Where the minor things are: a pan-eukaryotic survey suggests neutral processes may explain much of minor intron evolution

Abstract Spliceosomal introns are gene segments removed from RNA transcripts by ribonucleoprotein machineries called spliceosomes. In some eukaryotes a second ‘minor’ spliceosome is responsible for processing a tiny minority of introns. Despite its seemingly modest role, minor splicing has persisted for roughly 1.5 billion years of eukaryotic evolution. Identifying minor introns in over 3000 eukaryotic genomes, we report diverse evolutionary histories including surprisingly high numbers in some fungi and green algae, repeated loss, as well as general biases in their positional and genic distributions. We estimate that ancestral minor intron densities were comparable to those of vertebrates, suggesting a trend of long-term stasis. Finally, three findings suggest a major role for neutral processes in minor intron evolution. First, highly similar patterns of minor and major intron evolution contrast with both functionalist and deleterious model predictions. Second, observed functional biases among minor intron-containing genes are largely explained by these genes’ greater ages. Third, no association of intron splicing with cell proliferation in a minor intron-rich fungus suggests that regulatory roles are lineage-specific and thus cannot offer a general explanation for minor splicing’s persistence. These data constitute the most comprehensive view of minor introns and their evolutionary history to date, and provide a foundation for future studies of these remarkable genetic elements.

[1]  Susana M. Coelho,et al.  Uncovering gene-family founder events during major evolutionary transitions in animals, plants and fungi using GenEra , 2023, Genome Biology.

[2]  Susana M. Coelho,et al.  Uncovering gene-family founder events during major evolutionary transitions in animals, plants and fungi using GenEra , 2022, bioRxiv.

[3]  P. Bork,et al.  Interactive Tree Of Life (iTOL) v5: an online tool for phylogenetic tree display and annotation , 2021, Nucleic Acids Res..

[4]  I. Ruiz-Trillo,et al.  The Protistan Origins of Animal Cell Differentiation , 2021 .

[5]  H. Drost,et al.  Sensitive protein alignments at tree-of-life scale using DIAMOND , 2021, Nature Methods.

[6]  J. Diedrich,et al.  Coupling of spliceosome complexity to intron diversity , 2021, Current Biology.

[7]  S. Spirin,et al.  Is there any intron sliding in mammals? , 2020, BMC evolutionary biology.

[8]  M. Eliáš,et al.  Expansion and transformation of the minor spliceosomal system in the slime mold Physarum polycephalum , 2020, Current Biology.

[9]  R. Padgett,et al.  Comprehensive database and evolutionary dynamics of U12-type introns , 2020, Nucleic acids research.

[10]  B. Holland,et al.  Accuracy of ancestral state reconstruction for non-neutral traits , 2020, Scientific Reports.

[11]  V. Tarabykin,et al.  Srsf10 and the minor spliceosome control tissue-specific and dynamic SR protein expression , 2020, eLife.

[12]  T. Maeda,et al.  Structure-specific Regulation of Nutrient Transport and Metabolism in Arbuscular Mycorrhizal Fungi. , 2019, Plant & cell physiology.

[13]  N. King,et al.  Light-regulated collective contractility in a multicellular choanoflagellate , 2019, Science.

[14]  Boas Pucker,et al.  Animal, Fungi, and Plant Genome Sequences Harbor Different Non-Canonical Splice Sites , 2019, bioRxiv.

[15]  Boas Pucker,et al.  Genome-wide analyses supported by RNA-Seq reveal non-canonical splice sites in plant genomes , 2018, BMC Genomics.

[16]  T. Thomas,et al.  Early developmental arrest and impaired gastrointestinal homeostasis in U12-dependent splicing-defective Rnpc3-deficient mice , 2018, RNA: A publication of the RNA Society.

[17]  Sahar Al Seesi,et al.  Minor spliceosome inactivation causes microcephaly, owing to cell cycle defects and death of self-amplifying radial glial cells , 2018, Development.

[18]  R. Davenport,et al.  RNA Binding Motif Protein 48 Is Required for U12 Splicing and Maize Endosperm Differentiation , 2018, Plant Cell.

[19]  F. Spiegel,et al.  Variation in the SSUrDNA of the Genus Protostelium Leads to a New Phylogenetic Understanding of the Genus and of the Species Concept for Protostelium mycophaga (Protosteliida, Amoebozoa) , 2018, The Journal of eukaryotic microbiology.

[20]  Jernej Ule,et al.  A retained intron in the 3′‐UTR of Calm3 mRNA mediates its Staufen2‐ and activity‐dependent localization to neuronal dendrites , 2017, EMBO reports.

[21]  Anastasios D. Tsaousis,et al.  Extreme genome diversity in the hyper-prevalent parasitic eukaryote Blastocystis , 2017, PLoS biology.

[22]  E. Eyras,et al.  IRFinder: assessing the impact of intron retention on mammalian gene expression , 2017, Genome Biology.

[23]  Geet Duggal,et al.  Salmon: fast and bias-aware quantification of transcript expression using dual-phase inference , 2017, Nature Methods.

[24]  W. Barbazuk,et al.  Aberrant splicing in maize rough endosperm3 reveals a conserved role for U12 splicing in eukaryotic multicellular development , 2017, Proceedings of the National Academy of Sciences.

[25]  M. Csűrös,et al.  Splice Sites Seldom Slide: Intron Evolution in Oomycetes , 2016, Genome biology and evolution.

[26]  Christopher R. Sibley,et al.  Lessons from non-canonical splicing , 2016, Nature Reviews Genetics.

[27]  S. Roy How Common Is Parallel Intron Gain? Rapid Evolution Versus Independent Creation in Recently Created Introns in Daphnia. , 2016, Molecular biology and evolution.

[28]  M. Robinson,et al.  Differential analyses for RNA-seq: transcript-level estimates improve gene-level inferences , 2015, F1000Research.

[29]  Evgeny M. Zdobnov,et al.  BUSCO: assessing genome assembly and annotation completeness with single-copy orthologs , 2015, Bioinform..

[30]  R. Lanfear,et al.  Phylogenetic uncertainty can bias the number of evolutionary transitions estimated from ancestral state reconstruction methods. , 2015, Journal of experimental zoology. Part B, Molecular and developmental evolution.

[31]  W. Huber,et al.  Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2 , 2014, Genome Biology.

[32]  R. Munita,et al.  A comprehensive survey of non-canonical splice sites in the human transcriptome , 2014, Nucleic acids research.

[33]  M. Irimia,et al.  Origin of spliceosomal introns and alternative splicing. , 2014, Cold Spring Harbor perspectives in biology.

[34]  A. Gregory Matera,et al.  A day in the life of the spliceosome , 2014, Nature Reviews Molecular Cell Biology.

[35]  Sean R. Eddy,et al.  Infernal 1.1: 100-fold faster RNA homology searches , 2013, Bioinform..

[36]  M. Szcześniak,et al.  ERISdb: a database of plant splice sites and splicing signals. , 2013, Plant & cell physiology.

[37]  M. Moore,et al.  Introns in UTRs: Why we should stop ignoring them , 2012, BioEssays : news and reviews in molecular, cellular and developmental biology.

[38]  M. Frilander,et al.  The significant other: splicing by the minor spliceosome , 2012, Wiley interdisciplinary reviews. RNA.

[39]  Liran Carmel,et al.  Origin and evolution of spliceosomal introns , 2012, Biology Direct.

[40]  Chiao-Feng Lin,et al.  U12-type Spliceosomal Introns of Insecta , 2012, International journal of biological sciences.

[41]  Scott Federhen,et al.  The NCBI Taxonomy database , 2011, Nucleic Acids Res..

[42]  C. Will,et al.  Spliceosome structure and function. , 2011, Cold Spring Harbor perspectives in biology.

[43]  B. Langmead,et al.  Aligning Short Sequencing Reads with Bowtie , 2010, Current protocols in bioinformatics.

[44]  Sridhar Hannenhalli,et al.  Young proteins experience more variable selection pressures than old proteins. , 2010, Genome research.

[45]  Chiao-Feng Lin,et al.  Evolutionary dynamics of U12-type spliceosomal introns , 2010, BMC Evolutionary Biology.

[46]  T. Samuelsson,et al.  U12 type introns were lost at multiple occasions during evolution , 2010, BMC Genomics.

[47]  Ning Ma,et al.  BLAST+: architecture and applications , 2009, BMC Bioinformatics.

[48]  S. Roy Intronization, de-intronization and intron sliding are rare in Cryptococcus , 2009, BMC Evolutionary Biology.

[49]  Eugene V Koonin,et al.  The universal distribution of evolutionary rates of genes and distinct characteristics of eukaryotic genes of different apparent ages , 2009, Proceedings of the National Academy of Sciences.

[50]  P. Sharp,et al.  Proliferating Cells Express mRNAs with Shortened 3' Untranslated Regions and Fewer MicroRNA Target Sites , 2008, Science.

[51]  M. Jurica Detailed close-ups and the big picture of spliceosomes. , 2008, Current opinion in structural biology.

[52]  E. Koonin,et al.  U12 intron positions are more strongly conserved between animals and plants than U2 intron positions , 2008, Biology Direct.

[53]  Marcela Dávila López,et al.  Computational screen for spliceosomal RNA genes aids in defining the phylogenetic distribution of major and minor spliceosomal components , 2008, Nucleic acids research.

[54]  R. Qu,et al.  Gene expression enhancement mediated by the 5′ UTR intron of the rice rubi3 gene varied remarkably among tissues in transgenic rice plants , 2008, Molecular Genetics and Genomics.

[55]  F. Müller,et al.  Splicing Segregation: The Minor Spliceosome Acts outside the Nucleus and Controls Cell Proliferation , 2007, Cell.

[56]  E. Koonin,et al.  Patterns of intron gain and conservation in eukaryotic genes , 2007, BMC Evolutionary Biology.

[57]  Tyler S. Alioto,et al.  U12DB: a database of orthologous U12-type spliceosomal introns , 2006, Nucleic Acids Res..

[58]  A. Russell,et al.  An early evolutionary origin for the minor spliceosome , 2006, Nature.

[59]  Hung D. Nguyen,et al.  Phase distribution of spliceosomal introns: implications for intron origin , 2006, BMC Evolutionary Biology.

[60]  R. Sachidanandam,et al.  Comprehensive splice-site analysis using comparative genomics , 2006, Nucleic acids research.

[61]  Chris M. Brown,et al.  Effect of 5'UTR introns on gene expression in Arabidopsis thaliana , 2006, BMC Genomics.

[62]  Walter Gilbert,et al.  The evolution of spliceosomal introns: patterns, puzzles and progress , 2006, Nature Reviews Genetics.

[63]  R. Russell,et al.  Animal MicroRNAs Confer Robustness to Gene Expression and Have a Significant Impact on 3′UTR Evolution , 2005, Cell.

[64]  Kui Lin,et al.  The excess of 5′ introns in eukaryotic genomes , 2005, Nucleic acids research.

[65]  R. Padgett,et al.  A mutational analysis of U12-dependent splice site dinucleotides. , 2005, RNA.

[66]  E. Koonin,et al.  Conservation versus parallel gains in intron evolution , 2005, Nucleic acids research.

[67]  Walter Gilbert,et al.  Complex early genes. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[68]  Walter Gilbert,et al.  The pattern of intron loss. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[69]  E. Koonin,et al.  Remarkable Interkingdom Conservation of Intron Positions and Massive, Lineage-Specific Intron Loss and Gain in Eukaryotic Evolution , 2003, Current Biology.

[70]  Tobias Mourier,et al.  Eukaryotic Intron Loss , 2003, Science.

[71]  W. Gilbert,et al.  Large-scale comparison of intron positions in mammalian genes shows intron loss but no gain , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[72]  M. Tomita,et al.  On biased distribution of introns in various eukaryotes. , 2002, Gene.

[73]  Anton J. Enright,et al.  An efficient algorithm for large-scale detection of protein families. , 2002, Nucleic acids research.

[74]  R. Durbin,et al.  A computational scan for U12-dependent introns in the human genome sequence. , 2001, Nucleic acids research.

[75]  V. Solovyev,et al.  Analysis of canonical and non-canonical splice sites in mammalian genomes. , 2000, Nucleic acids research.

[76]  A. Vinogradov Intron–Genome Size Relationship on a Large Evolutionary Scale , 1999, Journal of Molecular Evolution.

[77]  C. Cunningham Some Limitations of Ancestral Character-State Reconstruction When Testing Evolutionary Hypotheses , 1999 .

[78]  J. Steitz,et al.  Initial recognition of U12-dependent introns requires both U11/5' splice-site and U12/branchpoint interactions. , 1999, Genes & development.

[79]  P. Sharp,et al.  Evolutionary fates and origins of U12-type introns. , 1998, Molecular cell.

[80]  J. Logsdon,et al.  The recent origins of spliceosomal introns revisited. , 1998, Current opinion in genetics & development.

[81]  Christopher B. Burge,et al.  Classification of Introns: U2-Type or U12-Type , 1997, Cell.

[82]  R. Padgett,et al.  Terminal intron dinucleotide sequences do not distinguish between U2- and U12-dependent introns. , 1997, Molecular cell.

[83]  J D Palmer,et al.  Intron "sliding" and the diversity of intron positions. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[84]  Woan-Yuh Tarn,et al.  Highly Diverged U4 and U6 Small Nuclear RNAs Required for Splicing Rare AT-AC Introns , 1996, Science.

[85]  Woan-Yuh Tarn,et al.  A Novel Spliceosome Containing U11, U12, and U5 snRNPs Excises a Minor Class (AT–AC) Intron In Vitro , 1996, Cell.

[86]  W. Gilbert,et al.  Intron phase correlations and the evolution of the intron/exon structure of genes. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[87]  R. Padgett,et al.  Conserved sequences in a class of rare eukaryotic nuclear introns with non-consensus splice sites. , 1994, Journal of molecular biology.

[88]  L. Derr,et al.  A role for reverse transcripts in gene conversion , 1993, Nature.

[89]  Ian J.Jackson,et al.  A reappraisal of non-consensus mRNA splice sites. , 1991, Nucleic acids research.

[90]  Stephen M. Mount,et al.  A catalogue of splice junction sequences. , 1982, Nucleic acids research.

[91]  W. Gilbert Why genes in pieces? , 1978, Nature.

[92]  S. Tonegawa,et al.  Variable and constant parts of the immunoglobulin light chain gene of a mouse myeloma cell are 1250 nontranslated bases apart. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[93]  A. Jeffreys,et al.  The rabbit β-globin gene contains a large insert in the coding sequence , 1977, Cell.

[94]  L. Carmel,et al.  The role of reverse transcriptase in intron gain and loss mechanisms. , 2012, Molecular biology and evolution.

[95]  Victor V. Solovyev,et al.  SpliceDB: database of canonical and non-canonical mammalian splice sites , 2001, Nucleic Acids Res..

[96]  P. Sharp,et al.  Splicing of messenger RNA precursors. , 1985, Harvey lectures.

[97]  P Chambon,et al.  Organization and expression of eucaryotic split genes coding for proteins. , 1981, Annual review of biochemistry.