Cargando…

Cyclin-Dependent Kinase CRK9, Required for Spliced Leader trans Splicing of Pre-mRNA in Trypanosomes, Functions in a Complex with a New L-Type Cyclin and a Kinetoplastid-Specific Protein

In eukaryotes, cyclin-dependent kinases (CDKs) control the cell cycle and critical steps in gene expression. The lethal parasite Trypanosoma brucei, member of the phylogenetic order Kinetoplastida, possesses eleven CDKs which, due to high sequence divergence, were generically termed CDC2-related kin...

Descripción completa

Detalles Bibliográficos
Autores principales: Badjatia, Nitika, Park, Sung Hee, Ambrósio, Daniela L., Kirkham, Justin K., Günzl, Arthur
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4783070/
https://www.ncbi.nlm.nih.gov/pubmed/26954683
http://dx.doi.org/10.1371/journal.ppat.1005498
_version_ 1782420065603289088
author Badjatia, Nitika
Park, Sung Hee
Ambrósio, Daniela L.
Kirkham, Justin K.
Günzl, Arthur
author_facet Badjatia, Nitika
Park, Sung Hee
Ambrósio, Daniela L.
Kirkham, Justin K.
Günzl, Arthur
author_sort Badjatia, Nitika
collection PubMed
description In eukaryotes, cyclin-dependent kinases (CDKs) control the cell cycle and critical steps in gene expression. The lethal parasite Trypanosoma brucei, member of the phylogenetic order Kinetoplastida, possesses eleven CDKs which, due to high sequence divergence, were generically termed CDC2-related kinases (CRKs). While several CRKs have been implied in the cell cycle, CRK9 was the first trypanosome CDK shown to control the unusual mode of gene expression found in kinetoplastids. In these organisms, protein-coding genes are arranged in tandem arrays which are transcribed polycistronically. Individual mRNAs are processed from precursor RNA by spliced leader (SL) trans splicing and polyadenylation. CRK9 ablation was lethal in cultured trypanosomes, causing a block of trans splicing before the first transesterification step. Additionally, CRK9 silencing led to dephosphorylation of RNA polymerase II and to hypomethylation of the SL cap structure. Here, we tandem affinity-purified CRK9 and, among potential CRK9 substrates and modifying enzymes, discovered an unusual tripartite complex comprising CRK9, a new L-type cyclin (CYC12) and a protein, termed CRK9-associated protein (CRK9AP), that is only conserved among kinetoplastids. Silencing of either CYC12 or CRK9AP reproduced the effects of depleting CRK9, identifying these proteins as functional partners of CRK9 in vivo. While mammalian cyclin L binds to CDK11, the CRK9 complex deviates substantially from that of CDK11, requiring CRK9AP for efficient CRK9 complex formation and autophosphorylation in vitro. Interference with this unusual CDK rescued mice from lethal trypanosome infections, validating CRK9 as a potential chemotherapeutic target.
format Online
Article
Text
id pubmed-4783070
institution National Center for Biotechnology Information
language English
publishDate 2016
publisher Public Library of Science
record_format MEDLINE/PubMed
spelling pubmed-47830702016-03-23 Cyclin-Dependent Kinase CRK9, Required for Spliced Leader trans Splicing of Pre-mRNA in Trypanosomes, Functions in a Complex with a New L-Type Cyclin and a Kinetoplastid-Specific Protein Badjatia, Nitika Park, Sung Hee Ambrósio, Daniela L. Kirkham, Justin K. Günzl, Arthur PLoS Pathog Research Article In eukaryotes, cyclin-dependent kinases (CDKs) control the cell cycle and critical steps in gene expression. The lethal parasite Trypanosoma brucei, member of the phylogenetic order Kinetoplastida, possesses eleven CDKs which, due to high sequence divergence, were generically termed CDC2-related kinases (CRKs). While several CRKs have been implied in the cell cycle, CRK9 was the first trypanosome CDK shown to control the unusual mode of gene expression found in kinetoplastids. In these organisms, protein-coding genes are arranged in tandem arrays which are transcribed polycistronically. Individual mRNAs are processed from precursor RNA by spliced leader (SL) trans splicing and polyadenylation. CRK9 ablation was lethal in cultured trypanosomes, causing a block of trans splicing before the first transesterification step. Additionally, CRK9 silencing led to dephosphorylation of RNA polymerase II and to hypomethylation of the SL cap structure. Here, we tandem affinity-purified CRK9 and, among potential CRK9 substrates and modifying enzymes, discovered an unusual tripartite complex comprising CRK9, a new L-type cyclin (CYC12) and a protein, termed CRK9-associated protein (CRK9AP), that is only conserved among kinetoplastids. Silencing of either CYC12 or CRK9AP reproduced the effects of depleting CRK9, identifying these proteins as functional partners of CRK9 in vivo. While mammalian cyclin L binds to CDK11, the CRK9 complex deviates substantially from that of CDK11, requiring CRK9AP for efficient CRK9 complex formation and autophosphorylation in vitro. Interference with this unusual CDK rescued mice from lethal trypanosome infections, validating CRK9 as a potential chemotherapeutic target. Public Library of Science 2016-03-08 /pmc/articles/PMC4783070/ /pubmed/26954683 http://dx.doi.org/10.1371/journal.ppat.1005498 Text en © 2016 Badjatia et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Badjatia, Nitika
Park, Sung Hee
Ambrósio, Daniela L.
Kirkham, Justin K.
Günzl, Arthur
Cyclin-Dependent Kinase CRK9, Required for Spliced Leader trans Splicing of Pre-mRNA in Trypanosomes, Functions in a Complex with a New L-Type Cyclin and a Kinetoplastid-Specific Protein
title Cyclin-Dependent Kinase CRK9, Required for Spliced Leader trans Splicing of Pre-mRNA in Trypanosomes, Functions in a Complex with a New L-Type Cyclin and a Kinetoplastid-Specific Protein
title_full Cyclin-Dependent Kinase CRK9, Required for Spliced Leader trans Splicing of Pre-mRNA in Trypanosomes, Functions in a Complex with a New L-Type Cyclin and a Kinetoplastid-Specific Protein
title_fullStr Cyclin-Dependent Kinase CRK9, Required for Spliced Leader trans Splicing of Pre-mRNA in Trypanosomes, Functions in a Complex with a New L-Type Cyclin and a Kinetoplastid-Specific Protein
title_full_unstemmed Cyclin-Dependent Kinase CRK9, Required for Spliced Leader trans Splicing of Pre-mRNA in Trypanosomes, Functions in a Complex with a New L-Type Cyclin and a Kinetoplastid-Specific Protein
title_short Cyclin-Dependent Kinase CRK9, Required for Spliced Leader trans Splicing of Pre-mRNA in Trypanosomes, Functions in a Complex with a New L-Type Cyclin and a Kinetoplastid-Specific Protein
title_sort cyclin-dependent kinase crk9, required for spliced leader trans splicing of pre-mrna in trypanosomes, functions in a complex with a new l-type cyclin and a kinetoplastid-specific protein
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4783070/
https://www.ncbi.nlm.nih.gov/pubmed/26954683
http://dx.doi.org/10.1371/journal.ppat.1005498
work_keys_str_mv AT badjatianitika cyclindependentkinasecrk9requiredforsplicedleadertranssplicingofpremrnaintrypanosomesfunctionsinacomplexwithanewltypecyclinandakinetoplastidspecificprotein
AT parksunghee cyclindependentkinasecrk9requiredforsplicedleadertranssplicingofpremrnaintrypanosomesfunctionsinacomplexwithanewltypecyclinandakinetoplastidspecificprotein
AT ambrosiodanielal cyclindependentkinasecrk9requiredforsplicedleadertranssplicingofpremrnaintrypanosomesfunctionsinacomplexwithanewltypecyclinandakinetoplastidspecificprotein
AT kirkhamjustink cyclindependentkinasecrk9requiredforsplicedleadertranssplicingofpremrnaintrypanosomesfunctionsinacomplexwithanewltypecyclinandakinetoplastidspecificprotein
AT gunzlarthur cyclindependentkinasecrk9requiredforsplicedleadertranssplicingofpremrnaintrypanosomesfunctionsinacomplexwithanewltypecyclinandakinetoplastidspecificprotein