Drosophila muscleblind is involved in troponin T alternative splicing and apoptosis.

<h4>Background</h4>Muscleblind-like proteins (MBNL) have been involved in a developmental switch in the use of defined cassette exons. Such transition fails in the CTG repeat expansion disease myotonic dystrophy due, in part, to sequestration of MBNL proteins by CUG repeat RNA. Four prot...

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Autores principales:	Marta Vicente-Crespo, Maya Pascual, Juan M Fernandez-Costa, Amparo Garcia-Lopez, Lidón Monferrer, M Eugenia Miranda, Lei Zhou, Ruben D Artero
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2008
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/2884507188d24920a3f657a3b0c91edf
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spelling	oai:doaj.org-article:2884507188d24920a3f657a3b0c91edf2021-11-25T06:13:22ZDrosophila muscleblind is involved in troponin T alternative splicing and apoptosis.1932-620310.1371/journal.pone.0001613https://doaj.org/article/2884507188d24920a3f657a3b0c91edf2008-02-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/18286170/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203<h4>Background</h4>Muscleblind-like proteins (MBNL) have been involved in a developmental switch in the use of defined cassette exons. Such transition fails in the CTG repeat expansion disease myotonic dystrophy due, in part, to sequestration of MBNL proteins by CUG repeat RNA. Four protein isoforms (MblA-D) are coded by the unique Drosophila muscleblind gene.<h4>Methodology/principal findings</h4>We used evolutionary, genetic and cell culture approaches to study muscleblind (mbl) function in flies. The evolutionary study showed that the MblC protein isoform was readily conserved from nematods to Drosophila, which suggests that it performs the most ancestral muscleblind functions. Overexpression of MblC in the fly eye precursors led to an externally rough eye morphology. This phenotype was used in a genetic screen to identify five dominant suppressors and 13 dominant enhancers including Drosophila CUG-BP1 homolog aret, exon junction complex components tsunagi and Aly, and pro-apoptotic genes Traf1 and reaper. We further investigated Muscleblind implication in apoptosis and splicing regulation. We found missplicing of troponin T in muscleblind mutant pupae and confirmed Muscleblind ability to regulate mouse fast skeletal muscle Troponin T (TnnT3) minigene splicing in human HEK cells. MblC overexpression in the wing imaginal disc activated apoptosis in a spatially restricted manner. Bioinformatics analysis identified a conserved FKRP motif, weakly resembling a sumoylation target site, in the MblC-specific sequence. Site-directed mutagenesis of the motif revealed no change in activity of mutant MblC on TnnT3 minigene splicing or aberrant binding to CUG repeat RNA, but altered the ability of the protein to form perinuclear aggregates and enhanced cell death-inducing activity of MblC overexpression.<h4>Conclusions/significance</h4>Taken together our genetic approach identify cellular processes influenced by Muscleblind function, whereas in vivo and cell culture experiments define Drosophila troponin T as a new Muscleblind target, reveal a potential involvement of MblC in programmed cell death and recognize the FKRP motif as a putative regulator of MblC function and/or subcellular location in the cell.Marta Vicente-CrespoMaya PascualJuan M Fernandez-CostaAmparo Garcia-LopezLidón MonferrerM Eugenia MirandaLei ZhouRuben D ArteroPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 3, Iss 2, p e1613 (2008)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Marta Vicente-Crespo Maya Pascual Juan M Fernandez-Costa Amparo Garcia-Lopez Lidón Monferrer M Eugenia Miranda Lei Zhou Ruben D Artero Drosophila muscleblind is involved in troponin T alternative splicing and apoptosis.
description	<h4>Background</h4>Muscleblind-like proteins (MBNL) have been involved in a developmental switch in the use of defined cassette exons. Such transition fails in the CTG repeat expansion disease myotonic dystrophy due, in part, to sequestration of MBNL proteins by CUG repeat RNA. Four protein isoforms (MblA-D) are coded by the unique Drosophila muscleblind gene.<h4>Methodology/principal findings</h4>We used evolutionary, genetic and cell culture approaches to study muscleblind (mbl) function in flies. The evolutionary study showed that the MblC protein isoform was readily conserved from nematods to Drosophila, which suggests that it performs the most ancestral muscleblind functions. Overexpression of MblC in the fly eye precursors led to an externally rough eye morphology. This phenotype was used in a genetic screen to identify five dominant suppressors and 13 dominant enhancers including Drosophila CUG-BP1 homolog aret, exon junction complex components tsunagi and Aly, and pro-apoptotic genes Traf1 and reaper. We further investigated Muscleblind implication in apoptosis and splicing regulation. We found missplicing of troponin T in muscleblind mutant pupae and confirmed Muscleblind ability to regulate mouse fast skeletal muscle Troponin T (TnnT3) minigene splicing in human HEK cells. MblC overexpression in the wing imaginal disc activated apoptosis in a spatially restricted manner. Bioinformatics analysis identified a conserved FKRP motif, weakly resembling a sumoylation target site, in the MblC-specific sequence. Site-directed mutagenesis of the motif revealed no change in activity of mutant MblC on TnnT3 minigene splicing or aberrant binding to CUG repeat RNA, but altered the ability of the protein to form perinuclear aggregates and enhanced cell death-inducing activity of MblC overexpression.<h4>Conclusions/significance</h4>Taken together our genetic approach identify cellular processes influenced by Muscleblind function, whereas in vivo and cell culture experiments define Drosophila troponin T as a new Muscleblind target, reveal a potential involvement of MblC in programmed cell death and recognize the FKRP motif as a putative regulator of MblC function and/or subcellular location in the cell.
format	article
author	Marta Vicente-Crespo Maya Pascual Juan M Fernandez-Costa Amparo Garcia-Lopez Lidón Monferrer M Eugenia Miranda Lei Zhou Ruben D Artero
author_facet	Marta Vicente-Crespo Maya Pascual Juan M Fernandez-Costa Amparo Garcia-Lopez Lidón Monferrer M Eugenia Miranda Lei Zhou Ruben D Artero
author_sort	Marta Vicente-Crespo
title	Drosophila muscleblind is involved in troponin T alternative splicing and apoptosis.
title_short	Drosophila muscleblind is involved in troponin T alternative splicing and apoptosis.
title_full	Drosophila muscleblind is involved in troponin T alternative splicing and apoptosis.
title_fullStr	Drosophila muscleblind is involved in troponin T alternative splicing and apoptosis.
title_full_unstemmed	Drosophila muscleblind is involved in troponin T alternative splicing and apoptosis.
title_sort	drosophila muscleblind is involved in troponin t alternative splicing and apoptosis.
publisher	Public Library of Science (PLoS)
publishDate	2008
url	https://doaj.org/article/2884507188d24920a3f657a3b0c91edf
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Drosophila muscleblind is involved in troponin T alternative splicing and apoptosis.

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