CRISPR/Cas9 in Leishmania mexicana: A case study of LmxBTN1

Result code in IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F60076658%3A12310%2F18%3A43897402" target="_blank" >RIV/60076658:12310/18:43897402 - isvavai.cz</a>

Alternative codes found

RIV/60077344:_____/18:00498707 RIV/61988987:17310/18:A1901WOT RIV/00216208:11310/18:10373352

Result on the web

<a href="https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0192723&type=printable" target="_blank" >https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0192723&type=printable</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1371/journal.pone.0192723" target="_blank" >10.1371/journal.pone.0192723</a>

Result language

angličtina

Original language name

CRISPR/Cas9 in Leishmania mexicana: A case study of LmxBTN1

Original language description

Leishmania parasites cause human cutaneous, mucocutaneous and visceral leishmaniasis. Several studies proposed involvement of certain genes in infectivity of these parasites based on differential mRNA expression data. Due to unusual gene expression mechanism, functions of such genes must be further validated experimentally. Here, we investigated a role of one of the putative virulence factors, LmxM. 22.0010-encoded BTN1 (a protein involved in Batten disease in humans), in L. mexicana infectivity. Due to the incredible plasticity of the L. mexicana genome, we failed to obtain a complete knock-out of LmxM. 22.0010 using conventional recombination-based approach even after ablating four alleles of this gene. To overcome this, we established a modified CRISPR-Cas9 system with genomic expression of Cas9 nuclease and gRNA. Application of this system allowed us to establish a complete BTN1 KO strain of L. mexicana. The mutant strain did not show any difference in growth kinetics and differentiation in vitro, as well as in the infectivity for insect vectors and mice hosts. Based on the whole-transcriptome profiling, LmxM. 22.0010-encoded BTN1 was considered a putative factor of virulence in Leishmania. Our study suggests that ablation of LmxM. 22.0010 does not influence L. mexicana infectivity and further illustrates importance of experimental validation of in silico-predicted virulence factors. Here we also describe the whole genome sequencing of the widely used model isolate L. mexicana M379 and report a modified CRISPR/Cas9 system suitable for complete KO of multi-copy genes in organisms with flexible genomes.

Czech name

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Czech description

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Type

J_imp - Article in a specialist periodical, which is included in the Web of Science database

CEP classification

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OECD FORD branch

10608 - Biochemistry and molecular biology

Project

Result was created during the realization of more than one project. More information in the Projects tab.

Continuities

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Publication year

2018

Confidentiality

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Name of the periodical

PLoS One

ISSN

1932-6203

e-ISSN

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Volume of the periodical

13

Issue of the periodical within the volume

2

Country of publishing house

US - UNITED STATES

Number of pages

17

Pages from-to

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UT code for WoS article

000425083400048

EID of the result in the Scopus database

2-s2.0-85042073182