Wong CH, Nguyen L, Peh J, Luu LM, Sanchez JS, Richardson SL, Tuccinardi T, Tsoi H, Chan WY, Chan HY, Baranger AM, Hergenrother PJ, Zimmerman SC. Targeting toxic RNAs that cause myotonic dystrophy type 1 (DM1) with a bisamidinium inhibitor. J Am Chem Soc. 2014 Apr 30;136(17):6355-61. PMID: 24702247; PMCID: PMC4015652.
From the abstract: "A working hypothesis for the pathogenesis of myotonic dystrophy type 1 (DM1) involves the aberrant sequestration of an alternative splicing regulator, MBNL1, by expanded CUG repeats, r(CUG)(exp). It has been suggested that a reversal of the myotonia and potentially other symptoms of the DM1 disease can be achieved by inhibiting the toxic MBNL1-r(CUG)(exp) interaction. Using rational design, we discovered an RNA-groove binding inhibitor (ligand 3) that contains two triaminotriazine units connected by a bisamidinium linker. ... Importantly, suppression of r(CUG)(exp) RNA-induced toxicity in a DM1 Drosophila model was observed after treatment with ligand 3. These results suggest ligand 3 as a lead for the treatment of DM1."
Monday, December 12, 2016
Using the fly to investigate what links xeroderma pigmentosum to cancer risk
Stettler K, Li X, Sandrock B, Braga-Lagache S, Heller M, Dümbgen L, Suter B. A Drosophila XPD model links cell cycle coordination with neuro-development and suggests links to cancer. Dis Model Mech. 2015 Jan;8(1):81-91. PMID: 25431422; PMCID: PMC4283652.
From the abstract: "XPD functions in transcription, DNA repair and in cell cycle control. Mutations in human XPD (also known as ERCC2) mainly cause three clinical phenotypes: xeroderma pigmentosum (XP), Cockayne syndrome (XP/CS) and trichothiodystrophy (TTD), and only XP patients have a high predisposition to developing cancer. Hence, we developed a fly model to obtain novel insights into the defects caused by individual hypomorphic alleles identified in human XP-D patients. This model revealed that the mutations that displayed the greatest in vivo UV sensitivity in Drosophila did not correlate with those that led to tumor formation in humans. Immunoprecipitations followed by targeted quantitative MS/MS analysis showed how different xpd mutations affected the formation or stability of different transcription factor IIH (TFIIH) subcomplexes. The XP mutants most clearly linked to high cancer risk, Xpd R683W and R601L, showed a reduced interaction with the core TFIIH and also an abnormal interaction with the Cdk-activating kinase (CAK) complex. ..."
From the abstract: "XPD functions in transcription, DNA repair and in cell cycle control. Mutations in human XPD (also known as ERCC2) mainly cause three clinical phenotypes: xeroderma pigmentosum (XP), Cockayne syndrome (XP/CS) and trichothiodystrophy (TTD), and only XP patients have a high predisposition to developing cancer. Hence, we developed a fly model to obtain novel insights into the defects caused by individual hypomorphic alleles identified in human XP-D patients. This model revealed that the mutations that displayed the greatest in vivo UV sensitivity in Drosophila did not correlate with those that led to tumor formation in humans. Immunoprecipitations followed by targeted quantitative MS/MS analysis showed how different xpd mutations affected the formation or stability of different transcription factor IIH (TFIIH) subcomplexes. The XP mutants most clearly linked to high cancer risk, Xpd R683W and R601L, showed a reduced interaction with the core TFIIH and also an abnormal interaction with the Cdk-activating kinase (CAK) complex. ..."
New fly model of myotonic dystrophy type 2
Yu Z, Goodman LD, Shieh SY, Min M, Teng X, Zhu Y, Bonini NM. A fly model for the CCUG-repeat expansion of myotonic dystrophy type 2 reveals a novel interaction with MBNL1. Hum Mol Genet. 2015 Feb 15;24(4):954-62. PMID: 25305073.
From the abstract: "Expanded non-coding RNA repeats of CUG and CCUG are the underlying genetic causes for myotonic dystrophy type 1 (DM1) and type 2 (DM2), respectively. A gain-of-function of these pathogenic repeat expansions is mediated at least in part by their abnormal interactions with RNA-binding proteins such as MBNL1 and resultant loss of activity of these proteins. To study pathogenic mechanisms of CCUG-repeat expansions in an animal model, we created a fly model of DM2 that expresses pure, uninterrupted CCUG-repeat expansions ranging from 16 to 720 repeats in length. We show that this fly model for DM2 recapitulates key features of human DM2 including RNA repeat-induced toxicity, ribonuclear foci formation and changes in alternative splicing. ... Our results suggest a novel mechanism for interaction between the pathogenic RNA repeat expansions of myotonic dystrophy and MBNL1."
From the abstract: "Expanded non-coding RNA repeats of CUG and CCUG are the underlying genetic causes for myotonic dystrophy type 1 (DM1) and type 2 (DM2), respectively. A gain-of-function of these pathogenic repeat expansions is mediated at least in part by their abnormal interactions with RNA-binding proteins such as MBNL1 and resultant loss of activity of these proteins. To study pathogenic mechanisms of CCUG-repeat expansions in an animal model, we created a fly model of DM2 that expresses pure, uninterrupted CCUG-repeat expansions ranging from 16 to 720 repeats in length. We show that this fly model for DM2 recapitulates key features of human DM2 including RNA repeat-induced toxicity, ribonuclear foci formation and changes in alternative splicing. ... Our results suggest a novel mechanism for interaction between the pathogenic RNA repeat expansions of myotonic dystrophy and MBNL1."
Fly as whole-animal petri dish: exploring pathogenesis related to Huntington's disease
Barbaro BA, Lukacsovich T, Agrawal N, Burke J, Bornemann DJ, Purcell JM, Worthge SA, Caricasole A, Weiss A, Song W, Morozova OA, Colby DW, Marsh JL. Comparative study of naturally occurring huntingtin fragments in Drosophila points to exon 1 as the most pathogenic species in Huntington's disease. Hum Mol Genet. 2015 Feb 15;24(4):913-25. PMID: 25305076; PMCID: PMC4834878.
From the abstract: "Although Huntington's disease is caused by the expansion of a CAG triplet repeat within the context of the 3144-amino acid huntingtin protein (HTT), studies reveal that N-terminal fragments of HTT containing the expanded PolyQ region can be produced by proteolytic processing and/or aberrant splicing. N-terminal HTT fragments are also prevalent in postmortem tissue, and expression of some of these fragments in model organisms can cause pathology. This has led to the hypothesis that N-terminal peptides may be critical modulators of disease pathology, raising the possibility that targeting aberrant splicing or proteolytic processing may present attractive therapeutic targets. ... We have used Drosophila as a model system to determine the relative toxicities of different naturally occurring huntingtin fragments in a system in which genetic background, transgene expression levels and post-translational proteolytic processing can be controlled. ... If this proves correct, efforts to specifically reduce the levels of exon 1 peptides or to target toxicity-influencing post-translational modifications that occur with the exon 1 context are likely to have the greatest impact on pathology."
From the abstract: "Although Huntington's disease is caused by the expansion of a CAG triplet repeat within the context of the 3144-amino acid huntingtin protein (HTT), studies reveal that N-terminal fragments of HTT containing the expanded PolyQ region can be produced by proteolytic processing and/or aberrant splicing. N-terminal HTT fragments are also prevalent in postmortem tissue, and expression of some of these fragments in model organisms can cause pathology. This has led to the hypothesis that N-terminal peptides may be critical modulators of disease pathology, raising the possibility that targeting aberrant splicing or proteolytic processing may present attractive therapeutic targets. ... We have used Drosophila as a model system to determine the relative toxicities of different naturally occurring huntingtin fragments in a system in which genetic background, transgene expression levels and post-translational proteolytic processing can be controlled. ... If this proves correct, efforts to specifically reduce the levels of exon 1 peptides or to target toxicity-influencing post-translational modifications that occur with the exon 1 context are likely to have the greatest impact on pathology."
Video methods: modeling TBI in the fly
Katzenberger RJ, Loewen CA, Bockstruck RT, Woods MA, Ganetzky B, Wassarman DA. A Method to Inflict Closed Head Traumatic Brain Injury in Drosophila. J Vis Exp. 2015 Jun 30;(100):e52905. doi: 10.3791/52905. Erratum in: J Vis Exp. 2015;(101):e5744. Ganetky, Barry [corrected to Ganetzky, Barry]. PMID: 26168076.
From the abstract: "Traumatic brain injury (TBI) affects millions of people each year, causing impairment of physical, cognitive, and behavioral functions and death. Studies using Drosophila have contributed important breakthroughs in understanding neurological processes. Thus, with the goal of understanding the cellular and molecular basis of TBI pathologies in humans, we developed the High Impact Trauma (HIT) device to inflict closed head TBI in flies. ... the HIT device can be used to perform large-scale genetic screens to understand the genetic basis of TBI pathologies."
From the abstract: "Traumatic brain injury (TBI) affects millions of people each year, causing impairment of physical, cognitive, and behavioral functions and death. Studies using Drosophila have contributed important breakthroughs in understanding neurological processes. Thus, with the goal of understanding the cellular and molecular basis of TBI pathologies in humans, we developed the High Impact Trauma (HIT) device to inflict closed head TBI in flies. ... the HIT device can be used to perform large-scale genetic screens to understand the genetic basis of TBI pathologies."
Results of fly study shed new light on cellular and organismal functions of ALS-related proteins
Coyne AN, Yamada SB, Siddegowda BB, Estes PS, Zaepfel BL, Johannesmeyer JS, Lockwood DB, Pham LT, Hart MP, Cassel JA, Freibaum B, Boehringer AV, Taylor JP, Reitz AB, Gitler AD, Zarnescu DC. Fragile X protein mitigates TDP-43 toxicity by remodeling RNA granules and restoring translation. Hum Mol Genet. 2015 Dec 15;24(24):6886-98. PMID: 26385636; PMCID: PMC5007633.
From the abstract: "RNA dysregulation is a newly recognized disease mechanism in amyotrophic lateral sclerosis (ALS). Here we identify Drosophila fragile X mental retardation protein (dFMRP) as a robust genetic modifier of TDP-43-dependent toxicity in a Drosophila model of ALS. We find that dFMRP overexpression (dFMRP OE) mitigates TDP-43 dependent locomotor defects and reduced lifespan in Drosophila. TDP-43 and FMRP form a complex in flies and human cells. ... Our data suggest a model whereby dFMRP is neuroprotective by remodeling TDP-43 containing RNA granules, reducing aggregation and restoring the translation of specific mRNAs in motor neurons."
From the abstract: "RNA dysregulation is a newly recognized disease mechanism in amyotrophic lateral sclerosis (ALS). Here we identify Drosophila fragile X mental retardation protein (dFMRP) as a robust genetic modifier of TDP-43-dependent toxicity in a Drosophila model of ALS. We find that dFMRP overexpression (dFMRP OE) mitigates TDP-43 dependent locomotor defects and reduced lifespan in Drosophila. TDP-43 and FMRP form a complex in flies and human cells. ... Our data suggest a model whereby dFMRP is neuroprotective by remodeling TDP-43 containing RNA granules, reducing aggregation and restoring the translation of specific mRNAs in motor neurons."
Tuesday, November 22, 2016
Review of involvement in disease of human orthologs of the Drosophila gene crumbs
Slavotinek AM. The Family of Crumbs Genes and Human Disease. Mol Syndromol. 2016 Oct;7(5):274-281. PMID: 27867342.
From the abstract: "The family of vertebrate Crumbs proteins, homologous to Drosophila Crumbs (Crb), share large extracellular domains with epidermal growth factor-like repeats and laminin-globular domains, a single transmembrane domain, and a short intracellular C-terminus containing a single membrane proximal 4.1/ezrin/radixin/moesin-binding domain and PSD-95/Discs large/ZO-1-binding motifs. There are 3 Crb genes in humans ... Bilallelic loss-of-function mutations in CRB1 cause visual impairment, with Leber's congenital amaurosis and retinitis pigmentosa, whereas CRB2 mutations are associated with raised maternal serum and amniotic fluid alpha feto-protein levels, ventriculomegaly/hydrocephalus, and renal disease, ranging from focal segmental glomerulosclerosis to congenital Finnish nephrosis. ... In this review, we summarize the phenotypic findings associated with deleterious sequence variants in CRB1 and CRB2. We discuss the mutational spectrum, animal models of loss of function for both genes and speculate on the likely mechanisms of disease."
From the abstract: "The family of vertebrate Crumbs proteins, homologous to Drosophila Crumbs (Crb), share large extracellular domains with epidermal growth factor-like repeats and laminin-globular domains, a single transmembrane domain, and a short intracellular C-terminus containing a single membrane proximal 4.1/ezrin/radixin/moesin-binding domain and PSD-95/Discs large/ZO-1-binding motifs. There are 3 Crb genes in humans ... Bilallelic loss-of-function mutations in CRB1 cause visual impairment, with Leber's congenital amaurosis and retinitis pigmentosa, whereas CRB2 mutations are associated with raised maternal serum and amniotic fluid alpha feto-protein levels, ventriculomegaly/hydrocephalus, and renal disease, ranging from focal segmental glomerulosclerosis to congenital Finnish nephrosis. ... In this review, we summarize the phenotypic findings associated with deleterious sequence variants in CRB1 and CRB2. We discuss the mutational spectrum, animal models of loss of function for both genes and speculate on the likely mechanisms of disease."
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