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Novel kinetoplastid-specific cAMP binding proteins identified by RNAi screening for cAMP resistance in Trypanosoma brucei

Cyclic AMP signalling in trypanosomes differs from most eukaryotes due to absence of known cAMP effectors and cAMP independence of PKA. We have previously identified four genes from a genome-wide RNAi screen for resistance to the cAMP phosphodiesterase (PDE) inhibitor NPD-001. The genes were named c...

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Autores principales: Bachmaier, Sabine, Gould, Matthew K., Polatoglou, Eleni, Omelianczyk, Radoslaw, Brennand, Ana E., Aloraini, Maha A., Munday, Jane C., Horn, David, Boshart, Michael, de Koning, Harry P.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10354285/
https://www.ncbi.nlm.nih.gov/pubmed/37475965
http://dx.doi.org/10.3389/fcimb.2023.1204707
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author Bachmaier, Sabine
Gould, Matthew K.
Polatoglou, Eleni
Omelianczyk, Radoslaw
Brennand, Ana E.
Aloraini, Maha A.
Munday, Jane C.
Horn, David
Boshart, Michael
de Koning, Harry P.
author_facet Bachmaier, Sabine
Gould, Matthew K.
Polatoglou, Eleni
Omelianczyk, Radoslaw
Brennand, Ana E.
Aloraini, Maha A.
Munday, Jane C.
Horn, David
Boshart, Michael
de Koning, Harry P.
author_sort Bachmaier, Sabine
collection PubMed
description Cyclic AMP signalling in trypanosomes differs from most eukaryotes due to absence of known cAMP effectors and cAMP independence of PKA. We have previously identified four genes from a genome-wide RNAi screen for resistance to the cAMP phosphodiesterase (PDE) inhibitor NPD-001. The genes were named cAMP Response Protein (CARP) 1 through 4. Here, we report an additional six CARP candidate genes from the original sample, after deep sequencing of the RNA interference target pool retrieved after NPD-001 selection (RIT-seq). The resistance phenotypes were confirmed by individual RNAi knockdown. Highest level of resistance to NPD-001, approximately 17-fold, was seen for knockdown of CARP7 (Tb927.7.4510). CARP1 and CARP11 contain predicted cyclic AMP binding domains and bind cAMP as evidenced by capture and competition on immobilised cAMP. CARP orthologues are strongly enriched in kinetoplastid species, and CARP3 and CARP11 are unique to Trypanosoma. Localization data and/or domain architecture of all CARPs predict association with the T. brucei flagellum. This suggests a crucial role of cAMP in flagellar function, in line with the cell division phenotype caused by high cAMP and the known role of the flagellum for cytokinesis. The CARP collection is a resource for discovery of unusual cAMP pathways and flagellar biology.
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spelling pubmed-103542852023-07-20 Novel kinetoplastid-specific cAMP binding proteins identified by RNAi screening for cAMP resistance in Trypanosoma brucei Bachmaier, Sabine Gould, Matthew K. Polatoglou, Eleni Omelianczyk, Radoslaw Brennand, Ana E. Aloraini, Maha A. Munday, Jane C. Horn, David Boshart, Michael de Koning, Harry P. Front Cell Infect Microbiol Cellular and Infection Microbiology Cyclic AMP signalling in trypanosomes differs from most eukaryotes due to absence of known cAMP effectors and cAMP independence of PKA. We have previously identified four genes from a genome-wide RNAi screen for resistance to the cAMP phosphodiesterase (PDE) inhibitor NPD-001. The genes were named cAMP Response Protein (CARP) 1 through 4. Here, we report an additional six CARP candidate genes from the original sample, after deep sequencing of the RNA interference target pool retrieved after NPD-001 selection (RIT-seq). The resistance phenotypes were confirmed by individual RNAi knockdown. Highest level of resistance to NPD-001, approximately 17-fold, was seen for knockdown of CARP7 (Tb927.7.4510). CARP1 and CARP11 contain predicted cyclic AMP binding domains and bind cAMP as evidenced by capture and competition on immobilised cAMP. CARP orthologues are strongly enriched in kinetoplastid species, and CARP3 and CARP11 are unique to Trypanosoma. Localization data and/or domain architecture of all CARPs predict association with the T. brucei flagellum. This suggests a crucial role of cAMP in flagellar function, in line with the cell division phenotype caused by high cAMP and the known role of the flagellum for cytokinesis. The CARP collection is a resource for discovery of unusual cAMP pathways and flagellar biology. Frontiers Media S.A. 2023-07-05 /pmc/articles/PMC10354285/ /pubmed/37475965 http://dx.doi.org/10.3389/fcimb.2023.1204707 Text en Copyright © 2023 Bachmaier, Gould, Polatoglou, Omelianczyk, Brennand, Aloraini, Munday, Horn, Boshart and de Koning https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Cellular and Infection Microbiology
Bachmaier, Sabine
Gould, Matthew K.
Polatoglou, Eleni
Omelianczyk, Radoslaw
Brennand, Ana E.
Aloraini, Maha A.
Munday, Jane C.
Horn, David
Boshart, Michael
de Koning, Harry P.
Novel kinetoplastid-specific cAMP binding proteins identified by RNAi screening for cAMP resistance in Trypanosoma brucei
title Novel kinetoplastid-specific cAMP binding proteins identified by RNAi screening for cAMP resistance in Trypanosoma brucei
title_full Novel kinetoplastid-specific cAMP binding proteins identified by RNAi screening for cAMP resistance in Trypanosoma brucei
title_fullStr Novel kinetoplastid-specific cAMP binding proteins identified by RNAi screening for cAMP resistance in Trypanosoma brucei
title_full_unstemmed Novel kinetoplastid-specific cAMP binding proteins identified by RNAi screening for cAMP resistance in Trypanosoma brucei
title_short Novel kinetoplastid-specific cAMP binding proteins identified by RNAi screening for cAMP resistance in Trypanosoma brucei
title_sort novel kinetoplastid-specific camp binding proteins identified by rnai screening for camp resistance in trypanosoma brucei
topic Cellular and Infection Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10354285/
https://www.ncbi.nlm.nih.gov/pubmed/37475965
http://dx.doi.org/10.3389/fcimb.2023.1204707
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