Genetic dissection of a Leishmania flagellar proteome demonstrates requirement for directional motility in sand fly infections
The protozoan parasite Leishmania possesses a single flagellum, which is remodelled during the parasite's life cycle from a long motile flagellum in promastigote forms in the sand fly to a short immotile flagellum in amastigotes residing in mammalian phagocytes. This study examined the protein...
Saved in:
Published in | bioRxiv |
---|---|
Main Authors | , , , , , , , , , , , , , , |
Format | Paper |
Language | English |
Published |
Cold Spring Harbor
Cold Spring Harbor Laboratory Press
27.11.2018
|
Subjects | |
Online Access | Get full text |
Cover
Loading…
Abstract | The protozoan parasite Leishmania possesses a single flagellum, which is remodelled during the parasite's life cycle from a long motile flagellum in promastigote forms in the sand fly to a short immotile flagellum in amastigotes residing in mammalian phagocytes. This study examined the protein composition and in vivo function of the promastigote flagellum. Protein mass spectrometry and label free protein enrichment testing of isolated flagella and deflagellated cell bodies defined a flagellar proteome for L. mexicana promastigote forms (available via ProteomeXchange with identifier PXD011057). This information was used to generate a CRISPR-Cas9 knockout library of 100 mutants to screen for flagellar defects. This first large-scale knockout screen in a Leishmania sp. identified 56 mutants with altered swimming speed (52 reduced and 4 increased) and defined distinct mutant categories (faster swimmers, slower swimmers, slow uncoordinated swimmers and paralysed cells, including aflagellate promastigotes and cells with curled flagella and disruptions of the paraflagellar rod). Each mutant was tagged with a unique 17-nt barcode, providing a simple barcode sequencing (bar-seq) method for measuring the relative fitness of L. mexicana mutants in vivo . In mixed infections of the permissive sand fly vector Lutzomyia longipalpis , paralysed promastigotes and uncoordinated swimmers were severely diminished in the fly after defecation of the bloodmeal. Subsequent examination of flies infected with a single mutant lacking the central pair protein PF16 showed that these paralysed promastigotes did not reach anterior regions of the fly alimentary tract. These data show that L. mexicana need directional motility for successful colonisation of sand flies. |
---|---|
AbstractList | The protozoan parasite Leishmania possesses a single flagellum, which is remodelled during the parasite's life cycle from a long motile flagellum in promastigote forms in the sand fly to a short immotile flagellum in amastigotes residing in mammalian phagocytes. This study examined the protein composition and in vivo function of the promastigote flagellum. Protein mass spectrometry and label free protein enrichment testing of isolated flagella and deflagellated cell bodies defined a flagellar proteome for L. mexicana promastigote forms (available via ProteomeXchange with identifier PXD011057). This information was used to generate a CRISPR-Cas9 knockout library of 100 mutants to screen for flagellar defects. This first large-scale knockout screen in a Leishmania sp. identified 56 mutants with altered swimming speed (52 reduced and 4 increased) and defined distinct mutant categories (faster swimmers, slower swimmers, slow uncoordinated swimmers and paralysed cells, including aflagellate promastigotes and cells with curled flagella and disruptions of the paraflagellar rod). Each mutant was tagged with a unique 17-nt barcode, providing a simple barcode sequencing (bar-seq) method for measuring the relative fitness of L. mexicana mutants in vivo . In mixed infections of the permissive sand fly vector Lutzomyia longipalpis , paralysed promastigotes and uncoordinated swimmers were severely diminished in the fly after defecation of the bloodmeal. Subsequent examination of flies infected with a single mutant lacking the central pair protein PF16 showed that these paralysed promastigotes did not reach anterior regions of the fly alimentary tract. These data show that L. mexicana need directional motility for successful colonisation of sand flies. |
Author | Jeffery, Heather Demay, Francois Gluenz, Eva Hookway, Edward Ashman, Nicole Wheeler, Richard J Sadlova, Jovana Volf, Petr Smith, James Myskova, Jitka Beneke, Tom Valli, Jessica Becvar, Tomas Lestinova, Tereza Shahaan Shafiq |
Author_xml | – sequence: 1 givenname: Tom surname: Beneke fullname: Beneke, Tom – sequence: 2 givenname: Francois surname: Demay fullname: Demay, Francois – sequence: 3 givenname: Edward surname: Hookway fullname: Hookway, Edward – sequence: 4 givenname: Nicole surname: Ashman fullname: Ashman, Nicole – sequence: 5 givenname: Heather surname: Jeffery fullname: Jeffery, Heather – sequence: 6 givenname: James surname: Smith fullname: Smith, James – sequence: 7 givenname: Jessica surname: Valli fullname: Valli, Jessica – sequence: 8 givenname: Tomas surname: Becvar fullname: Becvar, Tomas – sequence: 9 givenname: Jitka surname: Myskova fullname: Myskova, Jitka – sequence: 10 givenname: Tereza surname: Lestinova fullname: Lestinova, Tereza – sequence: 11 fullname: Shahaan Shafiq – sequence: 12 givenname: Jovana surname: Sadlova fullname: Sadlova, Jovana – sequence: 13 givenname: Petr surname: Volf fullname: Volf, Petr – sequence: 14 givenname: Richard surname: Wheeler middlename: J fullname: Wheeler, Richard J – sequence: 15 givenname: Eva surname: Gluenz fullname: Gluenz, Eva |
BookMark | eNotjzFPwzAUhD3AAAV-w5OYA3aeqeMRVVCQInWBuXKcZzBy7NZ2BhZ-O0FlutPp9Onukp3FFImxG8HvhODiXqq11vKC_WwpUvUWRl8K2epThOTAQE--fE4megMumA8KwWQ45FQpTQQjTSmWmk2lApmOs880UazgUl5Q-UQyAaZUffD1G3yEYuK4wP68OxXKFTt3JhS6_tcVe39-etu8NP1u-7p57JuDQC4bFCM32GqLiqyTzrpOOzVgazvbkXIC9aiNUh1xbSWhde1gaAkk0qAfNK7Y7Ym7PDjOVOr-K815GVj2rUC1xo63HH8B8-ZdSA |
ContentType | Paper |
Copyright | 2018. This article is published under http://creativecommons.org/licenses/by/4.0/ (“the License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License. |
Copyright_xml | – notice: 2018. This article is published under http://creativecommons.org/licenses/by/4.0/ (“the License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License. |
DBID | 8FE 8FH AAFGM AAMXL ABOIG ABUWG ADZZV AFKRA AFLLJ AFOLM AGAJT AQTIP AZQEC BBNVY BENPR BHPHI CCPQU DWQXO GNUQQ HCIFZ LK8 M7P PIMPY PQCXX PQEST PQQKQ PQUKI PRINS |
DOI | 10.1101/476994 |
DatabaseName | ProQuest SciTech Collection ProQuest Natural Science Collection ProQuest Central Korea - hybrid linking Natural Science Collection - hybrid linking Biological Science Collection - hybrid linking ProQuest Central (Alumni) ProQuest Central (Alumni) - hybrid linking ProQuest Central SciTech Premium Collection - hybrid linking ProQuest Central Student - hybrid linking ProQuest Central Essentials - hybrid linking ProQuest Women's & Gender Studies - hybrid linking ProQuest Central Essentials Biological Science Collection ProQuest Central Natural Science Collection ProQuest One Community College ProQuest Central Korea ProQuest Central Student SciTech Premium Collection Biological Sciences Biological Science Database Publicly Available Content Database ProQuest Central - hybrid linking ProQuest One Academic Eastern Edition (DO NOT USE) ProQuest One Academic ProQuest One Academic UKI Edition ProQuest Central China |
DatabaseTitle | Publicly Available Content Database ProQuest Central Student ProQuest Biological Science Collection ProQuest Central Essentials ProQuest One Academic Eastern Edition ProQuest Central (Alumni Edition) SciTech Premium Collection ProQuest One Community College ProQuest Natural Science Collection Biological Science Database ProQuest SciTech Collection ProQuest Central China ProQuest Central ProQuest One Academic UKI Edition Natural Science Collection ProQuest Central Korea Biological Science Collection ProQuest One Academic |
DatabaseTitleList | Publicly Available Content Database |
Database_xml | – sequence: 1 dbid: BENPR name: ProQuest Central url: https://www.proquest.com/central sourceTypes: Aggregation Database |
DeliveryMethod | fulltext_linktorsrc |
Genre | Working Paper/Pre-Print |
GroupedDBID | 8FE 8FH ABUWG AFKRA AZQEC BBNVY BENPR BHPHI CCPQU DWQXO GNUQQ HCIFZ LK8 M7P PIMPY PQEST PQQKQ PQUKI PRINS |
ID | FETCH-LOGICAL-p1304-31d0a329c37ecf4fcf89f7b32c8c8e7f139d9a778e09c4e3cf2baea7743eb9593 |
IEDL.DBID | BENPR |
IngestDate | Thu Oct 17 04:18:33 EDT 2024 |
IsDoiOpenAccess | true |
IsOpenAccess | true |
IsPeerReviewed | false |
IsScholarly | false |
Language | English |
LinkModel | DirectLink |
MergedId | FETCHMERGED-LOGICAL-p1304-31d0a329c37ecf4fcf89f7b32c8c8e7f139d9a778e09c4e3cf2baea7743eb9593 |
OpenAccessLink | https://www.proquest.com/docview/2137638020?pq-origsite=%requestingapplication% |
PQID | 2137638020 |
PQPubID | 2050091 |
ParticipantIDs | proquest_journals_2137638020 |
PublicationCentury | 2000 |
PublicationDate | 20181127 |
PublicationDateYYYYMMDD | 2018-11-27 |
PublicationDate_xml | – month: 11 year: 2018 text: 20181127 day: 27 |
PublicationDecade | 2010 |
PublicationPlace | Cold Spring Harbor |
PublicationPlace_xml | – name: Cold Spring Harbor |
PublicationTitle | bioRxiv |
PublicationYear | 2018 |
Publisher | Cold Spring Harbor Laboratory Press |
Publisher_xml | – name: Cold Spring Harbor Laboratory Press |
Score | 1.5887796 |
Snippet | The protozoan parasite Leishmania possesses a single flagellum, which is remodelled during the parasite's life cycle from a long motile flagellum in... |
SourceID | proquest |
SourceType | Aggregation Database |
SubjectTerms | Amastigotes CRISPR Defecation Flagella Gastrointestinal tract Leishmania Life cycles Mass spectroscopy Motility Phagocytes Promastigotes Protein composition Proteins Proteomes Protozoa Reproductive fitness Swimming |
Title | Genetic dissection of a Leishmania flagellar proteome demonstrates requirement for directional motility in sand fly infections |
URI | https://www.proquest.com/docview/2137638020 |
hasFullText | 1 |
inHoldings | 1 |
isFullTextHit | |
isPrint | |
link | http://utb.summon.serialssolutions.com/2.0.0/link/0/eLvHCXMwfV1NSwMxEA3aXrwpKn5UycHr0m2S3SQnQWkpoqWIhd5KNjvBgt2t3Xr1tzuTbhE8eAsbyCGTefN2MnnD2J10QWYuEwkI4RJlU48ulRZJagX-cOUBQpQUepnk45l6mmfzNuHWtGWVe0yMQF3WnnLkfTEgVzDIbu7Xnwl1jaLb1baFxiHr4rwxHdZ9GE6mr20TITxufaVzSx2J_yBtDB-jY9adujVsTtgBVKfsm6Se0V483oXHdwW8DtzxZ1g276RH4Xj4QD9HC214FFKoV8BLWBGXI2mHhm-AKnhjao8j7eS7yBTTepzK64hc82XFG1eVuBiNd0VXVXPGZqPh2-M4adsgJGsMMApRskydFNZLDT6o4IOxQRdSeOMN6IAcrrROawOp9QqkD6JwgB-UhIJ0h89Zp6oruGBcIJ3TFvIBKKl8gQtkNisQexWOAbJL1ttv06I9y83id-ev_p--ZkdIJwy91BO6xzrbzRfcYMjeFretXX4AkaGeLg |
link.rule.ids | 780,784,21388,27925,33744,43805 |
linkProvider | ProQuest |
linkToHtml | http://utb.summon.serialssolutions.com/2.0.0/link/0/eLvHCXMwfV09T8MwELWgHWADAeKjgAfWqKnjxPGEBGpVoK0q1ErdKsc5i0qQlKSs_Hbu3FRIDGyWI3nw-d69nM_vGLuLjItiE4sAhDCB1KFFlwqzINQCf7gSB85LCo0nyXAunxfxokm41U1Z5Q4TPVDnpaUceVf0yBVSZDf368-AukbR7WrTQmOftUk5PW6x9kN_Mn1tmgjhcetKlWjqSPwHaX34GByx9tSsoTpme1CcsG-SekZ7cX8X7t8V8NJxw0ewqt9Ij8Jw945-jhaquBdSKD-A5_BBXI6kHWpeAVXw-tQeR9rJt5HJp_U4ldcRueargtemyHExGm-Lror6lM0H_dnjMGjaIARrDDASUTIPTSS0jRRYJ511qXYqi4RNbQrKIYfLtVEqhVBbCZF1IjOAEzKCjHSHz1irKAs4Z1wgnVMakh7ISNoMF4h1nCH2ShwDxBess9umZXOW6-Xvzl_-__mWHQxn49Fy9DR5uWKHSC1SerUnVIe1NtUXXGP43mQ3jY1-ABf5oRY |
openUrl | ctx_ver=Z39.88-2004&ctx_enc=info%3Aofi%2Fenc%3AUTF-8&rfr_id=info%3Asid%2Fsummon.serialssolutions.com&rft_val_fmt=info%3Aofi%2Ffmt%3Akev%3Amtx%3Ajournal&rft.genre=article&rft.atitle=Genetic+dissection+of+a+Leishmania+flagellar+proteome+demonstrates+requirement+for+directional+motility+in+sand+fly+infections&rft.jtitle=bioRxiv&rft.au=Beneke%2C+Tom&rft.au=Demay%2C+Francois&rft.au=Hookway%2C+Edward&rft.au=Ashman%2C+Nicole&rft.date=2018-11-27&rft.pub=Cold+Spring+Harbor+Laboratory+Press&rft_id=info:doi/10.1101%2F476994 |