Post-transcriptional regulation of Gene Expression
Lead Research Organisation:
University of Edinburgh
Department Name: UNLISTED
Abstract
The flow of genetic information from DNA to RNA to protein involves complex mechanisms of regulation acting downstream of the process of transcription, which produces RNA molecules from a DNA template. Pre-mRNA splicing is the process by which non-coding intervening sequences (introns) are excised from precursor RNAs and coding sequences (exons) are joined to form the mature messenger RNA. A further level of complexity is added by alternative splicing, where a series of different mRNA molecules can be produced by the differential use of splice sites within a pre-mRNA, enabling a single gene to increase its coding capacity. We will study the regulation of Alternative splicing, in particular related to its coupling to the process of transcription. We are investigating a cellular process that controls the quality of RNA produced by cells, termed Nonsense-mediated decay (NMD), which degrades RNAs that encode harmful proteins for the cell. We will focus on a specialised NMD machinery that is localised to the endoplasmic reticulum (ER) and has a role during cellular stress. Finally, we will study the mechanism by which short non-coding RNAs (termed microRNAs) are produced from longer intricately folded precursors and regulate the expression of cellular mRNAs. Our research programme is at the basic end of the spectrum. We expect to contribute to a greater understanding on how the production of cellular RNAs is tightly controlled and how its dysregulation can contribute to human disease. We rely on a variety of experimental systems, including mammalian cell lines in culture, nematodes (C. elegans) and mouse models and we use cell biological, biochemical and single-molecule approaches.
Technical Summary
Gene expression is extensively regulated at the post-transcriptional level. RNAs associate with RNA-binding proteins (RBPs) to form ribonucleoprotein (RNP) complexes. There is a great variety of RBPs, each with unique RNA-binding activity that have a profound impact on gene expression networks, affecting processes as diverse as transcription, RNA splicing, microRNA (miRNA) biogenesis and function and mRNA translation.
The major aim of this programme is to study the mechanisms of post-transcriptional regulation of gene expression. We will address important questions concerning the role of RNA-binding proteins (RBPs) in Gene Expression at different levels during the RNA processing cascade from splicing in the nucleus to translation in the cytoplasm. This proposal will illustrate how alterations in RBP-mediated gene regulation can contribute to human disease.
We will study different aspects of RNA processing, including
- The Nonsense-mediated decay (NMD) pathway is a surveillance mechanism that selectively degrades mRNAs harbouring premature termination codons (PTCs), preventing the synthesis of truncated proteins. This pathway also controls the expression of naturally occurring transcripts and acts to modulate the phenotypic outcome of many inherited genetic disorders that arise as consequence of a frameshift or mutations that generate PTCs. Most studies to date have focused on cytoplasmic NMD; however, it is largely unknown how this mechanism operates on transcripts encoding secreted or ER-resident proteins. We have uncovered evidence that suggests the existence of an NMD pathway dedicated to Endoplasmic Reticulum (ER)-localised mRNAs, which involves two NMD factors that localize to the ER, NBAS and SEC13. We will focus on the functional characterisation of the ER-NMD pathway. We will use a variety of cells in culture, and will rely in cell biological, single molecule and biochemical approaches.
- Alternative splicing (AS) is a major contributor to protein isoform diversity. We will study the regulation of AS, in particular related to its coupling to transcription. We will focus on the role of RNA Polymerase II elongation rate in the regulation of Gene Expression and Alternative splicing in development and during differentiation. We will use mouse models and mouse embryonic stem cells (ESCs), both in an undifferentiated state and during neuronal differentiation.
- Due to the central role of miRNAs in the regulation of gene expression, their levels must be tightly controlled, since dysregulated miRNA expression can result in grossly aberrant gene expression and lead to human disease. We will investigate how RBPs influence miRNA biogenesis and will focus on the role of genetic variation in the production of miRNAs in human populations.
The major aim of this programme is to study the mechanisms of post-transcriptional regulation of gene expression. We will address important questions concerning the role of RNA-binding proteins (RBPs) in Gene Expression at different levels during the RNA processing cascade from splicing in the nucleus to translation in the cytoplasm. This proposal will illustrate how alterations in RBP-mediated gene regulation can contribute to human disease.
We will study different aspects of RNA processing, including
- The Nonsense-mediated decay (NMD) pathway is a surveillance mechanism that selectively degrades mRNAs harbouring premature termination codons (PTCs), preventing the synthesis of truncated proteins. This pathway also controls the expression of naturally occurring transcripts and acts to modulate the phenotypic outcome of many inherited genetic disorders that arise as consequence of a frameshift or mutations that generate PTCs. Most studies to date have focused on cytoplasmic NMD; however, it is largely unknown how this mechanism operates on transcripts encoding secreted or ER-resident proteins. We have uncovered evidence that suggests the existence of an NMD pathway dedicated to Endoplasmic Reticulum (ER)-localised mRNAs, which involves two NMD factors that localize to the ER, NBAS and SEC13. We will focus on the functional characterisation of the ER-NMD pathway. We will use a variety of cells in culture, and will rely in cell biological, single molecule and biochemical approaches.
- Alternative splicing (AS) is a major contributor to protein isoform diversity. We will study the regulation of AS, in particular related to its coupling to transcription. We will focus on the role of RNA Polymerase II elongation rate in the regulation of Gene Expression and Alternative splicing in development and during differentiation. We will use mouse models and mouse embryonic stem cells (ESCs), both in an undifferentiated state and during neuronal differentiation.
- Due to the central role of miRNAs in the regulation of gene expression, their levels must be tightly controlled, since dysregulated miRNA expression can result in grossly aberrant gene expression and lead to human disease. We will investigate how RBPs influence miRNA biogenesis and will focus on the role of genetic variation in the production of miRNAs in human populations.
Publications
Longman D
(2020)
Identification of a localized nonsense-mediated decay pathway at the endoplasmic reticulum.
in Genes & development
López-Perrote A
(2020)
Regulation of RUVBL1-RUVBL2 AAA-ATPases by the nonsense-mediated mRNA decay factor DHX34, as evidenced by Cryo-EM.
in eLife
Related Projects
Project Reference | Relationship | Related To | Start | End | Award Value |
---|---|---|---|---|---|
MC_UU_00007/1 | 31/03/2018 | 30/03/2023 | £662,000 | ||
MC_UU_00007/2 | Transfer | MC_UU_00007/1 | 31/03/2018 | 30/03/2023 | £3,730,000 |
MC_UU_00007/3 | Transfer | MC_UU_00007/2 | 31/03/2018 | 30/05/2022 | £3,053,000 |
MC_UU_00007/4 | Transfer | MC_UU_00007/3 | 31/03/2018 | 30/03/2023 | £1,772,000 |
MC_UU_00007/5 | Transfer | MC_UU_00007/4 | 31/03/2018 | 30/03/2023 | £4,524,000 |
MC_UU_00007/6 | Transfer | MC_UU_00007/5 | 31/03/2018 | 30/03/2023 | £2,878,000 |
MC_UU_00007/7 | Transfer | MC_UU_00007/6 | 31/03/2018 | 30/03/2023 | £2,829,000 |
MC_UU_00007/8 | Transfer | MC_UU_00007/7 | 31/03/2018 | 31/12/2022 | £4,072,000 |
MC_UU_00007/9 | Transfer | MC_UU_00007/8 | 31/03/2018 | 30/03/2023 | £3,137,000 |
MC_UU_00007/10 | Transfer | MC_UU_00007/9 | 31/03/2018 | 30/03/2023 | £6,948,000 |
MC_UU_00007/11 | Transfer | MC_UU_00007/10 | 31/03/2018 | 30/03/2023 | £2,421,000 |
MC_UU_00007/12 | Transfer | MC_UU_00007/11 | 31/03/2018 | 30/03/2023 | £1,205,000 |
MC_UU_00007/13 | Transfer | MC_UU_00007/12 | 31/03/2018 | 30/03/2023 | £1,174,000 |
MC_UU_00007/14 | Transfer | MC_UU_00007/13 | 31/03/2018 | 30/03/2023 | £1,838,000 |
MC_UU_00007/15 | Transfer | MC_UU_00007/14 | 31/03/2018 | 30/03/2023 | £2,551,000 |
MC_UU_00007/16 | Transfer | MC_UU_00007/15 | 31/03/2018 | 30/03/2023 | £1,496,000 |
MC_UU_00007/17 | Transfer | MC_UU_00007/16 | 31/03/2018 | 30/03/2023 | £1,886,000 |
Description | Llorca-Structural studies-Electron Microscopy (EM)-Nonsense-mediated decay (NMD) |
Organisation | Spanish National Cancer Research Center |
Country | Spain |
Sector | Public |
PI Contribution | Experimental design, biochemical experiments |
Collaborator Contribution | Experimental design, structural analysis, electron microscopy |
Impact | 1. Melero,R.±, Hug,N.±, Lopez-Perrote,A., Yamashita,A., Cáceres,J.F.* and Llorca,O*. (2016) The RNA helicase DHX34 functions as a scaffold for SMG1-mediated UPF1 phosphorylation. Nat. commun., 7,10585 | doi: 10.1038/ncomms10585 ±Joint First authors/*Co-corresponding authors 2. Lopez-Perrote A, Hug N, González-Corpas A, Rodriguez CF, Serna M, Garcia-Martin C, Boskovic J, Fernandez-Leiro R, Cáceres JF, Llorca O (2020) Regulation of RUVBL1-RUVBL2 AAA-ATPases by the nonsense-mediated decay factor DHX34, as evidenced by Cryo-EM. eLife 9: e63042. doi: 10.7554/eLife.63042. |
Start Year | 2013 |
Description | Michael Sattler-Structural studies-Crystallography-NMR of factors involved in biogenesis of miRNAs |
Organisation | Technical University of Munich |
Country | Germany |
Sector | Academic/University |
PI Contribution | Experimental design, biochemical experiments |
Collaborator Contribution | Experimental design, structural analysis, Crystallography, NMR |
Impact | Kooshapur,H., Choudhury,N.R., Simon,B., Mühlbauer,M., Jussupow,A., Fernandez,N., Dallmann,A., Gabel,F., Camilloni,C., Michlewski,G.*, Cáceres,J.F.* and Sattler,M.* (2018) Structural basis for terminal loop recognition and processing of pri-miRNA-18a by hnRNP A1. Nat commun., under revision *Co-corresponding authors |
Start Year | 2011 |
Description | Role of RNA helicases associated with Nonsense-mediated decay (NMD) and pre-mRNA splicing in Acute myeloid leukemia (AML) |
Organisation | Queen Mary University of London |
Department | Barts Cancer Institute |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | We are collaborating with the group of Professor Jude Fitzgibbon at the Barts Cancer Institute. His group is characterising novel genes mutated in familial AML (Acute myeloid leukemia) /MDS (Myelodysplastic syndrome). They have identified families harbouring variants in DHX34, an RNA helicase that my lab has shown to function in the Nonsense-mediated RNA decay (NMD) pathway and to co-purify with the spliceosome, suggesting its potential involvement in pre-mRNA splicing. |
Collaborator Contribution | We are providing functional assays to investigate whether mutations in DHX34 found in AML/MDS patients affect the function of DHX34 in Nonsense-mediated decay (NMD) and/or in pre-mRNA splicing |
Impact | 1] There is an Abstract to be presented at the European Hematology Association in Amsterdam in June 2019 Abstract Submission 3. Acute myeloid leukemia - Biology & Translational Research EHA-2619 THE GENETIC LANDSCAPE OF FAMILIAL MDS/AML; RECURRING MUTATIONS IN THE RNA HELICASE DHX34 LEADING TO DEFECTS IN NONSENSE-MEDIATED RNA DECAY Ana Rio-Machin* 1, Tom Vulliamy1, Nele Hug2, Javier F Caceres2, Jude Fitzgibbon1, Inderjeet Dokal1 1Queen Mary University of London, London, 2University of Edinburgh, Edinburgh, United Kingdom 2 MRC Human Genetics Unit, University of Edinburgh 2] There have been two publications 2.1. Rio-Machin A, Vulliamy T, Hug N, Walne A, Tawana K, Cardoso S, Ellison A, Pontikos N, Wang J, Tummala H, Al Seraihi AFH, Alnajar J, Bewicke-Copley F, Armes H, Barnett M, Bloor A, Bodor C, Bowen D, Fenaux P, Green A, Hallahan A, Hjorth-Hansen H, Hossain U, Killick S, Lawson S, Layton M, Male AM, Marsh J, Mehta P, Mous R, Nomdedéu JF, Owen C, Pavlu J, Payne EM, Protheroe RE, Preudhomme C, Pujol-Moix N, Renneville A, Russell N, Saggar A, Sciuccati G, Taussig D, Toze CL, Uyttebroeck A, Vandenberghe P, Schlegelberger B, Ripperger T, Steinemann D, Wu J, Mason J, Page P, Akiki S, Reay K, Cavenagh JD, Plagnol V, Cáceres JF, Fitzgibbon J Dokal I (2020) The complex genetic landscape of familial MDS and AML reveals pathogenic germline variants associated with the disease. Nat Commun 11: 1044. PMID: 32098966 PMCID: PMC7042299 DOI: 10.1038/s41467-020-14829-5 2.2. Hug N, Aitken S, Longman D, Raab M, Armes H, Mann AR, Rio-Machin A, Fitzgibbon J, Rouault-Pierre K, Cáceres JF (2022) A dual role for the RNA helicase DHX34 in NMD and pre-mRNA splicing and its function in hematopoietic differentiation. RNA 28: 1224-1238. PMID: 35768279 PMCID: PMC9380745 DOI: 10.1261/rna.079277.122 |
Start Year | 2018 |
Description | Role of the Nonsense-mediated pathway in models of Neurodegeneration in nematodes |
Organisation | University of Edinburgh |
Department | Centre for Integrative Physiology |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | We have uncovered evidence that suggests the existence of a branch of the Nonsense-mediated decay pathway that is dedicated to Endoplasmic Reticulum (ER)-localized mRNAs. We are focusing on a novel NMD factor that we identified in our lab, which is termed NBAS, for Neuroblatoma-amplified gene. Given the established link between NMD and ER stress, we are investigating whether there is a protective role for the NMD pathway in pathological conditions where there is chronic activation of the Unfolded Protein Response (UPR), using established models for this process in neurodegenerative disease. In particular, in this collaboration we are focusing on models of Neurodegeneration in the nematode, C. elegans. |
Collaborator Contribution | The lab of Maria Doitsidou is providing her expertise in C.elegans and in models of Neurodegeneration in this organism |
Impact | We generated a series of integrated transgenic C. elegans strains with pan-neuronal overexpression of core and ER-NMD factors, including nematode and human versions of SMG-2/UPF1; SMG-3/UPF2; SMGL-1/NBAS and NPP-20/SEC13. These strains will be crossed with established models of ALS, where worms with neuronal expression of a mutant form of hTDP-43 (G290A) develop severe progressive paralysis and exhibit neurodegeneration of motor neurons. We will monitor the effect of increased expression of NMD factors in neurons on the mobility and neurodegenerative phenotypes of ALS strains. |
Start Year | 2018 |
Description | Broughton high School: Genetic Testing & Whole Genome Sequencing (WGS) Workshop |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Schools |
Results and Impact | Our postdoc and student helped develop and deliver a workshop to 90+ S5/6 (15-17 years old) local High School Students on Genetic Testing & Whole Genome Sequencing. Students did small group activities and learned more about the technologies now available, and careers related to these. The students debated pros and cons of all people having their genome sequenced. Using mentimeter we probed their responses at the beginning and end of the session to the question "would you get your genome sequenced". Interestingly at the end of the session more pupils said they would have their genome sequenced. Following this visit one student attended the MRC IGMM for a tour and career discussion while applying for biomedical degree courses. |
Year(s) Of Engagement Activity | 2019 |
URL | https://twitter.com/MRC_IGMM/status/1187418436256894976 |