Internal transcribed spacer

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Internal transcribed spacer (ITS) is the spacer DNA situated between the small-subunit ribosomal RNA (rRNA) and large-subunit rRNA genes in the chromosome or the corresponding transcribed region in the polycistronic rRNA precursor transcript.

Contents

Across life domains

In bacteria and archaea, there is a single ITS, located between the 16S and 23S rRNA genes. Conversely, there are two ITSs in eukaryotes: ITS1 is located between 18S and 5.8S rRNA genes, while ITS2 is between 5.8S and 28S (in opisthokonts, or 25S in plants) rRNA genes. ITS1 corresponds to the ITS in bacteria and archaea, while ITS2 originated as an insertion that interrupted the ancestral 23S rRNA gene. [1] [2]

Organization

Organization of the eukaryotic nuclear ribosomal DNA tandem repeats Eucaryot rdna.png
Organization of the eukaryotic nuclear ribosomal DNA tandem repeats

In bacteria and archaea, the ITS occurs in one to several copies, as do the flanking 16S and 23S genes. When there are multiple copies, these do not occur adjacent to one another. Rather, they occur in discrete locations in the circular chromosome. It is not uncommon in bacteria to carry tRNA genes in the ITS. [3] [4]

In eukaryotes, genes encoding ribosomal RNA and spacers occur in tandem repeats that are thousands of copies long, each separated by regions of non-transcribed DNA termed intergenic spacer (IGS) or non-transcribed spacer (NTS).

Each eukaryotic ribosomal cluster contains the 5' external transcribed spacer (5' ETS), the 18S rRNA gene, the ITS1, the 5.8S rRNA gene, the ITS2, the 26S or 28S rRNA gene, and finally the 3' ETS. [5]

During rRNA maturation, ETS and ITS pieces are excised. As non-functional by-products of this maturation, they are rapidly degraded. [6]

Use in phylogenetic inference

Sequence comparison of the eukaryotic ITS regions is widely used in taxonomy and molecular phylogeny because of several favorable properties: [7]

For example, ITS markers have proven especially useful for elucidating phylogenetic relationships among the following taxa.

Taxonomic groupTaxonomic levelYearAuthors with references
Asteraceae: Compositae Species (congeneric)1992Baldwin et al. [9]
Viscaceae: Arceuthobium Species (congeneric)1994Nickrent et al. [10]
Poaceae: Zea Species (congeneric)1996Buckler & Holtsford [11]
Leguminosae: Medicago Species (congeneric)1998Bena et al. [5]
Orchidaceae: Diseae Genera (within tribes)1999Douzery et al. [12]
Odonata: Calopteryx Species (congeneric)2001Weekers et al. [13]
Yeasts of clinical importanceGenera2001Chen et al. [14]
Poaceae: SaccharinaeGenera (within tribes)2002Hodkinson et al. [15]
Plantaginaceae: Plantago Species (congeneric)2002Rønsted et al. [16]
Jungermanniopsida: Herbertus Species (congeneric)2004Feldberg et al. [17]
Pinaceae: Tsuga Species (congeneric)2008Havill et al. [18]
Chrysomelidae: Altica Genera (congeneric)2009Ruhl et al. [19]
Symbiodinium Clade2009Stat et al. [20]
Brassicaceae Tribes (within a family)2010Warwick et al. [21]
Ericaceae: Erica Species (congeneric)2011Pirie et al. [22]
Diptera: Bactrocera Species (congeneric)2014Boykin et al. [23]
Scrophulariaceae: Scrophularia Species (congeneric)2014Scheunert & Heubl [24]
Potamogetonaceae: Potamogeton Species (congeneric)2016Yang et al. [25]

ITS2 is known to be more conserved than ITS1 is. All ITS2 sequences share a common core of secondary structure, [26] while ITS1 structures are only conserved in much smaller taxonomic units. Regardless of the scope of conservation, structure-assisted comparison can provide higher resolution and robustness. [27]

Mycological barcoding

The ITS region is the most widely sequenced DNA region in molecular ecology of fungi [28] and has been recommended as the universal fungal barcode sequence. [29] It has typically been most useful for molecular systematics at the species to genus level, and even within species (e.g., to identify geographic races). Because of its higher degree of variation than other genic regions of rDNA (for example, small- and large-subunit rRNA), variation among individual rDNA repeats can sometimes be observed within both the ITS and IGS regions. In addition to the universal ITS1+ITS4 primers [30] [31] used by many labs, several taxon-specific primers have been described that allow selective amplification of fungal sequences (e.g., see Gardes & Bruns 1993 paper describing amplification of basidiomycete ITS sequences from mycorrhiza samples). [32] Despite shotgun sequencing methods becoming increasingly utilized in microbial sequencing, the low biomass of fungi in clinical samples make the ITS region amplification an area of ongoing research. [33] [34]

References

  1. ↑ Lafontaine, D. L. J.; Tollervey, D. (2001). "The function and synthesis of ribosomes". Nature Reviews Molecular Cell Biology. 2 (7): 514–520. doi:10.1038/35080045. hdl: 1842/729 . PMID   11433365. S2CID   2637106.
  2. ↑ Scott Orland Rogers (27 July 2011). Integrated Molecular Evolution. CRC Press. pp. 65–66. ISBN   978-1-4398-1995-1 . Retrieved 9 March 2015.
  3. ↑ Takada, Hiraku; Shimada, Tomohiro; Dey, Debashish; Quyyum, M. Zuhaib; Nakano, Masahiro; Ishiguro, Akira; Yoshida, Hideji; Yamamoto, Kaneyoshi; Sen, Ranjan; Ishihama, Akira (22 December 2016). "Differential Regulation of rRNA and tRNA Transcription from the rRNA-tRNA Composite Operon in Escherichia coli". PLOS ONE. 11 (12) e0163057. Bibcode:2016PLoSO..1163057T. doi: 10.1371/journal.pone.0163057 . PMC   5179076 . PMID   28005933.
  4. ↑ Stewart, Frank J.; Cavanaugh, Colleen M. (July 2007). "Intragenomic Variation and Evolution of the Internal Transcribed Spacer of the rRNA Operon in Bacteria". Journal of Molecular Evolution. 65 (1): 44–67. Bibcode:2007JMolE..65...44S. CiteSeerX   10.1.1.456.2659 . doi:10.1007/s00239-006-0235-3. PMID   17568983. S2CID   13536182.
  5. 1 2 Bena, Gilles; Jubier, Marie-France; Olivieri, Isabelle; Lejeune, Bernard (1998). "Ribosomal External and Internal Transcribed Spacers: Combined Use in the Phylogenetic Analysis of Medicago (Leguminosae)". Journal of Molecular Evolution. 46 (3): 299–306. Bibcode:1998JMolE..46..299B. doi:10.1007/PL00006306. ISSN   0022-2844. PMID   9502673. S2CID   38838013.
  6. ↑ Michot, Bernard; Bachellerie, Jean-Pierre; Raynal, Francoise (1983-05-25). "Structure of mouse rRNA precursors. Complete sequence and potential folding of the spacer regions between 18S and 28S rRNA". Nucleic Acids Research. 11 (10): 3375–3391. doi:10.1093/nar/11.10.3375. ISSN   0305-1048. PMC   325970 . PMID   6304630.
  7. ↑ Baldwin, Bruce G.; Sanderson, Michael J.; Porter, J. Mark; Wojciechowski, Martin F.; Campbell, Christopher S.; Donoghue, Michael J. (1995-01-01). "The ITS Region of Nuclear Ribosomal DNA: A Valuable Source of Evidence on Angiosperm Phylogeny". Annals of the Missouri Botanical Garden. 82 (2): 247–277. doi:10.2307/2399880. JSTOR   2399880.
  8. ↑ Song, Jingyuan; Shi, Linchun; Li, Dezhu; Sun, Yongzhen; Niu, Yunyun; Chen, Zhiduan; Luo, Hongmei; Pang, Xiaohui; Sun, Zhiying (2012-08-30). "Extensive Pyrosequencing Reveals Frequent Intra-Genomic Variations of Internal Transcribed Spacer Regions of Nuclear Ribosomal DNA". PLOS ONE. 7 (8) e43971. Bibcode:2012PLoSO...743971S. doi: 10.1371/journal.pone.0043971 . ISSN   1932-6203. PMC   3431384 . PMID   22952830.
  9. ↑ Baldwin, B.G. (1992). "Phylogenetic utility of the internal transcribed spacers of nuclear ribosomal DNA in plants: An example from the Compositae". Molecular Phylogenetics and Evolution. 1 (1): 3–16. doi:10.1016/1055-7903(92)90030-K. PMID   1342921.
  10. ↑ Nickrent, Daniel L.; Schuette, Kevin P.; Starr, Ellen M. (1994-01-01). "A Molecular Phylogeny of Arceuthobium (Viscaceae) Based on Nuclear Ribosomal DNA Internal Transcribed Spacer Sequences". American Journal of Botany. 81 (9): 1149–1160. doi:10.2307/2445477. JSTOR   2445477.
  11. ↑ Buckler, E. S.; Holtsford, T. P. (1996-04-01). "Zea systematics: ribosomal ITS evidence". Molecular Biology and Evolution. 13 (4): 612–622. doi:10.1093/oxfordjournals.molbev.a025621. ISSN   0737-4038. PMID   8882504.
  12. ↑ Douzery, Emmanuel J. P.; Pridgeon, Alec M.; Kores, Paul; Linder, H. P.; Kurzweil, Hubert; Chase, Mark W. (1999-06-01). "Molecular phylogenetics of Diseae (Orchidaceae): a contribution from nuclear ribosomal ITS sequences". American Journal of Botany. 86 (6): 887–899. doi:10.2307/2656709. ISSN   0002-9122. JSTOR   2656709. PMID   10371730.(subscription required)
  13. ↑ Weekers, Peter H. H.; De Jonckheere, Johan F.; Dumont, Henri J. (2001-07-01). "Phylogenetic Relationships Inferred from Ribosomal ITS Sequences and Biogeographic Patterns in Representatives of the Genus Calopteryx (Insecta: Odonata) of the West Mediterranean and Adjacent West European Zone". Molecular Phylogenetics and Evolution. 20 (1): 89–99. doi:10.1006/mpev.2001.0947. PMID   11421650.
  14. ↑ Chen, Y-C, J. D. Eisner, M. M. Kattar, S. L. Rassoulian-Barrett, K. Lafe, A. P. Limaye, and B. T. Cookson (2001). "Polymorphic Internal Transcribed Spacer Region 1 DNA Sequences Identify Medically Important Yeasts". J. Clin. Microbiol. 39 (11): 4042–4051. doi:10.1128/JCM.39.11.4042-4051.2001. PMC   88485 . PMID   11682528.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  15. ↑ Hodkinson, Trevor R.; Chase, Mark W.; Lledó, Dolores M.; Salamin, Nicolas; Renvoize, Stephen A. (2002). "Phylogenetics of Miscanthus, Saccharum and related genera (Saccharinae, Andropogoneae, Poaceae) based on DNA sequences from ITS nuclear ribosomal DNA and plastid trnL intron and trnL-F intergenic spacers". Journal of Plant Research. 115 (5): 381–392. Bibcode:2002JPlR..115..381H. doi:10.1007/s10265-002-0049-3. ISSN   0918-9440. PMID   12579363. S2CID   22971617.
  16. ↑ Rønsted, Nina; Chase, Mark W.; Albach, Dirk C.; Bello, Maria Angelica (2002-08-01). "Phylogenetic relationships within Plantago (Plantaginaceae): evidence from nuclear ribosomal ITS and plastid trnL-F sequence data". Botanical Journal of the Linnean Society. 139 (4): 323–338. doi: 10.1046/j.1095-8339.2002.00070.x . ISSN   1095-8339.
  17. ↑ Feldberg, K.; Groth, H.; Wilson, R.; Schäfer-Verwimp, A.; Heinrichs, J. (2004-11-04). "Cryptic speciation in Herbertus (Herbertaceae, Jungermanniopsida): Range and morphology of Herbertus sendtneri inferred from nrITS sequences". Plant Systematics and Evolution. 249 (3–4): 247–261. Bibcode:2004PSyEv.249..247F. doi:10.1007/s00606-004-0221-4. ISSN   0378-2697. S2CID   21538862.
  18. ↑ Havill, Nathan P.; Campbell, Christopher S.; Vining, Thomas F.; LePage, Ben; Bayer, Randall J.; Donoghue, Michael J. (2008-07-01). "Phylogeny and Biogeography of Tsuga (Pinaceae) Inferred from Nuclear Ribosomal ITS and Chloroplast DNA Sequence Data". Systematic Botany. 33 (3): 478–489. doi:10.1600/036364408785679770. S2CID   26668467.
  19. ↑ Ruhl, Michael W.; Wolf, Matthias; Jenkins, Tracie M. (2010). "Compensatory base changes illuminate morphologically difficult taxonomy". Molecular Phylogenetics and Evolution. 54 (2): 664–669. doi:10.1016/j.ympev.2009.07.036. PMID   19660561.
  20. ↑ Stat, Michael; Pochon, Xavier (2008-07-02). "Specificity in communities of Symbiodinium in corals from Johnston Atoll" (PDF). Marine Ecology Progress Series. 386: 83–96. doi: 10.3354/meps08080 .
  21. ↑ Warwick, Suzanne I.; Mummenhoff, Klaus; Sauder, Connie A.; Koch, Marcus A.; Al-Shehbaz, Ihsan A. (2010-04-13). "Closing the gaps: phylogenetic relationships in the Brassicaceae based on DNA sequence data of nuclear ribosomal ITS region". Plant Systematics and Evolution. 285 (3–4): 209–232. Bibcode:2010PSyEv.285..209W. doi:10.1007/s00606-010-0271-8. ISSN   0378-2697. S2CID   28199415.
  22. ↑ Pirie, Michael D.; Oliver, E. G. H.; Bellstedt, Dirk U. (2011-11-01). "A densely sampled ITS phylogeny of the Cape flagship genus Erica L. suggests numerous shifts in floral macro-morphology". Molecular Phylogenetics and Evolution. 61 (2): 593–601. doi:10.1016/j.ympev.2011.06.007. PMID   21722743.
  23. ↑ Boykin, L. M.; Schutze, M. K.; Krosch, M. N.; Chomič, A.; Chapman, T. A.; Englezou, A.; Armstrong, K. F.; Clarke, A. R.; Hailstones, D. (2014-05-01). "Multi-gene phylogenetic analysis of south-east Asian pest members of the Bactrocera dorsalis species complex (Diptera: Tephritidae) does not support current taxonomy". Journal of Applied Entomology. 138 (4): 235–253. doi:10.1111/jen.12047. ISSN   1439-0418. S2CID   82003038.
  24. ↑ Scheunert, Agnes; Heubl, Günther (2014-01-01). "Diversification of Scrophularia (Scrophulariaceae) in the Western Mediterranean and Macaronesia – Phylogenetic relationships, reticulate evolution and biogeographic patterns". Molecular Phylogenetics and Evolution. 70: 296–313. doi:10.1016/j.ympev.2013.09.023. PMID   24096055.
  25. ↑ Yang, Tao; Zhang, Tian-lei; Guo, You-hao; Liu, Xing (2016-11-17). "Identification of Hybrids in Potamogeton: Incongruence between Plastid and ITS Regions Solved by a Novel Barcoding Marker PHYB". PLOS ONE. 11 (11) e0166177. Bibcode:2016PLoSO..1166177Y. doi: 10.1371/journal.pone.0166177 . ISSN   1932-6203. PMC   5113904 . PMID   27855191.
  26. ↑ Schultz, J; Maisel, S; Gerlach, D; Müller, T; Wolf, M (April 2005). "A common core of secondary structure of the internal transcribed spacer 2 (ITS2) throughout the Eukaryota". RNA. 11 (4): 361–4. doi:10.1261/rna.7204505. PMC   1370725 . PMID   15769870.
  27. ↑ Koetschan, C; Kittelmann, S; Lu, J; Al-Halbouni, D; Jarvis, GN; Müller, T; Wolf, M; Janssen, PH (2014). "Internal transcribed spacer 1 secondary structure analysis reveals a common core throughout the anaerobic fungi (Neocallimastigomycota)". PLOS ONE. 9 (3) e91928. Bibcode:2014PLoSO...991928K. doi: 10.1371/journal.pone.0091928 . PMC   3963862 . PMID   24663345.
  28. ↑ Peay K.G.; Kennedy P.G.; Bruns T.D. (2008). "Fungal community ecology: a hybrid beast with a molecular master". BioScience. 58 (9): 799–810. doi: 10.1641/b580907 . S2CID   18363490.
  29. ↑ Schoch, C.L., Seifert, K.A., Huhndorf, S., Robert, V., Spouge, J.L., Levesque, C.A., Chen, W., Bolchacova, E., Voigt, K., Crous, P.W.; et al. (2012). "Nuclear Ribosomal Internal Transcribed Spacer (ITS) Region as a Universal DNA Barcode Marker for Fungi". PNAS. 109 (16): 6241–6246. doi: 10.1073/pnas.1117018109 . PMC   3341068 . PMID   22454494.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  30. ↑ White, T.J., Bruns, T., Lee, S., and Taylor, J. (1990). Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. PCR Protocols: a Guide to Methods and Applications 18, 315–322.
  31. ↑ The ITS1 primer covers ITS1-5.8S-ITS2 from the 5', and ITS4 covers the same area from the 3'.
  32. ↑ Gardes, M.; Bruns, T.D. (1993). "ITS primers with enhanced specificity for basidiomycetes: application to the identification of mycorrhiza and rusts". Molecular Ecology. 2 (2): 113–118. doi:10.1111/j.1365-294X.1993.tb00005.x. PMID   8180733. S2CID   24316407.
  33. ↑ Usyk, Mykhaylo; Zolnik, Christine P.; Patel, Hitesh; Levi, Michael H.; Burk, Robert D. (2017-12-13). Mitchell, Aaron P. (ed.). "Novel ITS1 Fungal Primers for Characterization of the Mycobiome". mSphere. 2 (6): e00488–17, /msphere/2/6/mSphere0488–17.atom. doi:10.1128/mSphere.00488-17. ISSN   2379-5042. PMC   5729218 . PMID   29242834.
  34. ↑ Nilsson, R. Henrik; Anslan, Sten; Bahram, Mohammad; Wurzbacher, Christian; Baldrian, Petr; Tedersoo, Leho (February 2019). "Mycobiome diversity: high-throughput sequencing and identification of fungi". Nature Reviews Microbiology. 17 (2): 95–109. doi:10.1038/s41579-018-0116-y. ISSN   1740-1534. PMID   30442909. S2CID   53438777.
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