Mechanisms and function of alternative splicing in the plant circadian clock

Lead Research Organisation: University of Dundee
Department Name: School of Life Sciences

Abstract

Circadian rhythms are ubiquitous in nature: most organisms exhibit robust daily rhythms in biological processes, growth or behaviour. The most familiar rhythm to us is our sleep/wake cycle which is set to local time and gives rise to jet lag when time-zones are traversed. Rhythms are driven by 'circadian clocks' located in every living cell that continue to run even in the absence of external signals. These biological devices allow an organism to anticipate predictable environmental changes such as the regular day to night transitions and to adjust its behaviour accordingly. Circadian clocks comprise molecular circuits organised into feedback loops that generate the oscillatory behaviour of clock components. By linking to different molecular pathways through the cycle, the clock is able to regulate the timing of key pathways (e.g. metabolic, signalling, growth and development) through the day. Clockwork failure can have a large fitness costs to the organism; in humans it contributes to several disease states.

Circadian clocks can operate over a temperature range. This is particularly important for plants that are exposed to the ever-changing environment. Chemical reaction rates are very sensitive to temperature change, yet the clock remains invariant in the face of daily temperature changes and a range of weather patterns through the seasons. Yet how this is achieved is not known. A key objective for this project is therefore to understand molecular processes that buffer the plant clock from temperature changes. This is important as maintaining clock function is essential for optimal photosynthesis, growth, and the timing of reproduction - factors that influence seed abundance (a yield output for crop plants).

When genes are expressed the DNA sequence is first copied into RNA (transcription), the RNA is processed and then it directs synthesis of the corresponding protein (translation). In this project we focus on deciphering the role of post-transcriptional RNA processing (alternative splicing: AS) in temperature-dependent clock function in the model plant Arabidopsis. AS generates different transcripts from the same gene and thereby can modulate transcript and protein levels and functions. We have shown that AS is important in controlling clock gene expression. We will establish the extent of AS in clock input genes, how this is affected at low temperature and their influence expression/AS of clock genes. Plants experience continued and variable temperature changes throughout the day/night cycle - we will investigate the degree and timing of temperature change which is able to elicit a temperature-dependent AS response. Light also has a major influence on the clock and we will identify which AS events respond to light intensity changes and whether they are different from temperature-dependent events.

A major question is how the different types of AS in different clock genes are regulated and how this mechanism contributes to temperature buffering of the circadian clockwork. We will use RNA sequencing to assess AS events during cooling. Co-expression and co-splicing network analysis will identify genes whose expression/splicing profiles correlate with those of the core clock and clock-associated genes to identify putative regulatory genes. We will also examine the natural genetic variation in this process and how this might aid our understanding.

The role of AS in regulation of clock gene expression is highly relevant to crop plants and yield. To utilise the knowledge and approaches from our Arabidopsis research, we have started to examine AS control in potato and barley to begin the translation toward application. We anticipate that this work will provide new insights into what controls phenotypes such as earliness in barley and endodormancy in potato and ultimately lead to strategies for the generation of new genotypes that have increased robustness to temperature and other stresses that can affect plants.

Technical Summary

Molecular clock circuits comprise a set of interlocked transcriptional feedback loops with imposed delays that generate a characteristic ~24h rhythm. Its machinery is controlled by several mechanisms, including gene expression, chromatin remodelling, protein phosphorylation and protein turnover. Our recent work has added a new dimension by showing that alternative splicing (AS) can play a significant role in determining the level of functional transcripts/proteins of key clock genes, particularly in rapid or long-term responses to temperature change.

We will identify AS in clock-associated genes which input signals to the clock and in selected regulatory genes. This will generate a panel of primers covering key AS events which will be used in our sensitive HR RT-PCR system to address different aspects of the project. The functional relevance of AS will be tested at the promoter, transcript, protein and whole plant levels using clock mutant lines complemented with gene versions where AS is compromised or limited. These lines will be analysed for physiological and molecular phenotypes. To address what factors regulate AS of the core clock and clock-associated genes, and identify the downstream effects of reduced clock protein levels at lower temperatures, we need a genome-wide approach. We will perform deep RNA-sequencing across multiple time-points in the diurnal cycle before and after transfer to low temperature. This will generate expression and AS information on genes expressed under these conditions. Gene expression and splicing network analysis will identify candidate genes which may regulate or be regulated by the clock. Studies of natural variation may pinpoint other regulatory genes. We will characterise selected regulatory candidate genes (e.g. splicing factors) using overexpressing lines or lines that carry mutations and testing for effects on AS in clock genes.

Planned Impact

Background
The research in this proposal is basic science on how alternative splicing (AS) regulates gene expression and thereby function in the circadian clock and its responses to external cues. Although at this stage the research is relatively far removed from direct application, the clock is so important for optimising plant growth and development in a changing environment that it is directly relevant to crop performance in the field. We have already made significant progress towards addressing similar questions in crop plants (potato and barley) in collaboration with scientists at the James Hutton Institute.

Who will benefit?
Understanding the molecular mechanisms that regulate the clock in a constantly changing environment could lead to new crop improvement strategies that mitigate the impact of predicted medium-term changes in seasonal temperatures, rainfall etc. The circadian clock is important to agricultural crops as it influences a range of processes that are important for productivity such as flowering time, starch production, disease resistance, stomatal movements, responses to stress and lignification. Understanding the basic molecular regulation of the clock will allow us to establish whether crop growth and productivity can be enhanced by controlling clock function. The work will therefore be of interest to crop scientists and plant biotechnology companies working on phenotypic traits and the underpinning genetics in crop species, and to government bodies responsible for future-proofing food production. Many of the principles governing the function of circadian clocks are broadly applicable across species; furthermore, there is an interactive chronobiology community which discusses ideas from many organisms. Hence our work is of potential interest to diverse communities such as human sleep researchers. The work will also be of interest to individuals (e.g. authors of textbooks) and organisations (e.g. Glasgow Science Centre) involved in science communication with schools and the general public. Our experience (e.g. at outreach activities such as the Glasgow Science Festival) is that the public can engage with such questions as 'can plants tell the time?' and that this can lead to discussion of why it is important to understand the timing mechanism.

How will they benefit?
Our work will be brought to the attention of industry and government through our interactions with crop geneticists and breeders at the James Hutton Institute. For example, JHI scientists are engaged in high throughput mapping of QTLs for traits in potato and barley. These scientists have interactions with the barley and potato breeding and processing communities in the UK, Europe and world-wide. Two key areas of interest are maturity determinants in barley and dormancy in potato both of which are influenced by the circadian clock. These areas of research are supported by the Scottish Government and policy groups within the government. We already have interactive research collaborations and have access to expertise, genetic resources and genomic information to catalyse translation of our basic research into the crop arena, so the long-term benefit will be in the broad area of food security.

We will disseminate the outcomes of our research at national and international plant, chronobiology and RNA meetings, and also present our work and publicise our research achievements e.g. to industry representatives who visit JHI or GU. Both universities seek to engage postdocs and PhDs to take part in the public engagement and impact agenda, for example by running Generic Skills programmes which also covers publicity activities. They also have Corporate Communications offices that regularly publicise research and promote engagement with the local media.

Publications

10 25 50
 
Description To analyse expression and alternative splicing in plants transferred from 20C to 4C we had performed a time-course of plants moved to the cold. samples were collected and RNA-sequencing performed. This dataset is probably one of the best datasets world-wide due to the high resolution of the time-course (every 3 hours for 26 time-points) and the depth of sequencing (ca. 180 million paired end reads per time-point). We have taken novel approaches to the analysis of this dataset which has required us to develop new approaches and methods.
We have developed a new Reference Transcript Dataset to allow us to analyse gene expression from a multiple time-point ultra-deep RNA-seq experiment. The RTD with new rapid analysis programmes represents an advance in gene expression analysis in Arabidopsis. We have developed a programme which allows the identification of isoform switches in the time-series data.
The key findings to date are that alternative splicing makes a significant contribution to re-programming of the transcriptome in response to cold stress; we have identified hundreds of novel cold response genes that are regulated by alternative splicing; mutants in some of these genes have been implicated in the alternative splicing of clock genes and some are required for freezing tolerance.
We have started to identify splicing factors that regulate alternative splicing of circadian clock genes in response to low temperatures which is a major objective of the grant.
Exploitation Route We have demonstrated the principle of analysing RNA-seq in a new way and have developed the RTD to allow this. Others in the Arabidopsis community in general can analyse or re-analyse their data. The principle can be applied to other plant species including crop plants and colleagues working on barley have begun to use this system.

We have interacted with colleagues working on barley and potato to generate RTDs for expression analysis. We have an EASTBIO PhD student who is developing an initial pipeline to construct an RTD for potato and this is progressing well. A publication on barley RTD will be submitted soon. Although occurring after this grant has finished, the research conducted under this grant was instrumental in devising strategies and methods to apply what we learned in Arabidopsis to crop species.
Sectors Agriculture, Food and Drink,Environment

 
Description BBSRC Response mode
Amount £1,000,000 (GBP)
Funding ID BB/P009751/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 04/2017 
End 03/2020
 
Description BBSRC Tools and Resources Development Fund
Amount £113,000 (GBP)
Funding ID BB/N022807/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 10/2016 
End 09/2017
 
Title 3D RNA-seq 
Description 3D RNA-seq is a program for accurate measurement of differential gene and transcript expression, differential alternative splicing, differential transcript usage and isoform switches from RNA-seq data. It is designed for use by biologists to overcome issues of access to bioinformaticians and to improve the accuracy of analysis over other available programs. The program already has had excellent uptake and excellent reviews (on social media). A paper has recently been submitted for publication. 
Type Of Material Technology assay or reagent 
Year Produced 2019 
Provided To Others? Yes  
Impact 3D has the potential to revolutionise RNA-seq analysis. Impact is measured by the growing number of users and requests from scientists for information and help in running. 
URL https://github.com/wyguo/ThreeDRNAseq
 
Title Arabidopsis Reference Transcript Dataset 
Description The Reference transcript dataset is a new set of transcripts for Arabidopsis which increases the diversity and number of known transcripts substantially. It can be used to analyse RNA-seq data rapidly and accurately to quantify expression levels of genes or individual transcripts. 
Type Of Material Technology assay or reagent 
Year Produced 2015 
Provided To Others? Yes  
Impact Other groups have begun to use it but it must be used in association with new analysis programmes and this is taking time to convert people. 
 
Title Arabidopsis thaliana Reference Transcript Dataset 2 
Description The tool is for analysis of RNA-seq data to quantify abundance of transcripts. The most accurate current programmes are Salmon and kallisto which require a reference transcriptome. We generated an earlier version which was an improvement on the available transcriptomes (TAIR10, Araport) but we realised that this could be significantly improved by assembling the extensive RNA-seq data which we and our collaborators in Vienna had generated. AtRTD2 was generated with a new pipeline of assembly and included many quality control filters to optimise the reference transcriptome. Its use with salmon on our data was validated by high resolution RT-PCR. AtRTD2 was pre-published in bioRxiv in May 2016. The paper is now published in Nucleic Acids Research. 
Type Of Material Technology assay or reagent 
Year Produced 2016 
Provided To Others? Yes  
Impact The major impact is in analysing our own dataset which is unique. The AtRTD2 was first made available to plant researchers in a pre-print in bioRxiv. A number of other groups are using AtRTD2. We have contacted Araport to get our assembly onto this database which has world-wide use. 
 
Title High resolution RT-PCR for alternative splicing 
Description The method allows us to examine alternative splicing in plant genes; it gives qualitative (discovery of new events) and quantitative data; it has been established for Arabidopsis but we have translated this to barley; it is a system to validate RNA-seq analysis. 
Type Of Material Technology assay or reagent 
Year Produced 2006 
Provided To Others? Yes  
Impact The method has been used by collaborators from around the world and has been established elsewhere by at least three other labs; it has led to many high quality pubications; 
 
Title TSIS - Time-Series Isoform Switch 
Description Gene expression is regulated by transcription and alternative splicing. In response to internal and external cues changes in AS occur. A particular type of change is an isoform switch where two AS transcripts from the same gene (isoform) change their relative abundance (switch). There are 2-3 programmes which can identify isoform switches and determine their significance. These function only on the basis of pairwise comparisons (e.g. wild-type vs treatment). We have an extensive time-course data set. We have developed Time-Series Isoform switch (TSIS) - a computational method to identify and evaluate isoform switches in time series data. Published in Bioinformatics and available on line. 
Type Of Material Technology assay or reagent 
Year Produced 2017 
Provided To Others? No  
Impact As the only available tool to analyse isoform switches in time-series data, we expect this to be used by researchers across the board who are using RNA-seq in time-course data. 
 
Title 3D RNA-seq - a program for biologists for easy and effective RNA-seq analysis 
Description RNA-seq analysis usually relies on bioinformaticians to find the time and will to analyse data generated by bench scientists. The analyses are not always optimum if the bioinformaticians do not fully understand what the biologists want. The time taken for getting such analyses done is a major source of frustration for biologists. We developed methods to analyse RNA-seq accurately and effectively. To meet the needs of biologists, we have developed an easy-to-use RNA-seq analysis program which can be used by biologists with minimal bioinformatics expertise or by bioinformaticians with limited experience in analysing gene expression. The program does the analysis and generates automatically the tables of differentially expressed, genes and transcripts, differentially alternatively spliced genes and differential transcript usage. The prototype was launched at the Cyverse Workshop at the Earlham Institute in December 2018. The updated version will be released in March/April 2019. 
Type Of Material Data analysis technique 
Year Produced 2019 
Provided To Others? No  
Impact The program will be made widely available soon. We expect it to have major impact. 
 
Title Arabidopsis thaliane Reference Transcript Dataset 2 (AtRTD2) 
Description The AtRTD2 is a very significant update over our previously released AtRTD. It has been assembled from over 9 billion paired end sequencing reads and many novel filters are used to ensure only high confidence transcripts are present It allows RNA-seq data from Arabidopsis to be analysed using programs such as salmon and kallisto. 
Type Of Material Database/Collection of data 
Year Produced 2016 
Provided To Others? Yes  
Impact The new AtRTD2 was released as a pre-print in bioRxiv in may 2016 so others could use it. It was published in march 2017 in Nucleic Acids Research. Papers from other groups are beginning to appear which have used this RTD. 
 
Title Arabidpsos Reference Transcript Dataset 
Description The RTD is a new collection of transcripts for Arabidopsis. It increases the number and diversity of transcripts for Arabidopsis substantially. It can be used with new rapid programmes to analyse or re-analyse RNA-seq data. 
Type Of Material Database/Collection of data 
Year Produced 2015 
Provided To Others? Yes  
Impact The RTD was released in August 2015 - it is being used by some groups but will get greater usage when we publish the main paper from the research which demonstrates its utility. 
 
Title New program to detect significant isoform switches in time-series data -Time-Series Isoform Switch (TSIS) 
Description One aspect of alternative splicing (AS) is isoform switching where the relative abundance of different isoforms of the same genes switches under different conditions. For example, isoform switches are used in cancer diagnostics. There are three programs to identify isoform switches in pairwise sample comparisons. TSIS is the only program which can identify significantly different isoform switches in time-series RNA-seq data. 
Type Of Material Data analysis technique 
Year Produced 2017 
Provided To Others? Yes  
Impact To date there are a small number of citations. However, it's application is clearly demonstrated in the Calixto et al (2018) paper and we expect more uptake. 
 
Description Alternative splicing and temperature control of the circadian clock 
Organisation University of Glasgow
Department Institute of Molecular Cell and Systems Biology
Country United Kingdom 
Sector Academic/University 
PI Contribution This is a joint grant with Prof Hugh Nimmo of Glasgow University and brings into collaboration our complementary skills in the circadian clock and in alternative splicing. We have specific goals and goals which we approach together as part of the strategy for delivery of the objectives. This was followed up by a further joint grant between the partners which has led directly to a number of publications and development of tools and methods for RNA-seq time-course analysis.
Collaborator Contribution We have analysed alternative splicing in clock and clock-associated genes; analysed the RNA-seq data and are looking at selected mutants for a clock AS phenotype. The RNA-seq time-course took a great deal of time to work out how best to do the analysis. This has proved extremely valuable and has made us one of the leading groups in such analysis. Additional outputs are tools and programs for specific types of analysis such as the new Arabidopsis transcriptome (AtRTD2), the R program Time-course Isoform Switch (TSIS)and £D RNA-seq - a program for biologists to analyse their own data. These advances have all emanated from the original grant and outputs in terms of the RNA-seq time-course dataset.
Impact To date, a number of highly cited publications have been produced along with tools and methods; we have also delivered seminars nationally and internationally.
Start Year 2013
 
Description Regulation of alternative splicing in plants 
Organisation Adam Mickiewicz University in Poznan
Country Poland 
Sector Academic/University 
PI Contribution Andrea Barta (Vienna), Artur Jarmolowski (Poznan) and I were the only plant groups to be invited to join the European Alternative Splicing Network of Excellence. We established a joint project to develop a high resolution RT-PCR system to analyse alternative splicing in plant genes which had been established initially in my lab. This system was used by us and opened up to other scientists around the world to use in collaborative research. Andrea Barta and Maria Kalyna (currently at the Agricultural University of Vienna continue to collaborate actively.
Collaborator Contribution Different mutants and plant material grown under stress conditions were supplied for analysis; in many cases PhD students or postdocs came to Dundee to run samples on the high resolution Rt-PCR system
Impact Publications; Conference proceedings; organisation of international meetings
Start Year 2006
 
Description Regulation of alternative splicing in plants 
Organisation Bielefeld University
Country Germany 
Sector Academic/University 
PI Contribution Andrea Barta (Vienna), Artur Jarmolowski (Poznan) and I were the only plant groups to be invited to join the European Alternative Splicing Network of Excellence. We established a joint project to develop a high resolution RT-PCR system to analyse alternative splicing in plant genes which had been established initially in my lab. This system was used by us and opened up to other scientists around the world to use in collaborative research. Andrea Barta and Maria Kalyna (currently at the Agricultural University of Vienna continue to collaborate actively.
Collaborator Contribution Different mutants and plant material grown under stress conditions were supplied for analysis; in many cases PhD students or postdocs came to Dundee to run samples on the high resolution Rt-PCR system
Impact Publications; Conference proceedings; organisation of international meetings
Start Year 2006
 
Description Regulation of alternative splicing in plants 
Organisation Leloir Institute
Country Argentina 
Sector Charity/Non Profit 
PI Contribution Andrea Barta (Vienna), Artur Jarmolowski (Poznan) and I were the only plant groups to be invited to join the European Alternative Splicing Network of Excellence. We established a joint project to develop a high resolution RT-PCR system to analyse alternative splicing in plant genes which had been established initially in my lab. This system was used by us and opened up to other scientists around the world to use in collaborative research. Andrea Barta and Maria Kalyna (currently at the Agricultural University of Vienna continue to collaborate actively.
Collaborator Contribution Different mutants and plant material grown under stress conditions were supplied for analysis; in many cases PhD students or postdocs came to Dundee to run samples on the high resolution Rt-PCR system
Impact Publications; Conference proceedings; organisation of international meetings
Start Year 2006
 
Description Regulation of alternative splicing in plants 
Organisation Medical University of Vienna
Country Austria 
Sector Academic/University 
PI Contribution Andrea Barta (Vienna), Artur Jarmolowski (Poznan) and I were the only plant groups to be invited to join the European Alternative Splicing Network of Excellence. We established a joint project to develop a high resolution RT-PCR system to analyse alternative splicing in plant genes which had been established initially in my lab. This system was used by us and opened up to other scientists around the world to use in collaborative research. Andrea Barta and Maria Kalyna (currently at the Agricultural University of Vienna continue to collaborate actively.
Collaborator Contribution Different mutants and plant material grown under stress conditions were supplied for analysis; in many cases PhD students or postdocs came to Dundee to run samples on the high resolution Rt-PCR system
Impact Publications; Conference proceedings; organisation of international meetings
Start Year 2006
 
Description Regulation of alternative splicing in plants 
Organisation National Center for Scientific Research (Centre National de la Recherche Scientifique CNRS)
Department Institute of Plant Sciences (ISV)
Country France 
Sector Academic/University 
PI Contribution Andrea Barta (Vienna), Artur Jarmolowski (Poznan) and I were the only plant groups to be invited to join the European Alternative Splicing Network of Excellence. We established a joint project to develop a high resolution RT-PCR system to analyse alternative splicing in plant genes which had been established initially in my lab. This system was used by us and opened up to other scientists around the world to use in collaborative research. Andrea Barta and Maria Kalyna (currently at the Agricultural University of Vienna continue to collaborate actively.
Collaborator Contribution Different mutants and plant material grown under stress conditions were supplied for analysis; in many cases PhD students or postdocs came to Dundee to run samples on the high resolution Rt-PCR system
Impact Publications; Conference proceedings; organisation of international meetings
Start Year 2006
 
Description Regulation of alternative splicing in plants 
Organisation University of Buenos Aires
Country Argentina 
Sector Academic/University 
PI Contribution Andrea Barta (Vienna), Artur Jarmolowski (Poznan) and I were the only plant groups to be invited to join the European Alternative Splicing Network of Excellence. We established a joint project to develop a high resolution RT-PCR system to analyse alternative splicing in plant genes which had been established initially in my lab. This system was used by us and opened up to other scientists around the world to use in collaborative research. Andrea Barta and Maria Kalyna (currently at the Agricultural University of Vienna continue to collaborate actively.
Collaborator Contribution Different mutants and plant material grown under stress conditions were supplied for analysis; in many cases PhD students or postdocs came to Dundee to run samples on the high resolution Rt-PCR system
Impact Publications; Conference proceedings; organisation of international meetings
Start Year 2006
 
Description Regulation of alternative splicing in plants 
Organisation University of California, Davis
Country United States 
Sector Academic/University 
PI Contribution Andrea Barta (Vienna), Artur Jarmolowski (Poznan) and I were the only plant groups to be invited to join the European Alternative Splicing Network of Excellence. We established a joint project to develop a high resolution RT-PCR system to analyse alternative splicing in plant genes which had been established initially in my lab. This system was used by us and opened up to other scientists around the world to use in collaborative research. Andrea Barta and Maria Kalyna (currently at the Agricultural University of Vienna continue to collaborate actively.
Collaborator Contribution Different mutants and plant material grown under stress conditions were supplied for analysis; in many cases PhD students or postdocs came to Dundee to run samples on the high resolution Rt-PCR system
Impact Publications; Conference proceedings; organisation of international meetings
Start Year 2006
 
Description Regulation of alternative splicing in plants 
Organisation University of Glasgow
Country United Kingdom 
Sector Academic/University 
PI Contribution Andrea Barta (Vienna), Artur Jarmolowski (Poznan) and I were the only plant groups to be invited to join the European Alternative Splicing Network of Excellence. We established a joint project to develop a high resolution RT-PCR system to analyse alternative splicing in plant genes which had been established initially in my lab. This system was used by us and opened up to other scientists around the world to use in collaborative research. Andrea Barta and Maria Kalyna (currently at the Agricultural University of Vienna continue to collaborate actively.
Collaborator Contribution Different mutants and plant material grown under stress conditions were supplied for analysis; in many cases PhD students or postdocs came to Dundee to run samples on the high resolution Rt-PCR system
Impact Publications; Conference proceedings; organisation of international meetings
Start Year 2006
 
Description 3D RNA-seq training workshops at University of Leeds (October) and Nottingham (November) - presented by Runxuan Zhang, Wenbin Guo, JC Entizne. 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Study participants or study members
Results and Impact We have developed 3D RNA-seq in oreder to analyse the RNA-seq data generated in time-courses of Arabidopsis plants exposed to cold. 3D RNA-seq is an analysis program for RNA-seq data analysis designed for use by biologists with minimal bioinformatics experience. The program is an easy-to-use tool that provides accurate differential gene and transcript expression and differential alternative splicing. It can be used for RNA-seq data from eukaryotes and has been successfully used with plants (Arabidopsis, potato, barley etc) and animals (human, mouse etc). It won the University of Dundee School of Life Sciences Best Innovation award in 2019. 3D RNA-seq was launched in May 2019 with publication in bioRxiv and has had great success with nearly 2,500 users. To help to have uptake in the UK, we have run training courses in the University of Leeds (supported by GARNet and the University of Nottingham. In addtion, we have trained individuals from human and medical sciences.
Year(s) Of Engagement Activity 2019
 
Description Alternative splicing and the cold transcriptome of Arabidopsis (John Brown/4th International Post-EURASNET meeting - Poznan, Poland/September 2016) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact The meeting is a follow-on from the very successful European Alternative Splicing Network of Excellence (EURASNET). The post-EURASNET meeting series brings together research groups working on alternative splicing in model species, plants, animals and humans. The most up-to-date research on AS is presented from molecular structures to gene therapy. The plant groups continue to be active in this meeting and to collaborate extensively with one another. The active collaboration among plant groups working on AS in the EU is a significant outcome.
Year(s) Of Engagement Activity 2016
 
Description Alternative splicing in the Arabidopsis thaliana circadian clock: an RNA-seq approach (Cristiane Calixto/FAPESP workshop, Brazil/2015) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Presentation by Dr. Cristiane Calixto at the FAPESP-British Council Workshop on Environmental and metabolic control of plant growth and development. 2015, Campinas, Brazil.
Year(s) Of Engagement Activity 2015
 
Description Alternative splicing in the Arabidopsis thaliana circadian clock: an RNA-seq approach (Cristiane Calixto/PRGEP meeting - Paris/2015) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk presented by Dr Cristiane Calixto at the Post-transcriptional Regulation of Gene Expression in Plants meeting. 2015, Paris, France.
Year(s) Of Engagement Activity 2015
 
Description Alternative splicing in the Arabidopsis thaliana circadian clock: an RNA-seq approach (Cristiane Calixto/RNA UK/2015) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Talk by Dr Cristiane Calixto at the RNA UK meeting, Windermere, UK.
Year(s) Of Engagement Activity 2016
 
Description Alternatively spliced genes as novel cold responsive genes in Arabidopsis (Cristiane Calixto/ASPB Hawaii/2017) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk in RNA Biology session of ASPB Plant Biology meeting Hawaii, USA 2017
Year(s) Of Engagement Activity 2017
 
Description Alternatively spliced genes as novel cold responsive genes in Arabidopsis (Cristiane Calixto/UK RNA Splicing/Lake district/January 2017) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact The meeting brings together research groups mainly from the UK working on RNA splicing in different organisms (yeast, plants, animals, human). Topics cover molecular mechanisms up to human disease and gene therapy. It is an opportunity to present our work to a very knowledgeable audience.
Year(s) Of Engagement Activity 2017
 
Description Alternatively spliced genes as novel cold responsive genes in Arabidopsis (J Brown/Amsterdam/2017) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited talk at Plant Genomics and Gene Editing Congress, 16-17 March 2017, Amsterdam, The Netherlands
Year(s) Of Engagement Activity 2017
 
Description Blog on clock research 
Form Of Engagement Activity Engagement focused website, blog or social media channel
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact The blog was to present some ideas from the collaborative groups from Glasgow and Dundee university on the clock, rhythms and alternative splicing
Year(s) Of Engagement Activity 2016
URL https://abouttimeresearch.com/
 
Description Family Fun Day and Fascination of Plants Day - Dundee Botanic Gardens 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Activities designed to provide information/interest in DNA, genetics, diversity

Excitement from children and interest from parents about DNA when children extracted DNA from raspberries.
Year(s) Of Engagement Activity 2011,2012,2013,2014,2015
URL http://www.lifesci.dundee.ac.uk/research/ps/engagement-plant-sciences
 
Description Garnet Newsletter - New Arabidopsis transcriptome AtRTD2 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Article about the new RTD2 transcriptome that we developed to analyse the RNA-seq data from the cold/clock experiment. Objective was to raise awareness among Arabidopsis scientists to use RTD2 instead of other references.
Year(s) Of Engagement Activity 2016
URL http://www.garnetcommunity.org.uk/newsletters
 
Description Interplay between gene expression and alternative splicing determines cold acclimation in Arabidopsis (John Brown/Cambridge University/November 2017) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Invited seminar - Plant Sciences, University of Cambridge
Year(s) Of Engagement Activity 2017
 
Description Invited talk at 60th Brazilian Genetics Congress, Guaraja, Brazil 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Invited seminar at the Congress to present results about alternative splicing in the circadian clock
Year(s) Of Engagement Activity 2014
 
Description Invited talk at Danforth Centre in St. Louis, USA 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Seminar about RNA-seq analysis and alternative splicing in the circadian clock to academics including students, PIs etc.
Year(s) Of Engagement Activity 2015
 
Description Open Doors Day 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Doors Open Day in the School of Life sciences has tours and exhibits and hands-on demonstrations for the general public. We attend to talk about our and other Plant Science research.
Year(s) Of Engagement Activity 2012,2013,2014,2015
 
Description Plant Power Day - Botanics, Dundee University 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Open day at the Botanics in Dundee to promote plant science to the general public. Number of exhibits presented throughout the day - opportunity to discuss issues with plant science e.g. GM with the public.
Year(s) Of Engagement Activity 2016
 
Description Presentaion at Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben, Germany - Rapid and dynamic alternative splicing impacts the cold response transcriptome in Arabidopsis - given by Prof John W S Brown 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited seminar at IPK-Gatersleben; multiple discussions with different group leaders and interested parties
Year(s) Of Engagement Activity 2018
 
Description Presentaion at Molecular Cell Physiology Department, University of Bielefeld, Bielefeld, Germany - Rapid and dynamic alternative splicing impacts the cold response transcriptome in Arabidopsis - given by Prof. John W S Brown 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited seminar - discussions with PhD students and research groups interested in our advanced approaches.
Year(s) Of Engagement Activity 2018
 
Description Presentation at 6th UK RNA Splicing Workshop - Rapid cold-induced alternative splicing in Arabidopsis involves a complex network of regulators - given by Prof John W S Brown 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Presentation covered our research on cold-induced changes in expression and alternative splicing. Also presented a new method/tool of RNA-seq analysis designed for biologists. massive interest in this.
Year(s) Of Engagement Activity 2018
 
Description Presentation at GARNet2018:a plant science showcase at University of York - in Large Scale Biology section - "Genome-wide alternative splicing" - given by Dr Cristane Calixto 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Invited to speak about oue genome-wide methods for analysis of RNA-seq for gene expression and alternative splicing
Year(s) Of Engagement Activity 2018
 
Description Presentation at SEB Annual Meeting, Florence - Rapid cold-induced alternative splicing in Arabidopsis involves a complex network of regulators - given by Dr Nikoleta Tzioutziou 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Presentation in section: Plant Temperature responses: Shaping Development and Enhancing Survival?

Presented out novel RNA-seq analysis methods for time-course analysis (paper published in Plant Cell - Calixto et al 2018.
Year(s) Of Engagement Activity 2018
 
Description RNA-squencing meeting (University of Dundee and James Hutton Institute) 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Professional Practitioners
Results and Impact We organised an "RNA-sequencing afternoon" workshop for people involved in RNA-seq analyses at the University of Dundee and the James Hutton Institute. the purpose was to raise awareness of new approaches in this very fast moving field.
Year(s) Of Engagement Activity 2017
 
Description Rapid and dynamic alternative splicing impacts the Arabidopsis cold response (Cristiane Calixto/IGC Symposium - Lisbon/2017) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited talk at the IGC Symposium 2017 - Plant RNA Biology. Lisbon, 27-28 September 2017 (talk given by Cristiane Calixto)
Year(s) Of Engagement Activity 2017
 
Description Rapid and dynamic alternative splicing impacts the cold response transcriptome in Arabidopsis (John Brown/Humboldt University, Berlin/January 2018) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited seminar at Humboldt University, Berlin, Germany
Year(s) Of Engagement Activity 2018
 
Description Rapid and dynamic alternative splicing impacts the cold response transcriptome in Arabidopsis (John Brown/Max Planck Institute, Golm, Germany/January 2018) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited seminar at MPI Golm, Germany
Year(s) Of Engagement Activity 2018
 
Description Re-programming of the cold transcriptome in Arabidopsis (John Brown/Invited Seminar - Durham University/January 2017) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact I gave a seminar at Durham University to update them on our methods of RNA-seq analysis and new pipelines for such analysis. There is a great deal of interest from 203 groups about our methods but also on cold responses in plants. This offers an opportunity for advertising our new Reference Transcript Dataset (AtRTD2) and other methods that we have developed to plant scientists likely to use them. This will bring new collaborations.
Year(s) Of Engagement Activity 2017
 
Description Regulation of Alternative Splicing of Arabidopsis Clock Genes in Cold Temperature Using Deep RNA-seq (Cristiane Calixto/Clock Networks in Plants and Algae - Edinburgh, UK/2016). 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Talk given by Dr Cristiane Calixto at the Clock Networks in Plants and Algae Meeting in Edinburgh, UK. Presented our new work on regulation of clock gene expression by tem[erature.
Year(s) Of Engagement Activity 2016
 
Description Seminar "Re-programming of the cold transcriptome in Arabidopsis" Sainsbury lab, Cambridge (November 2016) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact I gave a seminar at the Sainsbury lab in Cambridge to update them on our methods of RNA-seq analysis and new pipelines for such analysis. There is a great deal of interest. This offers an opportunity for advertising our new Reference Transcript Dataset (AtRTD2) and other methods that we have developed to plant scientists likely to use them.
Year(s) Of Engagement Activity 2016
 
Description Seminar - Alternative splicing and the cold transcriptome in Arabidopsis - Earlham Institute, Norwich (November 2016) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact I gave a seminar at the Earlham Institute in Norwich to update them on our methods of RNA-seq analysis and new pipelines for such analysis. This offers an opportunity for advertising our new Reference Transcript Dataset (AtRTD2) and other methods that we have developed to plant scientists likely to use them. The expertise in bioinformatics at Earlham makes EI an excellent place to discuss our topic. A major point is that EI need to engage with AS analysis.
Year(s) Of Engagement Activity 2016
 
Description Seminar at Central European Institute for Technology (CEITEC), Masaryk UniversityBrno, Czech Republic 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Seminar about our results in analysis of RNA-seq data and alternative splicing in the clock
Year(s) Of Engagement Activity 2015
 
Description Seminar atThe Sainsbury Lab, Cambridge University 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Postgraduate students
Results and Impact Seminar about methods of RNA-seq analysis and alternative splicing - to academics
Year(s) Of Engagement Activity 2016
 
Description The cold transcriptome of Arabidopsis using ultra-deep RNA-seq: regulation of AS of clock genes (Cristiane Calixto/PRGEP meeting - 2016, Austin, USA/2016) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Seminar by Dr Cristiane Calixto at meeting on Post-transcriptional Regulation of Gene Expression in Plants meeting. 2016, Austin, USA. This is a satellite meeting of the main ASPB meeting.
Year(s) Of Engagement Activity 2016
 
Description The cold transcriptome of Arabidopsis using ultra-deep RNA-seq: regulation of AS of clock genes (Poster/Cristiane calixto/ASPB USA 2016) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Poster presented by Dr Cristiane Calixto at the Plant Biology meeting, Austin, Texas, USA
Year(s) Of Engagement Activity 2016