Dogma in doubt

Scientists have found that alternative splicing does not lead to protein diversity

PR team of IBH RAS, "Open Science"

Staff The Institute of Bioorganic Chemistry of the Russian Academy of Sciences and the FNCC of Physico-Chemical Medicine of the FMBA of Russia found that alternative splicing of matrix RNAs makes a significantly smaller contribution to the diversity of proteins inside the cell. The results published in the journal Scientific Reports change our understanding of the main dogma of molecular biology "DNA-RNA-protein".

"Starting the study, we wanted to see what function alternative forms of protein (isoforms) that are formed as a result of alternative splicing can perform. However, during the study of cell proteins, we almost did not find the isoforms themselves. Apparently, the long-known process of alternative splicing is not so important for the production of new proteins, as they say in textbooks, but is needed for something else, for which we have yet to find out," says Igor Fesenko, Candidate of Biological Sciences, Employee Proteomics Laboratories M.M. Shemyakin and Yu.A. Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences.

Cells of all organisms, except bacteria and archaea, have a nucleus. Nuclear (eukaryotic) RNA molecules can undergo alternative splicing – the formation of several matrix RNAs from one that can be translated into different isoform proteins. Thus, it was previously assumed that one gene (a section of DNA) gives, thanks to splicing, several RNAs that form many proteins.

"When scientists discovered that eukaryotic genes can undergo alternative splicing, they assumed that due to this process, the gene can encode a huge number of different proteins. This is partly true: in any organism there are genes that encode many different protein isoforms. But in our study we have shown that in general this is not the case. Earlier, when studying the role of alternative splicing in various processes, biologists did not take this point into account in their work," says Igor.

Igor and his colleagues began their research with standard methods of gene expression analysis (transformation of DNA into RNA and protein): RNA was isolated from different cell types of a model object (Physcomitrella patens moss), the primary structure was determined (sequenced) and predicted on a computer which protein isoforms could be read from such RNAs. Then the scientists affected the moss cells with various stress factors, repeated all the manipulations and found that alternative splicing actively changes the matrix RNAs after stress. To confirm the hypothesis about the presence of predicted proteins in the cell and the participation of alternative splicing in changing their composition, scientists isolated proteins from moss cells and performed an analysis on a mass spectrometer - a device that "weighs" molecules and can identify proteins. It turned out that the role of alternative splicing in the formation of the protein composition of the cell is insignificant.

"The reviewers of the journal where we submitted the article suggested that the results we obtained were related to poorly performed mass spectrometric analysis," Igor continues. – To prove our case, we modeled an experiment in which we estimated the number of isoforms that we would have to identify if they were present in the cell. It turned out that if alternative splicing plays an important role in the formation of the protein composition of the cell, we would have seen dozens of times more isoforms in our mass spectrometric experiment than we actually received. So we proved that the role of alternative splicing is more complicated than previously thought."

A large variety of RNA leads to very small changes in proteins, therefore, according to the authors of the study, the canonical scheme of molecular biology "from DNA to protein" is violated. Now scientists have to understand what biological function alternative splicing has, what is the role of RNA-RNA interactions in this process and how all this affects the translation of proteins in the cell.

Portal "Eternal youth" http://vechnayamolodost.ru  13.06.2017