Marta Vicente-Crespo

Publications

• 5.20

  Impact points

  Drosophila Upf1 and Upf2 loss of function inhibits cell growth and causes animal death in a Upf3-independent manner.

  Paul Avery, Marta Vicente-Crespo, Deepthy Francis, Oxana Nashchekina, Claudio R Alonso, Isabel M Palacios

  RNA (New York, N.Y.). 02/2011; 17(4):624-38.

  Nonsense-mediated RNA decay (NMD) is a surveillance mechanism that degrades transcripts containing nonsense mutations, preventing the translation of truncated proteins. NMD also regulates the levels of many endogenous mRNAs. While the mechanism of NMD is gradually understood, its physiological role ... [more] Nonsense-mediated RNA decay (NMD) is a surveillance mechanism that degrades transcripts containing nonsense mutations, preventing the translation of truncated proteins. NMD also regulates the levels of many endogenous mRNAs. While the mechanism of NMD is gradually understood, its physiological role remains largely unknown. The core NMD genes upf1 and upf2 are essential in several organisms, which may reflect an important developmental role for NMD. Alternatively, the lethality of these mutants might arise from their function in NMD-independent processes. To analyze the developmental importance of NMD, we studied Drosophila mutants of the other core NMD gene, upf3. We compare the resulting upf3 phenotype with those defects observed in upf1 and upf2 loss-of-function mutants, as well as with flies expressing a mutant Upf2 protein unable to bind Upf3. Our results show that Upf3 is an NMD effector in the fly but, unlike Upf1 and Upf2, plays a peripheral role in the degradation of most NMD targets and is not required for development or viability. Furthermore, Upf1 and Upf2 loss-of-function inhibits cell growth and induces apoptosis through a Upf3-independent pathway. Accordingly, disruption of Upf2-Upf1 interaction causes death, while the Upf2-Upf3 complex is dispensable for viability. Our findings suggest that NMD is essential for cell growth and animal development, and that the lethality of upf1 and upf2 mutants is not due to disrupting their roles during NMD-independent processes, but to their function in the degradation of specific mRNAs by the NMD pathway. Furthermore, our results show that Upf3 is not always essential in NMD.
• 3.38

  Impact points

  Nonsense-mediated mRNA decay and development: shoot the messenger to survive?

  Marta Vicente-Crespo, Isabel M Palacios

  Biochemical Society transactions. 12/2010; 38(6):1500-5.

  NMD (nonsense-mediated mRNA decay) is a surveillance mechanism that degrades transcripts containing nonsense mutations, preventing the translation of potentially harmful truncated proteins. Although the mechanistic details of NMD are gradually being understood, the physiological role of this RNA sur... [more] NMD (nonsense-mediated mRNA decay) is a surveillance mechanism that degrades transcripts containing nonsense mutations, preventing the translation of potentially harmful truncated proteins. Although the mechanistic details of NMD are gradually being understood, the physiological role of this RNA surveillance pathway still remains largely unknown. The core NMD genes Upf1 (up-frameshift suppressor 1) and Upf2 are essential for animal viability in the fruitfly, mouse and zebrafish. These findings may reflect an important role for NMD during animal development. Alternatively, the lethal phenotypes of upf1 and upf2 mutants might be due to their function in NMD-independent processes. In the present paper, we describe the phenotypes observed when the NMD factors are mutated in various organisms, and discuss findings that might shed light on the function of NMD in cellular growth and development of an organism.
• 4.41

  Impact points

  Genetic and chemical modifiers of a CUG toxicity model in Drosophila.

  Amparo Garcia-Lopez, Lidon Monferrer, Irma Garcia-Alcover, Marta Vicente-Crespo, M. Carmen Alvarez-Abril, Ruben D Artero

  PLoS ONE. 02/2008; 3(2):e1595.

  Non-coding CUG repeat expansions interfere with the activity of human Muscleblind-like (MBNL) proteins contributing to myotonic dystrophy 1 (DM1). To understand this toxic RNA gain-of-function mechanism we developed a Drosophila model expressing 60 pure and 480 interrupted CUG repeats in the context... [more] Non-coding CUG repeat expansions interfere with the activity of human Muscleblind-like (MBNL) proteins contributing to myotonic dystrophy 1 (DM1). To understand this toxic RNA gain-of-function mechanism we developed a Drosophila model expressing 60 pure and 480 interrupted CUG repeats in the context of a non-translatable RNA. These flies reproduced aspects of the DM1 pathology, most notably nuclear accumulation of CUG transcripts, muscle degeneration, splicing misregulation, and diminished Muscleblind function in vivo. Reduced Muscleblind activity was evident from the sensitivity of CUG-induced phenotypes to a decrease in muscleblind genetic dosage and rescue by MBNL1 expression, and further supported by the co-localization of Muscleblind and CUG repeat RNA in ribonuclear foci. Targeted expression of CUG repeats to the developing eye and brain mushroom bodies was toxic leading to rough eyes and semilethality, respectively. These phenotypes were utilized to identify genetic and chemical modifiers of the CUG-induced toxicity. 15 genetic modifiers of the rough eye phenotype were isolated. These genes identify putative cellular processes unknown to be altered by CUG repeat RNA, and they include mRNA export factor Aly, apoptosis inhibitor Thread, chromatin remodelling factor Nurf-38, and extracellular matrix structural component Viking. Ten chemical compounds suppressed the semilethal phenotype. These compounds significantly improved viability of CUG expressing flies and included non-steroidal anti-inflammatory agents (ketoprofen), muscarinic, cholinergic and histamine receptor inhibitors (orphenadrine), and drugs that can affect sodium and calcium metabolism such as clenbuterol and spironolactone. These findings provide new insights into the DM1 phenotype, and suggest novel candidates for DM1 treatments.
• 4.41

  Impact points

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

  Marta Vicente-Crespo, Maya Pascual, Juan M Fernandez-Costa, Amparo Garcia-Lopez, Lidón Monferrer, M. Eugenia Miranda, Lei Zhou, Ruben D Artero

  PLoS ONE. 02/2008; 3(2):e1613.

  BACKGROUND: 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-... [more] BACKGROUND: 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. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSIONS/SIGNIFICANCE: 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.
• 3.31

  Impact points

  Muscleblind isoforms are functionally distinct and regulate alpha-actinin splicing.

  Marta Vicente, Lidón Monferrer, Michael G Poulos, Jonathan Houseley, Darren G Monckton, Kevin M C O'Dell, Maurice S Swanson, Rubén D Artero

  Differentiation; research in biological diversity. 07/2007; 75(5):427-40.

  Drosophila Muscleblind (Mbl) proteins control terminal muscle and neural differentiation, but their molecular function has not been experimentally addressed. Such an analysis is relevant as the human Muscleblind-like homologs (MBNL1-3) are implicated in the pathogenesis of the inherited muscular dev... [more] Drosophila Muscleblind (Mbl) proteins control terminal muscle and neural differentiation, but their molecular function has not been experimentally addressed. Such an analysis is relevant as the human Muscleblind-like homologs (MBNL1-3) are implicated in the pathogenesis of the inherited muscular developmental and degenerative disease myotonic dystrophy. The Drosophila muscleblind gene expresses four protein coding splice forms (mblA to mblD) that are differentially expressed during the Drosophila life cycle, and which vary markedly in their ability to rescue the embryonic lethal phenotype of muscleblind mutant flies. Analysis of muscleblind mutant embryos reveals misregulated alternative splicing of the transcripts encoding Z-band component alpha-Actinin, which can be replicated in human cells expressing a Drosophilaalpha-actinin minigene and epitope-tagged Muscleblind isoforms. MblC appreciably altered alpha-actinin splicing in this assay, whereas other isoforms had only a marginal or no effect, demonstrating functional specialization among Muscleblind proteins. To further analyze the molecular basis of these differences, we studied the subcellular localization of Muscleblind isoforms. Consistent with the splicing assay results, MblB and MblC were enriched in the nucleus while MblA was predominantly cytoplasmic. In myotonic dystrophy, transcripts bearing expanded non-coding CUG or CCUG repeats interfere with the function of human MBNL proteins. Co-expression of CUG repeat RNA with the alpha-actinin minigene altered splicing compared with that seen in muscleblind mutant embryos, indicating that CUG repeat expansion RNA also interferes with Drosophila muscleblind function. Moreover MblA, B, and C co-localize with CUG repeat RNA in nuclear foci in cell culture. Our observations indicate that Muscleblind isoforms perform different functions in vivo, that MblC controls muscleblind-dependent alternative splicing events, and establish the functional conservation between Muscleblind and MBNL proteins both over a physiological target (alpha-actinin) and a pathogenic one (CUG repeats).
• 3.31

  Impact points

  The Muscleblind family of proteins: an emerging class of regulators of developmentally programmed alternative splicing.

  Maya Pascual, Marta Vicente, Lidon Monferrer, Ruben Artero

  Differentiation; research in biological diversity. 04/2006; 74(2-3):65-80.

  Alternative splicing is widely used to generate protein diversity and to control gene expression in many biological processes, including cell fate determination and apoptosis. In this review, we focus on the Muscleblind family of tissue-specific alternative splicing regulators. Muscleblind proteins ... [more] Alternative splicing is widely used to generate protein diversity and to control gene expression in many biological processes, including cell fate determination and apoptosis. In this review, we focus on the Muscleblind family of tissue-specific alternative splicing regulators. Muscleblind proteins bind pre-mRNA through an evolutionarily conserved tandem CCCH zinc finger domain. Human Muscleblind homologs MBNL1, MBNL2 and MBNL3 promote inclusion or exclusion of specific exons on different pre-mRNAs by antagonizing the activity of CUG-BP and ETR-3-like factors (CELF proteins) bound to distinct intronic sites. The relative activities of Muscleblind and CELF proteins control a key developmental switch. Defined transcripts follow an embryonic splice pattern when CELF activity predominates, whereas they follow an adult pattern when Muscleblind activity prevails. Human MBNL proteins show functional specializations. While MBNL1 seems to promote muscle differentiation, MBNL3 appears to function in an opposing manner inhibiting expression of muscle differentiation markers. MBNL2, on the other hand, participates in a new RNA-dependent protein localization mechanism involving recruitment of integrin alpha3 protein to focal adhesions. Both muscleblind mutant Drosophila embryos and Mbnl1 knockout mice show muscle abnormalities and altered splicing of specific transcripts. In addition to regulating terminal muscle differentiation through alternative splicing control, results by several groups suggest that Muscleblind participates in the differentiation of photoreceptors, neurons, adipocytes and blood cell types. Misregulation of MBNL activity can lead to human pathologies. Through mechanisms not completely identified yet, expression of transcripts containing large non-coding CUG or CCUG repeat expansions mimics muscleblind loss-of-function phenotypes. Archetypical within this class of disorders are myotonic dystrophies. Our understanding of the biology of Muscleblind proteins has increased dramatically over the last few years, but several key issues remain unsolved. Defining the mechanism of the activity of Muscleblind proteins, their splicing partners, and the functional relevance of its several protein isoforms are just a few examples.