The article is truly spectacular, both for the novelty of the discoveries and for their quality, complexity and relevance, as well as for the amount of original information it offers and for the new ways and means of study and analysis it makes available to researchers to advance in this field.

I know from colleagues in this area that Juan Valcárcel's laboratory has been working for years to complete this study, which due to its magnitude and complexity has surely cost a great effort and a considerable investment.

They really manage to show for the first time the mapping of the fundamental nucleus of the human spliceosome. Because of its degree of detail and depth, this map, together with the new molecular 'laws' and interactions that it has allowed them to discover, represents a real pillar to consolidate this new field of study that we can call spliceosomics, in which Valcárcel's laboratory is an internationally recognized pioneer.

Until now we knew in a more or less partial or fragmentary way how a limited (although increasing) number of the multiple components of the spliceosome (the splicing factors) influence the mechanisms of splicing [the process of cutting and splicing of RNA after DNA transcription] in the outcome of splicing. However, with this work they have tackled a monumental task, experimentally and systematically reducing the levels of 305 proteins involved in splicing (either in the central core of the machinery, the spliceosome, or in the constellation of factors that dynamically interact with it and regulate the splicing process), and then analyze the result of the absence or decrease of these proteins, i.e., what type of splicing mechanisms (there are four fundamental subtypes) increase or decrease in the absence of each of these 305 factors, which ultimately results in the formation of different protein variants with different functions. This is known to result in functional alterations that can lead to diseases such as cancer.

As is to be expected from a study published in this journal, the work presents a multitude of experimental verifications confirming the findings with different types of assays, which endorses its quality. In addition, the study fits superbly well with the knowledge available so far, while providing numerous original and relevant findings. Much of what we know about the functional and dynamic architecture of the spliceosome comes from electron cryomicroscopy studies, studies of protein interactions, functional and database annotations, etc. This study confirms with experimental data many of the theoretical predictions previously made, but also provides a remarkable amount of new findings that illuminate previously unknown structural and functional relationships and that will serve as a basis for further deepening the understanding of the spliceosome and deciphering how the splicing process functions and is regulated.

This is very relevant because the decision taken by the spliceosome within a cell to make one variant or another can determine whether a normal cell becomes cancerous or whether a neuron degenerates or dies. Therefore, knowing how the splicing machinery makes these decisions is crucial to identify new treatment targets and develop original therapeutic strategies for cancer, neurodegenerative pathologies or rare diseases.

The study does not present significant limitations, although by opening up so many new avenues it generates a multitude of unknowns that will have to be explored from now on. In fact, this is one of the virtues of the work, offering a large amount of information and laying the foundations for a comprehensive study of a lesser-known facet of the life of cells, how they decide to transform the information contained in a gene, which can give rise to a multitude of different proteins, into just one or a few variants with a specific function, and how this splicing process is altered in diseases.

One of the most curious and interesting findings is the demonstration of something that was already suspected, and that is that the splicing machinery has an amazing ability to regulate itself in a very complex and interconnected way. The functional meaning of this self-regulation is a challenge to be deciphered.

In the future, for example, it would be ideal to also extend this strategy to RNA types other than messenger RNA (encoding proteins) that also undergo the splicing process but are still much less well known, despite growing evidence of their relevance both in normal conditions and in numerous diseases.

Furthermore, there is the extraordinary challenge of deploying and extending the results of this study to understand in a complete and integrated manner the ultimate consequences of splicing alterations in the production of protein variants with specific functions, a task that will undoubtedly require highly complex experimental, technological and computational approaches.

This is an extraordinary work in which the human spliceosome is studied by the procedure of inactivating its genetic components and, subsequently, its functionality is evaluated in the corresponding transcriptomes and the regulatory networks and associated decision-making processes that this machinery carries out.
This is a fundamental study that helps to understand, in a way that had not been done before, the intricate process of alternative processing that is so characteristic and fundamental to eukaryotic cells. Here they study, with such a basic approach to genetics as the systematic inactivation of components, how it affects function, understood as the product that finally results. But this product is examined by determining the complex interaction networks of the spliceosome, something that is a novelty with respect to previous studies.
In terms of its implications, this study actually dissects how this fundamental machinery works, evaluating in a very fine-grained way how the whole works and how it is affected when any component does not carry out its function, as well as the global effects that this has. As the authors themselves indicate at the end of their work, they evaluate the physiology of a eukaryotic machinery, but also its pathological effects when it does not work properly.

The work opens up important avenues of research. The authors focus on the human spliceosome, but this is a general machinery in eukaryotes, by no means as complex in other species. It remains to be seen what has happened to its evolution.

Let's imagine that our genetic information is stored in a book, and its chapters are our genes. Genes carry the information needed to make proteins, the molecules that do the actual work in the cell. When a protein is required, a chapter has to be transcribed into a molecule that carries the instructions to make the protein. However, contrary to how we read books, following pages sequentially, cells need to remove chunks of pages before the  transcript makes sense. This process is know a splicing, and it is catalyzed by the spliceosome. Very importantly as well, a given transcript may undergo diverse splicing events, producing a molecule that will encode a different protein. This is known as alternative splicing (AS), and it is at the heart of many essential regulatory schemes. 

Transcripts with sequence errors of a single base, or in the incorrect amount, can lead to disease and death. Thus is not surprising that the spliceosome includes more than a hundred factors, and hundreds more (in humans) can regulate it. However, how all this complexity works is not clear. What is the cellular code behind AS? Can it be tweaked to our advantage (e.g. to fight disease?) This question has been addressed a number of times, and we have learn about the function of some splicing factors at genomic level, or their role in all chapters (not a small feat). What it has been missing has been a more global approach. In this context, the work of Rogalska et al. is a very significant advance in our understanding of regulated splicing.  

Rogalska et al. have integrated the data from many experiments, each one being the output of the spliceosome (known as "transcriptome") when one of its components, or one of the splicing regulators, is missing. Each transcriptome is defined by the type and frequency of removed sets of pages. Thus, if the lack of two factors lead to similar transcriptomes, these factors must be functionally related. Following this rationale, and with a real tour de force, they have been able to produce a number of maps based on the closeness between transcriptomes. These maps,or networks of related factors, offer a blueprint of the inner works of the spliceosome and how they can be regulated. Importantly, they are consistent with previous knowledge of the spliceosome, which gives strong support to important predictions that can be made.

Examples of these findings are, a role in regulation for spliceosomal factors long thought to be known, or linking specific outcomes to regulatory splicing factors (surely very relevant to pharma research). Surely more will come, as their interaction maps are applied to other questions. The group of Juan Valcárcel has shown a number of times how dedication and courage to address complex questions can lead to significant discoveries.

It is important to keep in mind that this blueprint is based on looking at transcriptomes after a particular treatment to the cells. What other blueprints will be generated by other treatments? How robust are they? The sobering reality is that we are still far from fully understanding how genomes like ours work.  Without that, we will not be able to properly fight disease. The work of Rogalska et al. Is a significant step towards this goal.

In summary, Rogalska et al. have provided a blueprint of how the cellular collection of messages  to make proteins (transcriptome) is affected by changes in the machinery that  makes it. This blueprint includes the impact of hundreds of factors, thus providing a very valuable tool to develop drugs that can restore the proper transcriptome. Further research will be needed for this, but having a blueprint of effects and their interactions will be extremely valuable.