Cedric Notredame, a group leader at the CRG, tells us in his “Slow bioinformatics blog” his personal and interesting story behind the development of T-coffee, a method for multiple sequence alignment which he developed during his PhD and which is currently widely used.
“For those who have no clue what T-Coffee does, it is a multiple sequence aligner. It means that it takes a bunch of biological sequences – typically proteins – that have evolved from a common ancestor by accumulating mutations, insertions and deletions…”
If you want to know the real story behind the T-coffee success, read Notredame’s blog here!
“You went to high school and you learned genetics. You heard about a certain Gregor Mendel who crossed peas and came up with the idea that there is a dominant and a recessive allele. You did not particularly like the guy because there would always be a question about peas with recessive and dominant alleles at the exam. But you grew up, became wiser and just as you started to like him, you heard from someone that he faked his data….”
Did he or didn’t he?
You can read Guillaume Filion’s latest blog entry about the father of Mendelian genetics and statistics – but you will have to choose by yourself!
Different targeted strategies have recently emerged in the field of proteomics that enable the detection and quantification of a predetermined subset of proteins with a high degree of sensitivity and reproducibility across many samples. Major advances have been achieved in the targeted proteomics workflow, including advances in instrumentation, the generation of thousands of publicly available targeted assays, and the development of multiple computational tools for convenient data analysis.
This field is known as targeted proteomics and although it has successfully been applied in several research projects of molecular biology, systems biology and translational medicine, there is still a strong gap between theory and real application.
To address this mismatch and boost the applications of targeted proteomics, the PRBB held a new edition of the 6-day long EMBO Practical Course “Targeted proteomics: Experimental design and data analysis” co-organised by Eduard Sabidó, head of the CRG/UPF Proteomics Unit and Ruedi Aebersold from the ETH in Zurich.
Twenty-five participants coming from 20 countries from all continents attended the course, which offered a combination of keynotes, practical demonstrations and tutorials to provide the participants with the required knowledge and skills to design and analyse their own targeted proteomic experiments using the most advanced and state-of-the-art methods.
Each day started with a keynote open to all PRBB residents by different renowned proteomics researchers that reviewed the latest achievements in the field of targeted proteomics and introduced the “topic of the day”. The students also attended to several practical session to master the complete workflow associated to targeted proteomics, thus filling the gap between theory and the actual implementation of targeted proteomics experiments. During the practical sessions the students generated, refined and optimized targeted proteomics methods for a set of selected proteins of interest, and automated manual data analysis by reviewing concepts such as peak picking, quality assessment, and statistics for accurate protein quantitation. The program was complemented with poster sessions and several social events to foster informal scientific discussions and to adapt the technology to each participant’s particular interests.
The participants, ranging from PhD students to postdocs and senior researchers, gave a very positive feedback on the course, with an overall score of 4’75 on a scale from 0 to 5 and very encouraging comments:
“Very motivating meeting with very interesting researchers from very different fields but same interest in proteomics, and high quality seminars and tutorials that made me want to continue research in proteomics”
“I feel confident to start my own experiments now”
“Informative and engaging with strong relevance to my research”
“The most useful course so far and also great to share knowledge and tips between us!”
“Excellent speakers, plenty usufull, practical informations, nice atmosphere it is shortest description of the course”
“A wonderful course instructed by cutting edge and innovative field leading scientists whom together have passed on the necessary tools to enable me to design and implement successful targeted proteomic workflows”
This course disseminated the know-how present at the CRG/UPF Proteomics Unit to a broader scientific community and strengthened the interdisciplinary exchange of knowledge and ideas. Thus, by transferring the expertise on experimental design and data analysis for targeted proteomics, the CRG/UPF Proteomics Unit aims to facilitate a wider and more routine application of targeted proteomics among non-proteomic laboratories worldwide.
For those who could not attend this year, the EMBO Practical Course “Targeted proteomics: Experimental design and data analysis” will take place again at the PRBB in November 2016. You will soon be able to access this website to register for the 2016 edition – do not miss this opportunity!
You can see the videos of the 2015 edition here.
Course instructors that participated in the 2015 edition
Abersold, Ruedi; ETH Zürich, Switzerland
Altelaar, Maarten; Utrecht University, The Netherlands
Borràs, Eva; CRG-UPF Proteomics Unit, Spain
Bensimon, Ariel; ETH Zürich, Switzerland
Chiva, Cristina; CRG-UPF Proteomics Unit, Spain
Guillet, Ludovic; ETH Zürich, Switzerland
Ludwig, Christina; ETH Zürich, Switzerland
MacCoss, Michael; University of Washington, USA
MacLean, Brendan; University of Washington, USA
Reiter, Lukas; Biognosys GmbH, Switzerland
Sabidó, Eduard; CRG-UPF Proteomics Unit, Spain
Vitek, Olga; Northeastern University, USA
A couple of months ago, the CRG-UPF Proteomics Unit at the PRBB – run by the Centre for Genomic Regulation (CRG) and the Pompeu Fabra University (UPF) – announced the installation of one of the most precise mass spectrometers in the world, marketed as Orbitrap Fusion Lumos, becoming the first place in Europe and the third in the world to have this tribrid instrument.
You can read more about this innovative mass spectrometer – which can achieve simultaneous analysis and quantification of more than 10,000 proteins from ten different samples in a single day! – in a post entitled “The CRG Becomes the First European Centre to Have One of the Most Precise Mass Spectrometers in the World” and published on September 29th in Biocores – a directory of the core facilities, technological platforms, and scientific services in Barcelona.
You can also read an article published in Ellipse – the monthly newspaper of the PRBB – in which Eduard Sabidó, head of the Proteomics Unit, tells us about how mass spectrometry-based proteomics is helping to solve complex problems of molecular biology and translational research (see page 6 of this months’ edition of the Ellipse).
Transcription is a process that depends on many elements such as transcription factors, DNA methylation and chromatin structure. Histones are essential for the proper regulation of transcription, and the posttranslational modifications on their tails have been related both to activation (H3K4me3, H3K9ac and H3K36me3) and silencing (H3K27me3 and H3K9me3) of gene expression.
Using the fruit fly as model organism, in our lab at the CRG and in collaboration with Montserrat Corominas’ lab in the Universitat de Barcelona, we have found a set of genes (defined as developmentally regulated genes by their restricted expression to a short period of time) that are expressed without the canonical histone marks associated to activation of transcription. These findings have been published in the October’s issue of Nature Genetics, the cover of which also refers to our work and to Dalí’s butterflies –designed by Luisa Lente and Hagen Tilgner.
Why these developmentally regulated genes show a different pattern of histone modifications is not known, but we speculate that the need of rapid activation–deactivation of gene expression during development may be easier to achieve in absence of the histone posttranslational modifications. In this context, transcription factors would play a more important role. This hypothesis is reinforced by the fact that the binding of transcription factors is different between developmentally regulated and stable genes (those expressed throughout development).
There are previous reports claiming that some genes are transcribed in the absence of the canonical histone marks for gene expression (Hödl and Basler –2012–, Chen and collaborators –2013– and Zhang and collaborators –2014–), but our main contribution to the field is that we have seen that the lack of chromatin marks is a general feature of genes that are expressed for a short period of time, being the expression of these genes likely to be regulated mainly by the action of transcription factors.
We are often asked how come nobody had described these patterns before. Actually, the answer is probably because our approach was different from the beginning. When we started the project our main goal was to analyze the chromatin marks and the splicing of tissue- and time-specific genes in the fruit fly. The settings used to define the tissue- and time-specific genes were, then, very astringent, being our developmentally regulated genes only expressed in one time point throughout development. However, to our surprise, these particular genes did not show the expected histone marks when they were expressed. It was then that we focused our attention in these particular genes.
This work was presented in the 11th Transcription and Chromatin Conference (2014) at the EMBL in Heidelberg – where they were already introduced as being controversial- and was afterwards highlighted in the EpiGenie webpage and in the Epigenetics journal, in both cases by Sascha H. C. Duttke.
As we knew that these results are controversial and somehow challenge the classical association of histone marks with transcription, we put a great deal of effort in generating and analyzing all available data to produce as solid results as possible to demonstrate that our observations do not arise from a detection artifact. Besides, we think that with this work we open a new line of research as the results, so far observed in fly and worm, need to be further validated in mammalian systems. In this sense, our results so far with the ENCODE mouse tissues and human cell lines point out that this lack of chromatin marks in regulated genes may be a general feature along metazoans.
Marc A. Marti-Renom is interested in three-dimensional structures. After eight years in the US dedicated to the world of proteins, the biophysicist returned to his native country, first Valencia and then Barcelona, to specialise in RNA and DNA folding. In 2006 he set up his own group, which today is divided between the CNAG, where there are ten people, and the CRG, where there are two. “We do the experimental part, the sample preparation, here in the CRG, and the sequencing and analysis happens in the CNAG”, he explains. For his research he requires a large sequencing and computing capacity, which he can get at the CNAG, the second-most important sequencing analysis centre in Europe. “We are fortunate to be in one of the best places in the world to do these studies,” he says proudly.
Proteins with clinical application
Proteins caught his attention while he was doing his PhD, and in 2004, when he was at the University of California (UCSF), he collaborated in the creation of the “Tropical Disease Initiative,” a drug-discovery initiative linking people from both academia and companies to try to reposition drugs in favour of neglected diseases such as malaria and tuberculosis. “The idea was to make it all open source so everything we found was published directly to the web and couldn’t be patented”, says Marti-Renom.
The Structural Genomics group was a major player in one of the first instances that genome sequencing was used at the clinical level. “There was a patient with tuberculosis and a high resistance to antibiotics. We sequenced samples from the patient and found out he was infected by two different strains, and one of them was mutated. When we made models of the protein structure resulting from this mutation we saw how it was affecting the function”, explains the scientist. According to Marti-Renom, in a few years not only will everyone have their genome sequenced, but it will happen several times. “When someone develops a disease like cancer we will sequence them again to see what has changed and why”, he predicts.
Beyond proteins: RNA and DNA
Proteins, the cell’s building blocks, are not the be-all and end-all of life. Since the 1960s we have known that RNA has essential functions other than converting the information in DNA into proteins. But of its three-dimensional structure very little is known, and in the end, the function occurs in 3D. For this reason the group is developing computational tools to incorporate experimental data and make structural predictions.
The most recent biological component to enter the ‘3D world’ was the genome. In this case, too, little is known about how it folds in space. The group of Marti-Renom, along with three other groups at the CRG (Miguel Beato, Guillaume Fillion and Thomas Graf) is carrying out the 4DGenome project, which has a budget of 12.2 million euros, in order to understand the structure of the genome and how it changes over time. “We know the genome sequence very well, thanks to molecular biology and the big genome projects. We also understand the chromosomal macrostructure, thanks to advances in microscopy; but we can’t see the middle ground, the step between the tangled skein and the well-defined chromosome”, says the head of the group. In 2006 they began using Chromosome Conformation Capture (3C) data to develop software that allows you to view the entire genome at high-resolution, a kind of ‘molecular microscope’. With this and other technologies, like Hi-C, and using computational algorithms they have been able to observe how different regions of the same chromosome tend to interact with each other. They have also seen that the 3D ‘photo’ of a moment when, for example, there is high gene expression may be very different to another where the expression is low. “Without this three-dimensional information it is much more difficult to characterise how the genome works”, concludes the researcher.
This half-experimental half-computational laboratory works on how the architecture of the genome affects gene expression. How does a cell know how to read its DNA? Why some genes are or are not activated depending on the genomic context where they are located? The Filion’s lab studies the mystery of position effect variegation. The French scientist explains why this is important in one minute.
Video produced by the Barcelona Biomedical Research Park (www.prbb.org).
Xavier Estivill and his Genomics and Disease research group at the CRG are trying to find the genetic causes of complex diseases using the latest genomic technologies. Focused on central nervous system diseases and on non-coding RNAs, he is also involved in international sequencing projects such as the International Cancer Genome Consortium (ICGC). Hear him explain his research in this short video!
Methadone maintenance treatment (MMT) is the most widely-used therapy in opioid dependence, but it is not effective in some patients, who relapse or drop out from treatment. Researchers at the IMIM and Hospital del Mar led by Marta Torrens, in collaboration with colleagues at the CRG, have found a possible explanation of why some people may not respond well to this treatment.
As the authors explain in their paper published this month in the journal European Neuropsychopharmacology, they carried out a genetic analysis on several patients, focusing on the gene ALDH5A1. This enzyme is involved in the catabolism of the neurotransmitter gamma-aminobutyric acid (GABA), the major inhibitory neurotransmitter in the mammalian central nervous system. ALDH5A1 comes in many forms, and the scientists found that subjects carrying the T variant allele had a higher risk to be nonresponders to methadone treatment. They hypothesized that this could be due to a reduction in the ALDH5A1 enzyme activity, which would increase endogenous GABA levels and therefore induce symptoms such as sedation and impaired psychomotor performance. These neuropsychological effects related with the reduction in enzyme activity could be responsible for a higher propensity to relapse in these genetically predisposed patients.
The findings could be helpful to predict which subjects with opioid dependence problems would probably not benefit from methadone maintenance treatment and could use other treatments instead, such as diamorphine.
Fonseca F, Gratacòs M, Escaramís G, De Cid R, Martín-Santos R, Farré M, Estivill X, Torrens M. ALDH5A1 variability in opioid dependent patients could influence response to methadone treatment. Eur Neuropsychopharmacol. 2013 Oct 18;
Where do we come from? Why are we the way we are? Why aren’t there any 6-legged mammals? In this short video, Yogi Jaeger, head of the Comparative Analysis of Developmental Systems lab at the Centre for Genomic Regulation (CRG) talks about his research into these and other questions.