogenes. It seems that this mRNA pool constitutes a dynamic adaptation mechanism to cellular stress, in particular for MEKi drug tolerant cancer cell onset. In a second part of my thesis, in order to understand p-bodies formation, I studied exoribonuclease 1 (XRN1) localization. This crucial enzyme for RNAs decay is also found accumulated in p-bodies in an inactive form. In this context, to determine XRN1 role in p-bodies, I have been interested in p-bodies localization signal regions. Thus, I showed that region leading to XRN1 accumulation are found in C-terminal part of the protein at amino acid level: 1406-1468 et 1607-1706. These results indicate that these two conserved regions in vertebrates are indispensable for protein localization independently of the catalytic part localized in N-terminal part of XRN1 protein. To conclude, during my thesis I have worked on two p-bodies aspects: protein and RNA sequestration. These data will contribute to better understand p-bodies role in physiological and physiopathological diseases like cancer.Les « processing bodies » ou p-bodies sont des structures sans membranes (MLO : Membraneless Organelles) formĂ©es par un processus connu sous le nom de sĂ©paration de phase liquide-liquide (LLPS). Les p-bodies sont des agrĂ©gats de protĂ©ines et dâARNs localisĂ©s dans le cytoplasme des cellules. Ils ont Ă©tĂ© caractĂ©risĂ©s comme Ă©tant un lieu de stockage des ARNs rĂ©primĂ©s, puisque les ARNs qui y sont accumulĂ©s, sont stables et non traduits. Pour autant, lâabsence de membrane rend leur contenu rapidement mobilisable par les cellules, suggĂ©rant un rĂŽle important dans les rĂ©ponses immĂ©diates au stress. Au cours de ma thĂšse, je me suis intĂ©ressĂ©e, Ă la fois au mĂ©canisme de formation et de dissolution des p-bodies. Tout dâabord, jâai mis en Ă©vidence lâeffet de la dissolution des p-bodies sur les ARNm. Lâinduction de la dissolution des p-bodies conduit Ă une augmentation de la traduction des ARNm qui y sont stockĂ©s et rĂ©primĂ©s tels que les oncogĂšnes KRAS et NRAS. Il semble alors, que ce rĂ©servoir dâARNm constitue un mĂ©canisme dynamique dâadaptation au stress cellulaire, et en particulier pour le dĂ©veloppement de cellules rĂ©sistantes aux MEKi. Ensuite, afin de mieux comprendre la formation des p-bodies, jâai Ă©tudiĂ© la localisation de lâexoribonuclĂ©ase 1 (XRN1). Cette enzyme comme son nom lâindique est impliquĂ©e dans la dĂ©gradation des ARNs, mais est retrouvĂ©e accumulĂ©e dans les p-bodies sous forme inactive. Dans ce contexte, afin de dĂ©terminer le rĂŽle de la protĂ©ine XRN1 dans les p-bodies, je me suis intĂ©ressĂ©e aux rĂ©gions importantes pour son adressage dans les p-bodies. Ainsi, jâai pu montrer que les zones permettant lâaccumulation de XRN1 dans les p-bodies se trouvent dans la partie C-terminale de la protĂ©ine au niveau des acides aminĂ©s 1406-1468 et 1607-1706. Ces rĂ©sultats indiquent donc que ces deux rĂ©gions conservĂ©es chez les vertĂ©brĂ©s sont indispensables Ă la localisation de la protĂ©ine de façon indĂ©pendante de la partie catalytique situĂ©e en N-terminal. En conclusion, jâai, au cours de ma thĂšse, travaillĂ© sur deux aspects des p-bodies : la sĂ©questration protĂ©ique et la sĂ©questration des ARNs. Ces travaux permettent de mieux comprendre le rĂŽle jouĂ© par p-bodies dans des processus physiologiques ou physiopathologiques tels que le cancer
Mécanismes moléculaires et cellulaires de formation et de dissolution des « processing bodies » en conditions physiologiques et physiopathologiques
« Processing bodies » or p-bodies are Membraneless Organelles (MLO) formed by a processus called liquid liquid phase separation (LLPS). P-bodies are aggregates of proteins and RNAs localized in cell cytoplasm. They were characterized as a storage place of repressed RNAs, without degradation nor decay. However, membraneless properties makes their content quickly mobilizable by cells, suggesting an important role for immediate responses to stress. During my thesis, I was interested both in mechanisms of p-bodies formation and dissolution. First, I highlighted the impact of p-bodies dissolution on mRNAs translation. Indeed, P-bodies dissolution induction leads to increased translation of these repressed and stored mRNAs, like KRAS and NRAS oncogenes. It seems that this mRNA pool constitutes a dynamic adaptation mechanism to cellular stress, in particular for MEKi drug tolerant cancer cell onset. In a second part of my thesis, in order to understand p-bodies formation, I studied exoribonuclease 1 (XRN1) localization. This crucial enzyme for RNAs decay is also found accumulated in p-bodies in an inactive form. In this context, to determine XRN1 role in p-bodies, I have been interested in p-bodies localization signal regions. Thus, I showed that region leading to XRN1 accumulation are found in C-terminal part of the protein at amino acid level: 1406-1468 et 1607-1706. These results indicate that these two conserved regions in vertebrates are indispensable for protein localization independently of the catalytic part localized in N-terminal part of XRN1 protein. To conclude, during my thesis I have worked on two p-bodies aspects: protein and RNA sequestration. These data will contribute to better understand p-bodies role in physiological and physiopathological diseases like cancer.Les « processing bodies » ou p-bodies sont des structures sans membranes (MLO : Membraneless Organelles) formĂ©es par un processus connu sous le nom de sĂ©paration de phase liquide-liquide (LLPS). Les p-bodies sont des agrĂ©gats de protĂ©ines et dâARNs localisĂ©s dans le cytoplasme des cellules. Ils ont Ă©tĂ© caractĂ©risĂ©s comme Ă©tant un lieu de stockage des ARNs rĂ©primĂ©s, puisque les ARNs qui y sont accumulĂ©s, sont stables et non traduits. Pour autant, lâabsence de membrane rend leur contenu rapidement mobilisable par les cellules, suggĂ©rant un rĂŽle important dans les rĂ©ponses immĂ©diates au stress. Au cours de ma thĂšse, je me suis intĂ©ressĂ©e, Ă la fois au mĂ©canisme de formation et de dissolution des p-bodies. Tout dâabord, jâai mis en Ă©vidence lâeffet de la dissolution des p-bodies sur les ARNm. Lâinduction de la dissolution des p-bodies conduit Ă une augmentation de la traduction des ARNm qui y sont stockĂ©s et rĂ©primĂ©s tels que les oncogĂšnes KRAS et NRAS. Il semble alors, que ce rĂ©servoir dâARNm constitue un mĂ©canisme dynamique dâadaptation au stress cellulaire, et en particulier pour le dĂ©veloppement de cellules rĂ©sistantes aux MEKi. Ensuite, afin de mieux comprendre la formation des p-bodies, jâai Ă©tudiĂ© la localisation de lâexoribonuclĂ©ase 1 (XRN1). Cette enzyme comme son nom lâindique est impliquĂ©e dans la dĂ©gradation des ARNs, mais est retrouvĂ©e accumulĂ©e dans les p-bodies sous forme inactive. Dans ce contexte, afin de dĂ©terminer le rĂŽle de la protĂ©ine XRN1 dans les p-bodies, je me suis intĂ©ressĂ©e aux rĂ©gions importantes pour son adressage dans les p-bodies. Ainsi, jâai pu montrer que les zones permettant lâaccumulation de XRN1 dans les p-bodies se trouvent dans la partie C-terminale de la protĂ©ine au niveau des acides aminĂ©s 1406-1468 et 1607-1706. Ces rĂ©sultats indiquent donc que ces deux rĂ©gions conservĂ©es chez les vertĂ©brĂ©s sont indispensables Ă la localisation de la protĂ©ine de façon indĂ©pendante de la partie catalytique situĂ©e en N-terminal. En conclusion, jâai, au cours de ma thĂšse, travaillĂ© sur deux aspects des p-bodies : la sĂ©questration protĂ©ique et la sĂ©questration des ARNs. Ces travaux permettent de mieux comprendre le rĂŽle jouĂ© par p-bodies dans des processus physiologiques ou physiopathologiques tels que le cancer
KRAS and NRAS Translation Is Increased upon MEK Inhibitors-Induced Processing Bodies Dissolution
Overactivation of the mitogen-activated protein kinase (MAPK) pathway is a critical driver of many human cancers. However, therapies directly targeting this pathway lead to cancer drug resistance. Resistance has been linked to compensatory RAS overexpression, but the mechanisms underlying this response remain unclear. Here, we find that MEK inhibitors (MEKi) are associated with an increased translation of the KRAS and NRAS oncogenes through a mechanism involving dissolution of processing body (P-body) biocondensates. This effect is seen across different cell types and is extremely dynamic since removal of MEKi and ERK reactivation result in reappearance of P-bodies and reduced RAS-dependent signaling. Moreover, we find that P-body scaffold protein levels negatively impact RAS expression. Overall, we describe a new feedback loop mechanism involving biocondensates such as P-bodies in the translational regulation of RAS proteins and MAPK signaling
GC content shapes mRNA decay and storage in human cells
Control of protein expression results from the fine tuning of mRNA synthesis, decay and translation. These processes, which are controlled by a large number of RNA-binding proteins and by localization in RNP granules such as P-bodies, appear often intimately linked although the rules of this interplay are not well understood. In this study, we combined our recent P-body transcriptome with various transcriptomes obtained following silencing of broadly acting mRNA decay and repression factors. This analysis revealed the central role of GC content in mRNA fate, in terms of P-body localization, mRNA translation and mRNA decay. It also rationalized why PBs mRNAs have a strikingly low protein yield. We report too the existence of distinct mRNA decay pathways with preference for AU-rich or GC-rich transcripts. Compared to this impact of the GC content, sequence-specific RBPs and miRNAs appeared to have only modest additional effects on their bulk targets. Altogether, these results lead to an integrated view of post-transcriptional control in human cells where most regulation at the level of translation is dedicated to AU-rich mRNAs, which have a limiting protein yield, whereas regulation at the level of 5' decay applies to GC-rich mRNAs, whose translation is optimal