vavilovВавиловский журнал генетики и селекцииVavilov Journal of Genetics and Breeding2500-3259Institute of Cytology and Genetics of Siberian Branch of the RAS10.18699/VJ19.554vavilov-2332Research ArticleАктуальные технологии генетики растенийMainstream technologies in plant geneticsОбнаружение CRISPR-кассет и генов cas в геноме Arabidopsis thalianaDetection of CRISPR cassettes and cas genes in the Arabidopsis thaliana genomehttps://orcid.org/0000-0002-0601-2788КонстантиновЮ. М.KonstantinovYu. M.

Иркутск.

Irkutsk.

yukon@sifibr.irk.ruhttps://orcid.org/0000-0002-8788-5352ПетрушинИ. С.PetrushinI. S.

Иркутск.

Irkutsk.

Сибирский институт физиологии и биохимии растений Сибирского отделения Российской академии наук; Иркутский государственный университетРоссияSiberian Institute of Plant Physiology and Biochemistry, SB RAS; Irkutsk State UniversityRussian FederationИркутский государственный университетРоссияIrkutsk State UniversityRussian Federation201922112019237809816Copyright © Константинов Ю.М., Петрушин И.С., 20192019Константинов Ю.М., Петрушин И.С.Konstantinov Y.M., Petrushin I.S.This work is licensed under a Creative Commons Attribution 4.0 License.https://vavilov.elpub.ru/jour/article/view/2332

Современный уровень знаний в области эволюции растительных вирусов позволяет отнести проблему генетических основ противовирусного иммунитета высших растений (в том числе важнейших сельскохозяйственных культур) к разряду наиболее актуальных проблем генетики и селекции. В соответствии с эндосимбиотической теорией принято считать, что митохондрии произошли от альфа-протеобактерий, которые были поглощены, но не подвергнуты деструкции клеткой-хозяином. В связи с открытием у прокариот CRISPR-Cas (clustered regularly interspaced short palindromic repeats – CRISPR-associated proteins) систем, выполняющих функцию адаптивного иммунитета, возникает вопрос, мог ли подобный механизм противовирусной защиты быть подхвачен эволюцией и использован представителями эукариот, например растениями. Задачей настоящей работы был анализ полных последовательностей ядерного, митохондриального и хлоропластного геномов Arabidopsis thaliana с целью поиска генетических элементов, сходных с таковыми в CRISPR-Cas системах у бактерий и архей. В результате методами in silico в митохондриальном геноме экотипов A. thaliana обнаружен локус регулярно перемежающихся коротких прямых повторов, соответствующий по своей организации CRISPR-локусу адаптивного CRISPR-Cas иммунитета прокариот. На вероятную связь обнаруженного в митохондриальном геноме высшего растения локуса с функцией адаптивного иммунитета указывает наличие у спейсерных последовательностей в составе найденной СRISPR-кассеты гомологии с геномом вируса мозаики цветной капусты, поражающего растения арабидопсиса. У линий арабидопсиса С24 и Ler последовательности повторов и спейсеров CRISPR-кассеты полностью идентичны. В то же время локализация самого СRISPR-локуса в митохондриальном геноме этих линий существенно различается. Установлено, что у линии Col-0 в результате четырех делеций и одной инсерции CRISPR-кассета полностью нарушена. Хотя гены cas в митохондриальном геноме исследуемых экотипов арабидопсиса не были найдены, установлено их наличие в ядерном геноме. В ядерном геноме экотипа Col-0 на всех пяти хромосомах обнаружены гены cas и многочисленные CRISPR-кассеты. Полученные результаты позволяют предположить существование у растений системы адаптивного иммунитета, аналогичного CRISPR-иммунитету бактерий и архей.

The state of the art in the evolution of plant viruses allows the genetic foundations of antiviral immunity in higher (including the most important crops) plants to be categorized as one of the most pressing issues of genetics and selection. According to the endosymbiotic theory, mitochondria descended from alphaproteobacteria that had been absorbed but not degraded by the host cell. The discovery of CRISPR-Cas systems (clustered regularly interspaced short palindromic repeats (CRISPR)-associated proteins), which implement the adaptive immunity function in prokaryotes, raises the question whether such a mechanism of antiviral protection could be caught up by evolution and used by representatives of eukaryotes (in particular, plants). The purpose of this work was to analyze the complete sequences of nuclear, mitochondrial, and chloroplast genomes of Arabidopsis thaliana in order to search for genetic elements similar to those in CRISPR-Cas systems of bacteria and archaea. As a result, in silico methods helped us to detect a locus of regularly intermittent short direct repeats in the mitochondrial genome of A. thaliana ecotypes. The structure of this locus corresponds to the CRISPR locus of the prokaryotic adaptive antiviral immune system. The probable connection between the locus found in the mitochondrial genome of the higher plant and the function of adaptive immunity is indicated by a similarity between the spacer sequences in the CRISPR cassette found and the genome of Cauliflower mosaic virus affecting Arabidopsis plants. Sequences of repeats and spacers of CRISPR cassettes in Arabidopsis C24 and Ler lines are perfectly identical. However, the locations of the CRISPR locus in the mitochondrial genomes of these lines differ significantly. The CRISPR cassette in the Col-0 line was found to be completely broken as a result of four deletions and one insertion. Although cas genes were not detected in the mitochondrial genome of the studied Arabidopsis ecotypes, their presence was detected in the nuclear genome. Both cas genes and numerous CRISPR cassettes were found on all the five chromosomes in the nuclear genome of the Col-0 ecotype. The results suggest the existence of a system of adaptive immunity in plants, which is similar to the CRISPR immunity of bacteria and archaea.

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The authors declare that there are no conflicts of interest present.