Human Molecular Genetics (Hum Mol Genet)

Publications in this journal

• Article: MicroRNA-22 and promoter motif polymorphisms at the Chga locus in genetic hypertension: Functional and therapeutic implications for gene expression and the pathogenesis of hypertension.

  Ryan S Friese, Angelina E Altshuler, Kuixing Zhang, Jose Pablo Miramontes-Gonzalez, C Makena Hightower, Martin L Jirout, Rany M Salem, Jiaur R Gayen, Nitish R Mahapatra, Nilima Biswas, [......], Sucheta Vaingankar, Hyung-Suk Kim, Maïté Courel, Laurent Taupenot, Michael G Ziegler, Nicholas J Schork, Michal Pravenec, Sushil K Mahata, Geert W Schmid-Schönbein, Daniel T O'Connor
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  ABSTRACT: Hypertension is a common hereditary syndrome with unclear pathogenesis. Chromogranin A (Chga), which catalyzes formation and cargo storage of regulated secretory granules in neuroendocrine cells, contributes to blood pressure homeostasis centrally and peripherally. Elevated Chga occurs in Spontaneously Hypertensive Rat (SHR) adrenal glands and plasma, but central expression is unexplored. In this report, we measured SHR and WKY (control) Chga expression in central and peripheral nervous systems, and found Chga protein to be decreased in SHR brainstem yet increased in adrenal and plasma. By re-sequencing, we systematically identified 5 promoter, 2 coding, and one 3'-untranslated-region (3'-UTR) polymorphism at the SHR (versus WKY or BN) Chga locus. Using HXB/BXH recombinant inbred (RI) strain linkage and correlations, we demonstrated genetic determination of Chga expression in SHR, including a cis-QTL (i.e., at the Chga locus), and such expression influenced biochemical determinants of blood pressure, including a cascade of catecholamine biosynthetic enzymes, catecholamines themselves, and steroids. Luciferase reporter assays demonstrated that the 3'-UTR polymorphism (which disrupts microRNA miR-22 motif) and promoter polymorphisms altered gene expression consistent with the decline in SHR central Chga expression. Coding region polymorphisms did not account for changes in Chga expression or function. Thus we hypothesized that the 3'-UTR and promoter mutations lead to dysregulation (diminution) of Chga in brainstem cardiovascular control nuclei, ultimately contributing to the pathogenesis of hypertension in SHR. Accordingly, we demonstrated that in vivo administration of miR-22 antagomir to SHR causes substantial (∼18 mmHg) reductions in blood pressure, opening a novel therapeutic avenue for hypertension.
  Human Molecular Genetics 05/2013;
• Article: Pharmacological chaperones as a potential therapeutic option in methylmalonic aciduria cblB type.

  Ana Jorge-Finnigan, Sandra Brasil, Jarl Underhaug, Pedro Ruíz-Sala, Begoña Merinero, Ruma Banerjee, Lourdes R Desviat, Magdalena Ugarte, Aurora Martinez, Belén Pérez
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  ABSTRACT: Methylmalonic aciduria (MMA) cblB type is caused by mutations in the MMAB gene. This encodes the enzyme ATP:cob(I)alamin adenosyltransferase (ATR), which converts reduced cob(I)alamin to an active adenosylcobalamin cofactor. We recently reported the presence of destabilizing pathogenic mutations that retain some residual ATR activity. The aim of the present study was to seek pharmacological chaperones as a tailored therapy for stabilizing the ATR protein. High-throughput ligand screening of over 2000 compounds was performed; six were found to enhance the thermal stability of purified recombinant ATR. Further studies using a well-established bacterial system in which the recombinant ATR protein was expressed in the presence of these six compounds, showed them all to increase the stability of the wild-type ATR and the p.Ile96Thr mutant proteins. Compound V (N-{[(4-chlorophenyl)carbamothioyl]amino}-2-phenylacetamide) significantly increased this stability and did not act as an inhibitor of the purified protein. Importantly, compound V increased the activity of ATR in patient-derived fibroblasts harboring the destabilizing p.Ile96Thr mutation in a hemizygous state to within control range. When cobalamin was coadministrated with compound V, mutant ATR activity further improved. Oral administration of low doses of compound V to C57BL/6J mice for 12 days, led to increase in steady-state levels of ATR protein in liver and brain (disease-relevant organs). These results hold promise for the clinical use of pharmacological chaperones in MMA cblB type patients harboring chaperone-responsive mutations.
  Human Molecular Genetics 05/2013;
• Article: Enhanced Optineurin E50 K-TBK1 interaction evokes protein insolubility and initiates familial primary open-angle glaucoma.

  Yuriko Minegishi, Daisuke Iejima, Hiroaki Kobayashi, Zai-Long Chi, Kazuhide Kawase, Tetsuya Yamamoto, Tomohisa Seki, Shinsuke Yuasa, Keiichi Fukuda, Takeshi Iwata
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  ABSTRACT: Glaucoma is the leading cause for blindness affecting 60 million people worldwide. The optineurin (OPTN) E50 K mutation was first identified in familial primary open-angle glaucoma (POAG), the onset of which is not associated with intraocular pressure (IOP) elevation, and is classified as normal-tension glaucoma (NTG). Optineurin (OPTN) is a multifunctional protein and its mutations are associated with neurodegenerative diseases such as POAG and amyotrophic lateral sclerosis (ALS). We have previously described an E50 K mutation-carrying transgenic (E50 K(-tg)) mouse that exhibited glaucomatous phenotypes of decreased retinal ganglion cells (RGCs) and surrounding cell death at normal IOP. Further phenotypic analysis of these mice revealed persistent reactive gliosis and E50 K mutant protein deposits in the outer plexiform layer (OPL). Over-expression of E50 K in HEK293 cells indicated accumulation of insoluble OPTN in the endoplasmic reticulum (ER). This phenomenon was consistent with the results seen in neurons derived from induced pluripotent stem cells (iPSCs) from E50 K mutation-carrying NTG patients. The E50 K mutant strongly interacted with TANK-binding kinase 1 (TBK1), which prohibited the proper oligomerization and solubility of OPTN, both of which are important for OPTN intracellular transition. Treatment with a TBK1 inhibitor, BX795, abrogated the aberrant insolubility of the E50 K mutant. Here, we delineated the intracellular dynamics of the endogenous E50 K mutant protein for the first time and demonstrated how this mutation causes OPTN insolubility, in association with TBK1, to evoke POAG.
  Human Molecular Genetics 05/2013;
• Article: Genome-wide Loss of 5-hmC is a Novel Epigenetic Feature of Huntington's Disease.

  Fengli Wang, Yeran Yang, Xiwen Lin, Jiu-Qiang Wang, Yong-Sheng Wu, Wenjuan Xie, Dandan Wang, Shu Zhu, You-Qi Liao, Qinmiao Sun, Yun-Gui Yang, Caixia Guo, Chunsheng Han, Tie-Shan Tang
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  ABSTRACT: 5-hydroxymethylcytosine (5-hmC) may represent a new epigenetic modification of cytosine. While the dynamics of 5-hmC during neurodevelopment have recently been reported, little is known about its genomic distribution and function(s) in neurodegenerative diseases such as Huntington's disease (HD). We here observed a marked reduction of 5-hmC signal in YAC128 (yeast artificial chromosome transgene with 128 CAG repeats) HD mouse brain tissues when compared with age-matched wild type (WT) mice, suggesting a deficiency of 5-hmC reconstruction in HD brains during postnatal development. Genome-wide distribution analysis of 5-hmC further confirmed the diminishment of 5-hmC signal in striatum and cortex in YAC128 HD mice. General genomic features of 5-hmC are highly conserved, not being affected by either disease or brain regions. Intriguingly, we have identified disease-specific (YAC128 versus WT) differentially hydroxymethylated regions (DhMRs), and found that acquisition of DhmRs in gene body is a positive epigenetic regulator for gene expression. Ingenuity pathway analysis of genotype-specific DhMR-annotated genes revealed that alternation of a number of canonical pathways involving neuronal development/differentiation (Wnt/β-catenin/Sox pathway, axonal guidance signaling pathway) and neuronal function/survival (glutamate receptor/calcium/CREB, GABA receptor signaling, dopamine-DARPP32 feedback pathway etc) could be important for the onset of HD. Our results indicate that loss of the 5-hmC marker is a novel epigenetic feature in Huntington's disease, and that this aberrant epigenetic regulation may impair the neurogenesis, neuronal function and survival in HD brain. Our study also opens a new avenue for HD treatment; re-establishing the native 5-hmC landscape may have the potential to slow/halt the progression of HD.
  Human Molecular Genetics 05/2013;
• Article: Bcl6a function is required during optic cup formation to prevent p53-dependent apoptosis and colobomata.

  Jiwoon Lee, Bum-Kyu Lee, Jeffrey M Gross
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  ABSTRACT: Mutations in BCOR (Bcl6 corepressor) are found in patients with oculo-facio-cardio-dental (OFCD) syndrome, a congenital disorder affecting visual system development, and loss of function studies in zebrafish and Xenopus demonstrate a role for Bcor during normal optic cup development in preventing colobomata. The mechanism whereby BCOR functions during eye development to prevent colobomata is not known, but in other contexts it serves as a transcriptional corepressor that potentiates transcriptional repression by B cell leukemia/lymphoma 6 (BCL6). Here, we have explored the function of the zebrafish ortholog of Bcl6, Bcl6a, during eye development, and our results demonstrate that Bcl6a, like Bcor, is required to prevent colobomata during optic cup formation. Our data demonstrate that Bcl6a acts downstream of Vax1 and Vax2, known regulators of ventral optic cup formation and choroid fissure closure, and that bcl6a is a direct target of Vax2. Together, this regulatory network functions to repress p53 expression and thereby suppress apoptosis in the developing optic cup. Furthermore, our data demonstrate that Bcl6a functions cooperatively with Bcor, Rnf2 and Hdac1 in a common gene regulatory network that acts to repress p53 and prevent colobomata. Together, these data support a model in which p53-dependent apoptosis needs to be tightly regulated for normal optic cup formation and that Bcl6a, Bcor, Rnf2 and Hdac1 activities mediate this regulation.
  Human Molecular Genetics 05/2013;
• Article: PGC-1α is a male-specific disease modifier of human and experimental amyotrophic lateral sclerosis.

  Judith Eschbach, Birgit Schwalenstöcker, Selma Soyal, Hanna Bayer, Diana Wiesner, Chizuru Akimoto, Anne-Charloth Nilsson, Anna Birve, Thomas Meyer, Luc Dupuis, Karin Danzer, Peter M Andersen, Anke Witting, Albert C Ludolph, Wolfgang Patsch, Patrick Weydt
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  ABSTRACT: Amyotrophic lateral sclerosis (ALS) is a devastating, adult-onset neurodegenerative disorder of the upper and lower motor systems. It leads to paresis, muscle wasting and inevitably to death, typically within 3 to 5 years. However, disease onset and survival vary considerably ranging in extreme cases from a few months to several decades. The genetic and environmental factors underlying this variability are of great interest as potential therapeutic targets. In ALS men are affected more often and have an earlier age of onset than women. This gender difference is recapitulated in transgenic rodent models, but no underlying mechanism has been elucidated. Here we report, that SNPs in the brain-specific promoter region of the transcriptional co-activator PGC-1α, a master regulator of metabolism, modulate age of onset and survival in two large and independent ALS populations and this occurs in a strictly male-specific manner. In complementary animal studies we show that deficiency of full-length Pgc-1α leads to a significantly earlier age of onset and a borderline shortened survival in male, but not in female ALS-transgenic mice. In the animal model full-length Pgc-1α-loss is associated with reduced mRNA levels of the trophic factor Vegf-A in males, but not in females. In summary we indentify PGC-1α as a novel and clinically relevant disease modifier of human and experimental ALS and report a sex-dependent effect of PGC-1α in this neurodegenerative disorder.
  Human Molecular Genetics 05/2013;
• Article: The Lysosomal Inhibitor, Chloroquine, Increases Cell Surface BMPR-II levels and Restores BMP9 Signalling in Endothelial Cells Harbouring BMPR-II Mutations.

  Benjamin J Dunmore, Kylie M Drake, Paul D Upton, Mark R Toshner, Micheala A Aldred, Nicholas W Morrell
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  ABSTRACT: Pulmonary arterial hypertension (PAH) is characterised by dysregulated pulmonary artery endothelial cell (PAEC) proliferation, apoptosis and permeability. Loss of function mutations in the bone morphogenetic protein receptor type-II (BMPR-II) are the commonest cause of heritable PAH, usually resulting in haploinsufficiency. We previously showed that BMPR-II expression is regulated via a lysosomal degradative pathway. Here we show that the antimalarial drug, chloroquine, markedly increased cell surface expression of BMPR-II protein independent of transcription in PAECs. Inhibition of protein synthesis experiments revealed a rapid turnover of cell surface BMPR-II, which was inhibited by chloroquine treatment. Chloroquine enhanced PAEC expression of BMPR-II following siRNA knockdown of the BMPR-II transcript. Using blood outgrowth endothelial cells (BOECs), we confirmed that signalling in response to the endothelial BMPR-II ligand, BMP9, is compromised in BOECs from patients harbouring BMPR-II mutations, and in BMPR-II mutant pulmonary artery endothelial cells (PAECs). Chloroquine significantly increased gene expression of BMP9-BMPR-II signalling targets Id1, miR21 and miR27a in both mutant BMPR-II PAECs and BOECs. These findings provide support for the restoration of cell surface BMPR-II with agents such as chloroquine as a potential therapeutic approach for heritable PAH.
  Human Molecular Genetics 05/2013;
• Article: Whole-exome sequencing identifies ADAM10 mutations as a cause of reticulate acropigmentation of Kitamura, a clinical entity distinct from Dowling-Degos disease.

  Michihiro Kono, Kazumitsu Sugiura, Mutsumi Suganuma, Masahiro Hayashi, Hiromichi Takama, Tamio Suzuki, Kayoko Matsunaga, Yasushi Tomita, Masashi Akiyama
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  ABSTRACT: Reticulate acropigmentation of Kitamura (RAK) is a rare genetic disorder of cutaneous pigmentation with an autosomal dominant pattern of inheritance and a high penetration rate. The characteristic skin lesions are reticulate, slightly depressed pigmented macules mainly affecting the dorsa of the hands and feet, which first appear before puberty and subsequently expand to the proximal limb and the trunk. To identify mutations that cause RAK, we performed exome sequencing of 4 family members in a pedigree with RAK. 53 SNV/Indels were considered as candidate mutations after some condition narrowing. We confirmed the mutation status in each candidate gene of 4 other members in the same pedigree to find the gene that matched the mutation status and phenotype of each member. A mutation in ADAM10 encoding a zinc metalloprotease, a disintegrin and metalloprotease domain-containing protein 10 (ADAM10) was identified in the RAK family. ADAM10 is known to be involved in the ectodomain shedding of various substrates in the skin. Sanger sequencing of 4 additional unrelated RAK patients revealed 4 additional ADAM10 mutations. We identified a total of 3 truncating mutations, a splice site mutation and a missense mutation in ADAM10. We searched for mutations in the KRT5 gene, a causative gene for the similar pigmentation disorder Dowling-Degos disease, in all the patients and found no KRT5 mutation. These results reveal that mutations in ADAM10 are a cause of RAK and that RAK is an independent clinical entity distinct from Dowling-Degos disease.
  Human Molecular Genetics 05/2013;
• Article: In vitro-differentiated neural cell cultures progress towards donor-identical brain tissue.

  Brooke E Hjelm, Bodour Salhia, Ahmet Kurdoglu, Szabolcs Szelinger, Rebecca A Reiman, Lucia I Sue, Thomas G Beach, Matthew J Huentelman, David W Craig
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  ABSTRACT: Multiple research groups have observed neuropathological phenotypes and molecular symptoms in vitro using induced pluripotent stem cell (iPSC)-derived neural cell cultures (i.e., patient-specific neurons and glia). However, the global differences/similarities that may exist between in vitro neural cells and their tissue-derived counterparts remain largely unknown. In this study, we compared temporal series of iPSC-derived in vitro neural cell cultures to endogenous brain tissue from the same autopsy donor. Specifically, we utilized RNA sequencing (RNA-Seq) to evaluate the transcriptional progression of in vitro-differentiated neural cells (over a timecourse of 0, 35, 70, 105, and 140 days), and compared this to donor-identical temporal lobe tissue. We observed in vitro progression towards the reference brain tissue, and the following three results support this conclusion: (1) there was a significant increasing monotonic correlation between the days of our timecourse and the number of actively transcribed protein-coding genes and long intergenic noncoding RNAs (lincRNAs) (p<0.05), consistent with the transcriptional complexity of the brain; (2) there was an increase in CpG methylation after neural differentiation that resembled the epigenomic signature of the endogenous tissue; and (3) there was a significant decreasing monotonic correlation between the days of our timecourse and the percent of in vitro to brain tissue differences (p<0.05) for tissue-specific protein-coding genes and all putative lincRNAs. Taken together, these results are consistent with in vitro neural development and physiological progression occurring predominantly by transcriptional activation of downregulated genes rather than deactivation of upregulated genes.
  Human Molecular Genetics 05/2013;
• Article: GATA3 controls the specification of prosensory domain and neuronal survival in the mouse cochlea.

  Xiong-Jian Luo, Min Deng, Xiaoling Xie, Liang Huang, Hui Wang, Lichun Jiang, Guoqing Liang, Fang Hu, Roger Tieu, Rui Chen, Lin Gan
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  ABSTRACT: HDR syndrome (also known as Barakat syndrome) is a developmental disorder characterized by hypoparathyroidism, sensorineural deafness, and renal disease. Although genetic mapping and subsequent functional studies indicate that GATA3 haplo-insufficiency causes human HDR syndrome, the role of Gata3 in sensorineural deafness and auditory system development is largely unknown. In this study, we show that Gata3 is continuously expressed in the developing mouse inner ear. Conditional knockout of Gata3 in the developing inner ear disrupts the morphogenesis of mouse inner ear, resulting in a disorganized and shortened cochlear duct with significant fewer hair cells and supporting cells. Loss of Gata3 function leads to the failure in the specification of prosensory domain and subsequently, to increased cell death in the cochlear duct. Moreover, though the initial generation of cochleovestibular ganglion (CVG) cells is not affected in Gata3-null mice, spiral ganglion neurons (SGNs) are nearly depleted due to apoptosis. Our results demonstrate Gata3's essential role in specifying the prosensory domain in the cochlea and in regulating the survival of SGNs, thus identifying a molecular mechanism underlying human HDR syndrome.
  Human Molecular Genetics 05/2013;
• Article: Efficacy of N-acetylcysteine in phenotypic suppression of mouse models of Niemann-Pick disease, type C1.

  Rao Fu, Christopher A Wassif, Nicole M Yanjanin, Dawn E Watkins-Chow, Laura L Baxter, Art Incao, Laura Liscum, Rohini Sidhu, Sally Firnkes, Mark Graham, Daniel S Ory, Forbes D Porter, William J Pavan
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  ABSTRACT: Niemann-Pick disease, type C1 (NPC1), which arises from mutation in the NPC1 gene, is characterized by abnormal cellular storage and transport of cholesterol and other lipids that leads to hepatic disease and progressive neurological impairment. Oxidative stress has been hypothesized to contribute to the NPC1 disease pathological cascade. To determine if treatments reducing oxidative stress could alleviate NPC1 disease phenotypes, the in vivo effects of the antioxidant N-acetylcysteine (NAC) on two mouse models for NPC1 disease were studied. NAC was able to partially suppress phenotypes in both antisense-induced (NPC1ASO) and germline (Npc1-/-) knockout genetic mouse models, confirming the presence of an oxidative stress-related mechanism in progression of NPC1 phenotypes and suggesting NAC as a potential molecule for treatment. Gene expression analyses of NAC-treated NPC1ASO mice suggested NAC affects pathways distinct from those initially altered by Npc1 knockdown, data consistent with NAC achieving partial disease phenotype suppression. In a therapeutic trial of short-term NAC administration to NPC1 patients, no significant effects on oxidative stress in these patients were identified other than moderate improvement of the fraction of reduced CoQ10, suggesting limited efficacy of NAC monotherapy. However, the mouse model data suggest that the distinct antioxidant effects of NAC could provide potential treatment of NPC1 disease, possibly in concert with other therapeutic molecules at earlier stages of disease progression. This paper also validated the NPC1ASO mouse as an efficient model for candidate NPC1 drug screening, and demonstrated similarities in hepatic phenotypes and genome-wide transcript expression patterns between the NPC1ASO and Npc1-/- models.
  Human Molecular Genetics 05/2013;
• Article: Nilotinib reverses loss of dopamine neurons and improves motor behavior via autophagic degradation of α-synuclein in Parkinson's disease models.

  Michaeline L Hebron, Irina Lonskaya, Charbel E-H Moussa
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  ABSTRACT: Parkinson's disease is a movement disorder characterized by death of dopaminergic substantia nigra (SN) neurons and brain accumulation of α-synuclein. The tyrosine kinase Abl is activated in neurodegeneration. Here, we show that lentiviral expression of α-synuclein in the mouse SN leads to Abl activation (phosphorylation) and lentiviral Abl expression increases α-synuclein levels, in agreement with Abl activation in PD brains. Administration of the tyrosine kinase inhibitor nilotinib decreases Abl activity and ameliorates autophagic clearance of α-synuclein in transgenic and lentiviral gene transfer models. Subcellular fractionation shows accumulation of α-synuclein and hyper-phosphorylated Tau (p-Tau) in autophagic vacuoles in α-synuclein expressing brains, but nilotinib enhances protein deposition into the lysosomes. Nilotinib is used for adult leukemia treatment and it enters the brain within US Food and Drug Administration approved doses, leading to autophagic degradation of α-synuclein, protection of SN neurons and amelioration of motor performance. These data suggest that nilotinib may be a therapeutic strategy to degrade α-synuclein in PD and other α-synucleinopathies.
  Human Molecular Genetics 05/2013;
• Article: Dyskeratosis congenita mutations in dyskerin SUMOylation consensus sites lead to impaired telomerase RNA accumulation and telomere defects.

  Marie Eve Brault, Catherine Lauzon, Chantal Autexier
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  ABSTRACT: Mutations in the dyskerin gene (DKC1) cause X-linked dyskeratosis congenita (DC), a rare and fatal premature aging syndrome characterized by defective telomere maintenance. Dyskerin is a highly conserved nucleolar protein, and a component of the human telomerase complex that is essential for human telomerase RNA (hTR) stability. However, its regulation remains poorly understood. Here, we report that dyskerin can be modified by small ubiquitin-like modifiers (SUMO). We find that human DC-causing mutations in highly conserved dyskerin SUMOylation consensus sites lead to impaired hTR accumulation, telomerase activity and telomere maintenance. Finally, we show that modification of dyskerin by SUMOylation is required for its stability. Our findings provide the first evidence that dyskerin stability is regulated by SUMOylation and that mutations altering dyskerin SUMOylation can lead to defects in telomere maintenance that are characteristic of DC.
  Human Molecular Genetics 05/2013;
• Article: Progressive Impairment of Muscle Regeneration in Muscleblind-like 3 Isoform Knockout Mice.

  Michael G Poulos, Ranjan Batra, Moyi Li, Yuan Yuan, Chaolin Zhang, Robert B Darnell, Maurice S Swanson
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  ABSTRACT: The muscleblind-like (MBNL) genes encode alternative splicing factors that are essential for postnatal development of multiple tissues and inhibition of MBNL activity by toxic C(C)UG repeat RNAs is a major pathogenic feature of the neuromuscular disease myotonic dystrophy. While MBNL1 controls fetal-to-adult splicing transitions in muscle and MBNL2 serves a similar role in the brain, the function of MBNL3 in vivo is unknown. Here, we report that mouse Mbnl3, which encodes protein isoforms that differ in the number of tandem zinc-finger RNA-binding motifs and subcellular localization, is expressed primarily during embryonic development but also transiently during injury-induced adult skeletal muscle regeneration. Mbnl3 expression is required for normal C2C12 myogenic differentiation and high-throughput sequencing combined with crosslinking/immunoprecipitation analysis indicates Mbnl3 binds preferentially to the 3' untranslated regions of genes implicated in cell growth and proliferation. In addition, Mbnl3(ΔE2) isoform knockout mice, which fail to express the major Mbnl3 nuclear isoform, show age-dependent delays in injury-induced muscle regeneration and impaired muscle function. These results suggest that Mbnl3 inhibition by toxic RNA expression may be a contributing factor to the progressive skeletal muscle weakness and wasting characteristic of myotonic dystrophy.
  Human Molecular Genetics 05/2013;
• Article: Celecoxib increases SMN and survival in a severe spinal muscular atrophy mouse model via p38 pathway activation.

  Faraz Farooq, Francisco Abadía-Molina, Duncan Mackenzie, Jeremiah Hadwen, Fahad Shamim, Sean O' Reilly, Martin Holcik, Alex Mackenzie
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  ABSTRACT: The loss of functional Survival Motor Neuron (SMN) protein due to mutations or deletion in the SMN1 gene causes autosomal recessive neurodegenerative spinal muscle atrophy (SMA). A potential treatment strategy for SMA is to upregulate the amount of SMN protein originating from the highly homologous SMN2 gene, compensating in part for the absence of the functional SMN1 gene. We have previously shown in vitro activation of the p38 pathway stabilizes and increases SMN mRNA levels leading to increased SMN protein levels. In this report we explore the impact of the FDA approved, blood brain barrier permeating compound celecoxib and known p38 activator on SMN levels in vitro and in a mouse model of SMA. We demonstrate a significant induction of SMN mRNA and protein levels in human and mouse neuronal cells upon treatment with celecoxib. We show that activation of the p38 pathway by low doses celecoxib increases SMN mRNA in a HuR protein dependent manner. Furthermore, celecoxib treatment induces SMN expression in brain and spinal cord samples of wild type mice in vivo. Critically, celecoxib treatment increased SMN levels, improved motor function and enhanced survival in a severe SMA mouse model. Our re sults identify low dose celecoxib as a potential new member of the SMA therapeutic armamentarium.
  Human Molecular Genetics 05/2013;
• Article: APP/PS1 mice overexpressing SREBP-2 exhibit combined Aβ accumulation and tau pathology underlying Alzheimer's disease.

  Elisabet Barbero-Camps, Anna Fernández, Laura Martínez, Jose C Fernández-Checa, Anna Colell
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  ABSTRACT: Current evidence indicates that excess brain cholesterol regulates amyloid-β (Aβ) deposition, which in turn can regulate cholesterol homeostasis. Moreover, Aβ neurotoxicity is potentiated, in part, by mitochondrial glutathione (mGSH) depletion. To better understand the relationship between alterations in cholesterol homeostasis and Alzheimer's disease (AD), we generated a triple transgenic mice featuring sterol regulatory element-binding protein-2 (SREBP-2) overexpression in combination with APPswe/PS1ΔE9 mutations (APP/PS1) to examine key biochemical and functional characteristics of AD. Unlike APP/PS1 mice, APP/PS1/SREBP-2 mice exhibited early mitochondrial cholesterol loading and mGSH depletion. Moreover, β-secretase activation and Aβ accumulation, correlating with oxidative damage and neuroinflammation, were accelerated in APP/PS1/SREBP-2 mice compared to APP/PS1 mice. Triple transgenic mice displayed increased synaptotoxicity reflected by loss of synaptophysin and neuronal death, resulting in early object-recognition memory impairment associated with deficits in spatial memory. Interestingly, tau pathology was present in APP/PS1/SREBP-2 mice, manifested by increased tau hyperphosphorylation and cleavage, activation of tau kinases and neurofibrillary tangle formation without expression of mutated tau. Importantly, in vivo treatment with the cell permeable GSH ethyl ester, which restored mGSH levels in APP/PS1/SREBP-2 mice, partially prevented the activation of tau kinases, reduced abnormal tau aggregation and Aβ deposition, resulting in attenuated synaptic degeneration. Taken together, these results show that cholesterol-mediated mGSH depletion is a key event in AD progression, accelerating the onset of key neuropathological hallmarks of the disease. Thus, therapeutic approaches to recover mGSH may represent a relevant strategy in the treatment of AD.
  Human Molecular Genetics 05/2013;
• Article: Functional implications of splicing polymorphisms in the human genome.

  Yerbol Z Kurmangaliyev, Roman A Sutormin, Sergey A Naumenko, Georgii A Bazykin, Mikhail S Gelfand
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  ABSTRACT: Proper splicing is often crucial for gene functioning and its disruption may be strongly deleterious. Nevertheless, even the essential for splicing canonical dinucleotides of the splice sites are often polymorphic. Here, we use data from The 1000 Genomes Project to study SNPs in the canonical dinucleotides. Splice sites carrying SNPs are enriched in weakly expressed genes and in rarely-used alternative splice sites. Genes with disrupted splice sites tend to have low selective constraint, and the splice sites disrupted by SNPs are less likely to be conserved in mouse. Furthermore, SNPs are enriched in splice sites whose effects on gene function are minor: splice sites located outside of protein-coding regions, in shorter exons, closer to the 3'-ends of proteins, and outside of functional protein domains. Most of these effects are more pronounced for high-frequency SNPs. Despite these trends, many of the polymorphic sites may still substantially affect the function of the corresponding genes. A number of the observed splice site-disrupting SNPs, including several high-frequency ones, were found among mutations described in OMIM.
  Human Molecular Genetics 05/2013;
• Article: Investigation of six testicular germ cell tumor susceptibility genes suggests a parent-of-origin effect in SPRY4.

  Robert Karlsson, Kristine E Andreassen, Wenche Kristiansen, Elin L Aschim, Roy M Bremnes, Olav Dahl, Sophie D Fosså, Olbjørn Klepp, Carl W Langberg, Arne Solberg, Steinar Tretli, Patrik K E Magnusson, Hans-Olov Adami, Trine B Haugen, Tom Grotmol, Fredrik Wiklund
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  ABSTRACT: Recent genome-wide association studies have identified single nucleotide polymorphisms (SNPs) associated with testicular germ cell tumour (TGCT) risk in the genes ATF7IP, BAK1, DMRT1, KITLG, SPRY4, and TERT. In the present study we validate these associations in a Scandinavian population, and explore effect modification by parental sex and differences in associations between the major histological subtypes seminoma and non-seminoma.A total of 118 SNPs in the six genes were genotyped in a population-based Swedish-Norwegian sample comprising 831 TGCT case-parent triads, 474 dyads, 712 singletons and 3919 population controls. 734 additional SNPs were imputed using reference haplotypes from the 1000 genomes project. SNP-TGCT association was investigated using a likelihood-based association test for nuclear families and unrelated subjects implemented in the software package UNPHASED. Forward stepwise regression within each gene was applied to determine independent association signals. Effect modification by parent-of-origin and effect differences between histological subtypes were explored.We observed strong association between SNPs in all six genes and TGCT (lowest P-value per gene: ATF7IP 6.2×10(-6); BAK1 2.1×10(-10); DMRT1 6.7×10(-25); KITLG 2.1×10(-48); SPRY4 1.4×10(-29); TERT 1.8×10(-18)). Stepwise regression indicated three independent signals for BAK1 and TERT, two for SPRY4, and one each for DMRT1, ATF7IP and KITLG. A significant parent-of-origin effect was observed for rs10463352 in SPRY4 (maternal odds ratio=1.72, paternal odds ratio=0.99, interaction P=0.0013). No significant effect differences between seminomas and non-seminomas were found.In summary, we validated previously reported genetic associations with TGCT in a Scandinavian population, and observed suggestive evidence of a parent-of-origin effect in SPRY4.
  Human Molecular Genetics 05/2013;
• Article: Venous malformation -causative TIE2-mutations mediate an AKT-dependent decrease in PDGFB.

  Melanie Uebelhoer, Marjut Nätynki, Jaakko Kangas, Antonella Mendola, Ha-Long Nguyen, Julie Soblet, Catherine Godfraind, Laurence M Boon, Lauri Eklund, Nisha Limaye, Miikka Vikkula
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  ABSTRACT: Mutations in the endothelial cell tyrosine kinase receptor TIE2 cause inherited and sporadic forms of venous malformation. The recurrent somatic mutation L914F and common germline mutation R849W differ in terms of phosphorylation-level, as well as sub-cellular localization and trafficking of the receptor. Previous studies have shed light on certain pathogenic properties of R849W, but the mechanisms of action of L914F are unknown. We used global gene expression profiling to study the effects of L914F on endothelial cells. We found that L914F strongly dysregulated genes involved in vascular development, cell migration, and extracellular matrix processing, while R849W had weak effects. We also demonstrated, for the first time, that TIE2-mutant endothelial cells were deficient in the production of PDGFB, both in vitro and ex vivo in patient tissues. This defect was mediated by the chronic, ligand-independent activation of AKT by the mutant receptors. Inadequate secretion of the major mural cell attractant likely plays an important role in the development of abnormal vascular channels, contributing to the characteristic paucity of surrounding vascular smooth muscle cells.
  Human Molecular Genetics 04/2013;
• Article: A novel function of Ataxin-1 in the modulation of PP2A activity is dysregulated in the spinocerebellar ataxia type 1.

  Ivelisse Sánchez, Patricia Piñol, Marc Corral-Juan, Massimo Pandolfo, Antoni Matilla-Dueñas
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  ABSTRACT: An expansion of glutamines within the human ataxin-1 protein underlies spinocerebellar ataxia type 1 (SCA1), a dominantly inherited neurodegenerative disorder characterized by ataxia and loss of cerebellar Purkinje neurons. Although the mechanisms linking the mutation to the disease remain unclear, evidence indicates it involves a combination of both gain and loss of functions of ataxin-1. We previously showed that the mutant ataxin-1 interacts with Anp32a, a potent and selective PP2A inhibitor, suggesting a role of PP2A in SCA1. Herein we found a new function of ataxin-1 (Atxn1): the modulation of Pp2a activity and the regulation of its holoenzyme composition, with the polyglutamine-mutation within Atxn1 altering this function in the SCA1 mouse cerebellum before disease onset. We show that ataxin-1 enhances Pp2a-bβ expression and down-regulates Anp32a levels without affecting post-translational modifications of Pp2a catalytic subunit (Pp2a-c) known to regulate Pp2a activity. In contrast, mutant Atxn1 induces a decrease in Y307-phosphorylation in Pp2a-c, known to enhance its activity, while reducing Pp2a-b expression and inhibiting Anp32a levels. qRT-PCR and ChIP analyses show that ataxin-1-mediated regulations of the Pp2a-bβ subunit, specifically bβ2, and of Anp32a occur at the transcriptional level. The Pp2a pathway alterations were confirmed by identified phosphorylation changes of the known Pp2a-substrates, Erk2 and Gsk3β. Similarly, mutant ataxin-1-expressing SH-SY5Y cells exhibit abnormal neuritic morphology, decreased levels of both PP2A-Bβ and ANP32A, and PP2A pathway alterations, all of which are ameliorated by overexpressing ANP32A. Our results point to dysregulation of this newly-assigned function of ataxin-1 in SCA1 uncovering new potential targets for therapy.
  Human Molecular Genetics 04/2013;