Periodic Reporting for period 3 - MyeRIBO (Deconstructing the Translational Control of Myelination by Specialized Ribosomes)
Okres sprawozdawczy: 2022-07-01 do 2023-12-31
The myelin sheath is essential for neuronal function and health: myelinating glial cells speed up propagation of axonal potentials, fuel the energetic demands and regulate the ionic environment of neurons. Myelination involves a striking expansion of the glial cell membrane that relies on an exceptional increase in protein and lipid synthesis rates. Decades of dedicated research have uncovered a complex transcriptional program, driven by an intricate network of transcription factors, epigenetic modifiers and non-coding RNAs, that coordinate this process. In contrast, much less is known about the translational control of myelination. Studies directed at exploring specific mechanisms regulating mRNA translation during myelination have barely been pursued, nor have their implications in myelin disorders been examined. The mission of MyeRIBO is to deconstruct the translational control of myelination and examine its implication in myelin disorders.
There is emerging evidence, enabled by modern techniques, that ribosomes, typically viewed as invariant, passive molecular machines, may instead be heterogeneous in composition, with particular ribosomal components having a ‘specialized’ regulatory capacity for preferential translation of specific mRNAs. In MyeRIBO, I propose that translation control by specialized ribosomes is a novel layer of regulation that shapes the proteome of the myelinating glial cell. MyeRIBO is designed to discover the nature and diversity of the ribosome population in myelinating Schwann cells, and the mechanistic and functional role of these specialized ribosomes in regulating myelination. Building on these results, I will determine the contribution of ribosome heterogeneity in pathogenesis of myelin disorders.
Lesions to the myelin sheath result in devastating neurological disorders that include multiple sclerosis, diabetic neuropathy and Charcot-Marie-Tooth disease. Unfortunately, no therapeutical strategies are available for the vast majority of demyelinating neuropathies, as the precise molecular mechanisms of disease progression and functional failure remain poorly understood. MyeRIBO will push further the boundaries of our current understanding of the molecular control of myelination, which could have a profound impact for understanding neural development and myelin disorders.
There is emerging evidence, enabled by modern techniques, that ribosomes, typically viewed as invariant, passive molecular machines, may instead be heterogeneous in composition, with particular ribosomal components having a ‘specialized’ regulatory capacity for preferential translation of specific mRNAs. In MyeRIBO, I propose that translation control by specialized ribosomes is a novel layer of regulation that shapes the proteome of the myelinating glial cell. MyeRIBO is designed to discover the nature and diversity of the ribosome population in myelinating Schwann cells, and the mechanistic and functional role of these specialized ribosomes in regulating myelination. Building on these results, I will determine the contribution of ribosome heterogeneity in pathogenesis of myelin disorders.
Lesions to the myelin sheath result in devastating neurological disorders that include multiple sclerosis, diabetic neuropathy and Charcot-Marie-Tooth disease. Unfortunately, no therapeutical strategies are available for the vast majority of demyelinating neuropathies, as the precise molecular mechanisms of disease progression and functional failure remain poorly understood. MyeRIBO will push further the boundaries of our current understanding of the molecular control of myelination, which could have a profound impact for understanding neural development and myelin disorders.
During this first period of the proposal, we have addressed our outstanding question of ribosome heterogeneity during Schwann cell differentiation. We uncovered extensive remodelling of actively translating ribosome composition, and in particular, identified a specific PTM that has profound effects on Schwann cell myelination and in demyelinating pathologies, through a mechanism that could involve changes in ribosome function (manuscript under preparation).
In summary, we have made significant progress in the following areas:
As proposed, we performed quantitative mass spectrometry (SWATH-MS) of purified ribosomal fractions from Schwann cells cultured under control and myelinogenic conditions and found widespread differences in ribosome composition in myelinating Schwann cells, as well as dynamic changes as these cells differentiate. We identified several ribosomal proteins that were substoichiometric in the actively translating polysomes in these cells, and showed dynamic changes compared to control cells or even when monosomes were compared to polysomes in the same cell type. In addition, we also identified several ribosome associated proteins (RAPs; proteins that associate physically with ribosomes connecting the ribosome to other cell signalling pathways but also enabling the translation of specific RNAs) that are differentially expressed in myelinating conditions comparted to control conditions. Most of the proteins appear exclusively in one of the groups (Monosomes, Light polysomes or heavy polysomes), indicating that the system is very heterogeneous and supports the theory that each of these groups may be translating different transcripts. Several of these RAPs are associated with specific protein classes that we are further pursuing. We are at present validating these findings obtained from cultured cells, in vivo using genetic models and are designing the tools to study the biological and functional role of these ribosomal proteins and RAPs in selective mRNA translation during Schwann cell myelination.
As a specific example of ribosomal specialization, we identified a RAP as an enzyme that catalyses a particular post-translational modification, which we have identified to be a key regulator of Schwann cell myelination and is implicated in demyelinating disorders (manuscript in preparation). Our aim now is to discover whether this PTM regulates ribosome function and selective mRNA translation that we would connect to our findings on its functional role in Schwann cell biology and pathology. This is an unplanned result but has opened an exciting line of research in the lab that we are currently actively pursuing.
In addition, we report other publications e.g. reviews or the development of complementary projects by our collaborators. In turn, we have several other exciting projects related to other gene control mechanisms that are ongoing and will advance our understanding on myelin biology.
In summary, we have made significant progress in the following areas:
As proposed, we performed quantitative mass spectrometry (SWATH-MS) of purified ribosomal fractions from Schwann cells cultured under control and myelinogenic conditions and found widespread differences in ribosome composition in myelinating Schwann cells, as well as dynamic changes as these cells differentiate. We identified several ribosomal proteins that were substoichiometric in the actively translating polysomes in these cells, and showed dynamic changes compared to control cells or even when monosomes were compared to polysomes in the same cell type. In addition, we also identified several ribosome associated proteins (RAPs; proteins that associate physically with ribosomes connecting the ribosome to other cell signalling pathways but also enabling the translation of specific RNAs) that are differentially expressed in myelinating conditions comparted to control conditions. Most of the proteins appear exclusively in one of the groups (Monosomes, Light polysomes or heavy polysomes), indicating that the system is very heterogeneous and supports the theory that each of these groups may be translating different transcripts. Several of these RAPs are associated with specific protein classes that we are further pursuing. We are at present validating these findings obtained from cultured cells, in vivo using genetic models and are designing the tools to study the biological and functional role of these ribosomal proteins and RAPs in selective mRNA translation during Schwann cell myelination.
As a specific example of ribosomal specialization, we identified a RAP as an enzyme that catalyses a particular post-translational modification, which we have identified to be a key regulator of Schwann cell myelination and is implicated in demyelinating disorders (manuscript in preparation). Our aim now is to discover whether this PTM regulates ribosome function and selective mRNA translation that we would connect to our findings on its functional role in Schwann cell biology and pathology. This is an unplanned result but has opened an exciting line of research in the lab that we are currently actively pursuing.
In addition, we report other publications e.g. reviews or the development of complementary projects by our collaborators. In turn, we have several other exciting projects related to other gene control mechanisms that are ongoing and will advance our understanding on myelin biology.
Our research has unravelled many sources of ribosome heterogeneity in myelinating Schwann cells and show that there are dynamic changes in this heterogeneity as the Schwann cells differentiate from their progenitors, implicating specialized mRNA translation in the regulation of the myelination process. Our aim for the next period of the proposal is to decipher the biological and functional role of these specialized ribosomal components in myelination as well as their implications in demyelinating pathologies. We have also already identified a key PTM that is a novel essential regulator of Schwann cell myelination and is implicated in myelin disorders, both breakthrough findings in our field of research. Our aim now is to discover whether this PTM is involved in modulating ribosome function in myelinating Schwann cells, as our studies suggest, thus regulating selective mRNA translation in myelination and in myelin disorders.