Alternative splicing and its impact on molecular interaction networks during stem cell differentiation.
PTDC/BIA-GEN/116519/2010


Summary

Stem cells have been the focus of intense research in recent years. Their unique dual capacity to self-renew and to differentiate into other cell types represents an attractive feature, not only for potential applications in regenerative medicine, but also for the study of fundamental processes in embryology such as the development of complex multi-cellular organs. Moreover, strong indications exist that stem cells play critical roles in neurodegenerative diseases, diabetes and cancer. Despite the great progress achieved in stem cell biology in the last decade, however, many characteristics and features of the stem cells still await full clarification.

Of key importance for safe application of stem cells in medicine is the dissection and determination of the genetic programs that govern maintenance and differentiation of stem cells. These programs are executed by complex molecular networks, which combine internal and external cues to determine the fate of the cells. In this project, we aim to reconstruct and analysis the molecular networks defining stem cell identify and to model their dynamic changes during differentiation. A specific focus is set on the impact of alternative splicing on the structure and function of molecular networks that drive the drive the differentiation of stem cells towards heart muscle cells (cardiomyocytes). Alternative splicing enables the synthesis of multiple transcripts from a single gene and is a major mechanism for the expansion, diversification and regulation of gene function in higher organims. Although alternative splicing has been intensively studied, how it can modify the function of individual genes, its concerted impact on complex molecular networks remains largely to be explored. From the medical perspective, the study of differentiation of stem cells into cardiomyocytes is particularly interestingly, since it might give us new indications for innovative therapies for heart failure - the leading cause of death worldwide.

As a basis for our studies, we have considerably extended our UniHI database (NAR, 2014), a comprehensive resource for molecular interaction networks in human. To make the networks more specific to stem cell biology, we collected a large number of physical and regulatory molecular interactions for human and mouse, and integrated them in our newly established StemCellNet webserver (NAR, 2014). To assess the relevance of the individual components for the unique characteristic of stem cells i.e. the capacity for self-renewal and for generation of differentiated progeny or in short, their stemness, we have also curated and integrated numerous published stem cell signatures resulting in our StemChecker web-tool (NAR, 2015). Additionally, we have collaborated with the group led by Paul de Sousa (MRC Centre for Regenerative Medicine, University of Edinburgh, UK) to identify epigenetic signatures for human stem cells (PloS ONE, 2015). The established resources enable us to build comprehensive molecular networks for stem cells and to study their changes during differentiation. Currently, we are using RNA-Seq data to identify splicing events, which can potentially alter structure and function of relevant molecular networks. Identified events will subsequently be validated and further characterised in collaboration with other research groups. Importantly, they could give us key molecular switches and eventually potential novel drug targets for the control of the the behaviour and function of networks (Frontiers in Genetics, 2014).


Project Team



Established Databases & Software

StemCellNet

StemcCellNet is an interactive web server for network analysis and visualization in stem cell biology. It gives access to a large collection of curated physical and regulatory interactions identified in human and murine stem cells and features various easy-to-use tools for selection and prioritization of network components, as well as for integration of expression data. StemCellNet can indicate novel candidate genes by evaluating their connectivity patterns. It is the only current platform, which allows the screening of networks for stemness-associated genes and potential target candidates. With its comprehensive coverage of the human interactome, it is a powerful tool not only for stem cell researchers, but also for researchers working on degenerative diseases and on cancer to identify stemness signatures in molecular networks of interest.

StemChecker

StemChecker is a web-based tool that enables researchers to rapidly check whether a given list of genes can be linked to stemness. For this purpose, we curated numerous published stemness signatures derived by alternative approaches. StemChecker examines whether genes uploaded by the user are included in the curated set of stemness signatures and evaluates the statistical significance. The results are displayed in alternative formats, showing the potential association with stemness signatures of individual genes, as well as of the whole set of inputted genes. Additionally, StemChecker indicates whether genes are targeted by a set of transcription factors linked to pluripotency and stem cell maintenance.


HeartEXpress

HeartEXpress is a web-based platform for the analysis of integrated expression datasets associated with cardiomyogenesis. The current version comprises data from independent microarray experiments for stem cell differentiation, in vitro or in vivo reprogramming, in the context of cardiomyogenesis, and heart development.


Publications

Stephen Pells, Eirini Koutsouraki, Sofia Morfopoulou, Sara Valencia-Cadavid, Simon Tomlinson, Ravi Kalathur, Matthias Futschik and Paul De Sousa (2015) Novel human embryonic stem cell regulators identified by conserved and distinct CpG island methylation state, PLoS ONE 10(7): e0131102. (pdf + html)

José P. Pinto, Ravi K. Kalathur, Daniel V. Oliveira, Tânia Barata, Rui S. R. Machado, Susana Machado, Ivette Pacheco-Leyva, Isabel Duarte and Matthias E. Futschik (2015) StemChecker: a web-based tool to discover and explore stemness signatures in gene sets, Nucleic Acids Research, 43 (W1): W72-W77 (pdf + html)

José P. Pinto, Ravi K.R. Kalathur, Rui S. R. Machado, Joana M. Xavier, José Braganaça, and Matthias E. Futschik (2014) StemCellNet: an interactive platform for network-oriented investigations in stem cell biology. Nucleic Acids Research 42:W154-W160. (pdf+html)

José P. Pinto, Rui Machado, Joana Xavier and Matthias E. Futschik (2014) Targeting molecular networks for drug research. Frontiers in Genetics 5:160 (pdf + html )

Ravi K. Kalathur, José P. Pinto, Miguel A. Hernández-Prieto, Rui S.R. Machado, Dulce Almeida, Gautam Chaurasia and Matthias Futschik (2014) UniHI 7: an enhanced database for retrieval and interactive analysis of human molecular interaction networks, Nucleic Acids Research, Database issue, 42 (D1): D408-D414 (html, pdf)



Presentations

Isabel Duarte, D.Oliveira, T. Justo, J. Braganša and M. Futschik (2015) Predicting Cardiac Novel Splice Variantes from RNAseq data, Bioinformatics for the Life Sciences, Universidade do Minho, Braga, Portugal

DV Oliveira, I Duarte, RS Machado, I Pacheco-Leyva, T Justo, JP Pinto, S Machado, JA Belo, J Braganša & ME Futschik (2015) From Stem Cells 2 Heart: Finding novel candidate isoforms and genes for cardiomyogenesis. 9th International Meeting of the Portuguese society for Stem cells and Cell therapies. Oeiras. Portugal.

RS Machado, JP Pinto, DV Oliveira, I. Duarte, S Machado, J Braganša & M Futschik (2015) A meta-analysis study of stem cell differentiation in the context of cardiogenesis.9th International Meeting of the Portuguese society for Stem cells and Cell therapies. Oeiras. Portugal.

S Machado, R Kalathur, DV Oliveira, T Barata, R Machado, I Pacheco-Leyva, I Duarte, M Futschik, J Pinto (2015) StemChecker: a web-based tool to discover and explore stemness signatures in gene sets.9th International Meeting of the Portuguese society for Stem cells and Cell therapies. Oeiras. Portugal.

JP Pinto, R Kalathur, RS Machado, J Xavier, J Braganša & ME Futschik (2015) StemCellNet: an interactive platform for network -oriented investigations in stem cell biology. 9th International Meeting of the Portuguese society for Stem cells and Cell therapies. Oeiras. Portugal.

Isabel Duarte, D.Oliveira, T. Justo, J. Braganša, J Belo and M. Futschik (2015) From Stem Cells 2 Heart: Estudo do Impacto do Splice Alternativo durante o Desenvolvimento Cardiaco, Molecular Genetics Anual Seminar Series, Universidade do Algarve, Faro, Portugal

J. Pinto, R. Kalathur, R. Machado , J. Xavier, J. Bragança , and M.E Futschik (2014) StemCellNet: an interactive platform for network-oriented investigations in stem cell biology, Heart without Boarders, International Conference, Porto, Portugal (abstract)

I. Duarte, T. Justo, J. Pinto, D.Oliveira, J. Braganaça, J. Belo and M. Futschik (2014) From Stem Cells 2 heart: Elucidating the role of alternative splicing in cardiomyogenesis, Heart without Boarders, International Conference, Porto, Portugal (abstract)

D Oliveira, R. Machado, I. Pacheco-Leyva, I. Duarte, T. Justo , J. Pinto, J. Belo, J. Braganaça and M. Futschik (2014) From Stem Cells 2 heart: Finding novel candidate genes for cardiomyogenesis, Heart without Boarders, International Conference, Porto, Portugal (abstract)

Rui S.R. Machado, José Pedro B. G. P. Pinto, José Braganaça and Matthias E. Futschik (2013) What a heart makes or breaks: A meta-analysis of gene expression profiles , 8th International Meeting of the Portuguese Society for Stem Cells and Cell Therapies, Faro, Portugal

José Pedro B. G. P. Pinto, Rui S.R. Machado<, Ravi K.R. Kalathur, and Matthias E. Futschik (2013) StemCellNet: A web-based Platform for Assembly and Analysis of Molecular Interaction Networks in Stem Cells, 8th International Meeting of the Portuguese Society for Stem Cells and Cell Therapies, Faro, Portugal