Journal of Molecular Biology
Alternative Splicing Mechanisms for the Modulation of Protein Function: Conservation Between Human and Other Species
Introduction
Alternative splicing (AS) is one of the most important sources of protein functional diversity in eukaryotes.1., 2. At the protein sequence level, AS generates this diversity using two different mechanisms:3 insertions/deletions (IDs) and substitutions (Ss). The biological impact of AS may go from drastic switch-off effects, as those produced by large IDs, to smoother functional changes usually associated with Ss. For example, AS of transcription factors contributes to the control of gene expression during developmental processes4 by changing DNA-binding properties,5 by introducing or eliminating activating domains6 or by increasing the in vivo stability of a given isoform.7 In the case of Drosophila Dscam,8 it is believed that mutually exclusive substitution of exons may result in protein isoforms with different recognition properties. For the main neurotransmitter receptors, it is AS that determines their cellular location.9
In general, all these mechanisms are used by the organism to modulate protein function within the context of biological processes that are relevant to the organism's complexity, like tissue differentiation or developmental processes.2., 10. It is then interesting to know their degree of conservation among different species, as it may provide valuable information for understanding the phenotypic differences between them.
Experimental evidence on this point is scarce, but suggests that conservation of AS patterns among species is probably related to the distance between the organisms,11., 12. and to the relevance for the organism's survival of the AS event.13., 14. Recent bioinformatics studies15 indicate that there is a high degree of AS conservation between human and mouse, at least for the major isoforms.16 However, to the extent of our knowledge, no comparative study has been done on the use and conservation of AS mechanisms for protein function modulation between species. Here, we address this issue, studying two different aspects of these mechanisms: (i) their frequency of use among four species with clear phenotypic differences, human, mouse, rat and fruitfly; (ii) their degree of conservation in pairs of biologically equivalent AS events, between human and mouse, and human and rat. Our results show that the different AS mechanisms for modulating protein function are present with similar frequencies both in human and in the three remaining species. We also have found that biologically equivalent AS events in human and mouse, and human and rat, display a high degree of similarity at the protein level.
Section snippets
Results
The mechanisms of AS for the modulation of protein function may be very different. In this work, we have characterized them using a series of protein sequence and structure properties that are normally utilised to assess the impact of protein sequence/structure changes in protein function: size,17., 18. solvent accessibility and secondary structure location,17., 19. and nature of the amino acid changes.18 Size was chosen because it is related to the effect of protein sequence changes in protein
Discussion
AS is an important process for modulating protein function in eukaryotic organisms.1., 2., 10. This is done through a rich variety of mechanisms, including deletion of activation domains,6 creation of additional binding sites,21 substitution of sequence stretches,8 etc. Here, we have studied the degree of conservation of these mechanisms among species, concentrating on two aspects. First, we have studied the frequency with which the different AS mechanisms for modulating protein function are
Protein isoforms
A set of different proteins together with their known isoforms, for human, mouse, rat and fruitfly, was obtained after searching the SwissProt27 database version 39, using as query keywords VARSPLIC and HUMAN, or MOUSE, or RAT, or DROME, respectively. AS events annotated by similarity were excluded. A total of 802, 367, 220 and 121 human, mouse, rat and fruitfly proteins, respectively, were kept; with a total of 2218, 972, 624 and 314 isoforms.
Accessibility and secondary structure predictions
Secondary structure and accessibility predictions
Acknowledgements
The authors acknowledge one of the referees for his insightful comments on the manuscript. They also acknowledge Miquel Calvo for helpful comments on the statistical treatment of the data. X.C. acknowledges the economical support of the Spanish Ministry of Science and Technology (grant GEN2001-4758-C07-07).
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A.V. and D.T. contributed equally to this work.