Loss-of-function mutations in the gene encoding filaggrin underlie a Japanese family with food-dependent exercise-induced anaphylaxis

Abstract Variants in the GABRB3 gene encoding the β3-subunit of the γ-aminobutyric acid type A ( receptor are associated with various developmental and epileptic encephalopathies. Typically, these variants cause a loss-of-function molecular phenotype whereby γ-aminobutyric acid has reduced inhibitory effectiveness leading to seizures. Drugs that potentiate inhibitory GABAergic activity, such as nitrazepam, phenobarbital or vigabatrin, are expected to compensate for this and thereby reduce seizure frequency. However, vigabatrin, a drug that inhibits γ-aminobutyric acid transaminase to increase tonic γ-aminobutyric acid currents, has mixed success in treating seizures in patients with GABRB3 variants: some patients experience seizure cessation, but there is hypersensitivity in some patients associated with hypotonia, sedation and respiratory suppression. A GABRB3 variant that responds well to vigabatrin involves a truncation variant (p.Arg194*) resulting in a clear loss-of-function. We hypothesized that patients with a hypersensitive response to vigabatrin may exhibit a different γ-aminobutyric acid A receptor phenotype. To test this hypothesis, we evaluated the phenotype of de novo variants in GABRB3 (p.Glu77Lys and p.Thr287Ile) associated with patients who are clinically hypersensitive to vigabatrin. We introduced the GABRB3 p.Glu77Lys and p.Thr287Ile variants into a concatenated synaptic and extrasynaptic γ-aminobutyric acid A receptor construct, to resemble the γ-aminobutyric acid A receptor expression by a patient heterozygous for the GABRB3 variant. The mRNA of these constructs was injected into Xenopus oocytes and activation properties of each receptor measured by two-electrode voltage clamp electrophysiology. Results showed an atypical gain-of-function molecular phenotype in the GABRB3 p.Glu77Lys and p.Thr287Ile variants characterized by increased potency of γ-aminobutyric acid A without change to the estimated maximum open channel probability, deactivation kinetics or absolute currents. Modelling of the activation properties of the receptors indicated that either variant caused increased chloride flux in response to low concentrations of γ-aminobutyric acid that mediate tonic currents. We therefore propose that the hypersensitivity reaction to vigabatrin is a result of GABRB3 variants that exacerbate GABAergic tonic currents and caution is required when prescribing vigabatrin. In contrast, drug strategies increasing tonic currents in loss-of-function variants are likely to be a safe and effective therapy. This study demonstrates that functional genomics can explain beneficial and adverse anti-epileptic drug effects, and propose that vigabatrin should be considered in patients with clear loss-of-function GABRB3 variants.

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The Multifaceted Roles of MicroRNAs in Cystic Fibrosis

Diagnostics ◽

10.3390/diagnostics10121102 ◽

2020 ◽

Vol 10 (12) ◽

pp. 1102

Author(s):

Fatima Domenica Elisa De Palma ◽

Valeria Raia ◽

Guido Kroemer ◽

Maria Chiara Maiuri

Keyword(s):

Cystic Fibrosis ◽

Respiratory Infections ◽

Current Knowledge ◽

Pancreatic Insufficiency ◽

Clinical Manifestations ◽

Predictive Biomarkers ◽

Loss Of Function ◽

Gene Encoding ◽

Multisystemic Disease

Cystic fibrosis (CF) is a lifelong disorder affecting 1 in 3500 live births worldwide. It is a monogenetic autosomal recessive disease caused by loss-of-function mutations in the gene encoding the chloride channel cystic fibrosis transmembrane conductance regulator (CFTR), the impairment of which leads to ionic disequilibria in exocrine organs. This translates into a chronic multisystemic disease characterized by airway obstruction, respiratory infections, and pancreatic insufficiency as well as hepatobiliary and gastrointestinal dysfunction. Molecular characterization of the mutational heterogeneity of CFTR (affected by more than 2000 variants) improved the understanding and management of CF. However, these CFTR variants are linked to different clinical manifestations and phenotypes, and they affect response to treatments. Expanding evidence suggests that multisystemic disease affects CF pathology via impairing either CFTR or proteins regulated by CFTR. Thus, altering the expression of miRNAs in vivo could constitute an appealing strategy for developing new CF therapies. In this review, we will first describe the pathophysiology and clinical management of CF. Then, we will summarize the current knowledge on altered miRNAs in CF patients, with a focus on the miRNAs involved in the deregulation of CFTR and in the modulation of inflammation. We will highlight recent findings on the potential utility of measuring circulating miRNAs in CF as diagnostic, prognostic, and predictive biomarkers. Finally, we will provide an overview on potential miRNA-based therapeutic approaches.

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Genomic Variation in IbA10G2 and Other Patient-Derived Cryptosporidium hominis Subtypes

Journal of Clinical Microbiology ◽

10.1128/jcm.01798-16 ◽

2016 ◽

Vol 55 (3) ◽

pp. 844-858 ◽

Cited By ~ 16

Author(s):

Per Sikora ◽

Sofia Andersson ◽

Jadwiga Winiecka-Krusnell ◽

Björn Hallström ◽

Cecilia Alsmark ◽

...

Keyword(s):

Parasite Burden ◽

Genomic Variation ◽

Loss Of Function ◽

Oocyst Wall ◽

Gene Encoding ◽

Cryptosporidium Hominis ◽

Content Type ◽

Successful Isolation ◽

Target Dna ◽

Stool Samples

ABSTRACTIn order to improve genotyping and epidemiological analysis ofCryptosporidiumspp., genomic data need to be generated directly from a broad range of clinical specimens. Utilizing a robust method that we developed for the purification and generation of amplified target DNA, we present its application for the successful isolation and whole-genome sequencing of 14 differentCryptosporidium hominispatient specimens. Six isolates of subtype IbA10G2 were analyzed together with a single representative each of 8 other subtypes: IaA20R3, IaA23R3, IbA9G3, IbA13G3, IdA14, IeA11G3T3, IfA12G1, and IkA18G1. Parasite burden was measured over a range of more than 2 orders of magnitude for all samples, while the genomes were sequenced to mean depths of between 17× and 490× coverage. Sequence homology-based functional annotation identified several genes of interest, including the gene encodingCryptosporidiumoocyst wall protein 9 (COWP9), which presented a predicted loss-of-function mutation in all the sequence subtypes, except for that seen with IbA10G2, which has a sequence identical to theCryptosporidium parvumreference Iowa II sequence. Furthermore, phylogenetic analysis showed that all the IbA10G2 genomes form a monophyletic clade in theC. hoministree as expected and yet display some heterogeneity within the IbA10G2 subtype. The current report validates the aforementioned method for isolating and sequencingCryptosporidiumdirectly from clinical stool samples. In addition, the analysis demonstrates the potential in mining data generated from sequencing multiple whole genomes ofCryptosporidiumfrom human fecal samples, while alluding to the potential for a higher degree of genotyping withinCryptosporidiumepidemiology.

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Functional characterization of a loss-of-function mutant I324M of arginine vasopressin receptor 2 in X-linked nephrogenic diabetes insipidus

Scientific Reports ◽

10.1038/s41598-021-90736-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Lixia Wang ◽

Weihong Guo ◽

Chunyun Fang ◽

Wenli Feng ◽

Yumeng Huang ◽

...

Keyword(s):

Diabetes Insipidus ◽

Arginine Vasopressin ◽

Nephrogenic Diabetes Insipidus ◽

Functional Characterization ◽

Gene Mutations ◽

Biochemical Properties ◽

Vasopressin Receptor ◽

Inherited Disease ◽

Loss Of Function ◽

Gene Encoding

AbstractX-linked nephrogenic diabetes insipidus (X-linked NDI) is a rare inherited disease mainly caused by lost-of-function mutations in human AVPR2 gene encoding arginine vasopressin receptor 2 (V2R). Our focus of the current study is on exploration of the functional and biochemical properties of Ile324Met (I324M) mutation identified in a pedigree showing as typical recessive X-linked NDI. We demonstrated that I324M mutation interfered with the conformation of complex glycosylation of V2R. Moreover, almost all of the I324M-V2R failed to express on the cell surface due to being captured by the endoplasmic reticulum control system. We further examined the signaling activity of DDAVP-medicated cAMP and ERK1/2 pathways and the results revealed that the mutant receptor lost the ability in response to DDAVP stimulation contributed to the failure of accumulation of cAMP and phosphorylated ERK1/2. Based on the characteristics of molecular defects of I324M mutant, we selected two reagents (SR49059 and alvespimycin) to determine whether the functions of I324M-V2R can be restored and we found that both compounds can significantly “rescue” I324M mutation. Our findings may provide further insights for understanding the pathogenic mechanism of AVPR2 gene mutations and may offer some implications on development of promising treatments for patients with X-linked NDI.

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WED 202 Lamb-shaffer syndrome: importance of snp array in diagnosing neurodevelopmental syndromes

Journal of Neurology Neurosurgery & Psychiatry ◽

10.1136/jnnp-2018-abn.102 ◽

2018 ◽

Vol 89 (10) ◽

pp. A29.4-A30 ◽

Cited By ~ 1

Author(s):

Ela M Akay ◽

Ian S Schofield ◽

Ming H Lai ◽

Rhys H Thomas

Keyword(s):

Intellectual Disability ◽

Language Impairment ◽

Cervical Vertebrae ◽

Snp Array ◽

Global Developmental Delay ◽

Prior History ◽

Loss Of Function ◽

Mild Intellectual Disability ◽

Gene Encoding ◽

Original Cohort

We describe the seizure phenotype of a 26 year old lady who presented with a probable photic-induced convulsion on a background of mild intellectual disability, facial dysmorphia, fused cervical vertebrae and ventricular septal defect. There was no prior history of seizures.Routine EEG was polyrhythmic with a prominent photoparoxysmal response at 14 Hz and 40 Hz. CT head was normal. A SNP array demonstrated a rare 51 kb deletion at 12 p12.1 which disrupts the SOX5 gene.SOX5 is a developmentally important gene encoding a transcription factor that plays a role in multiple developmental pathways including of the nervous system. Loss of function of this gene is associated with Lamb-Shaffer syndrome, first characterised in 2012 with global developmental delay, intellectual disability, mild dysmorphic facies, language impairment and variable skeletal abnormalities.3 of the original cohort of 16 patients described experienced seizures and the nature of their epilepsy was not further defined. Only a further 7 cases have been reported to date, none of whom experienced seizures. Our case helps to broaden the phenotype of Lamb-Shaffer syndrome, highlights the importance of looking for copy number variation and poses questions regarding the neurobiology of photo-sensitivity.

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Homozygous loss-of-function mutations in the gene encoding the dopamine transporter are associated with infantile parkinsonism-dystonia

Journal of Clinical Investigation ◽

10.1172/jci39060 ◽

2009 ◽

Cited By ~ 20

Author(s):

Manju A. Kurian ◽

Juan Zhen ◽

Shu-Yuan Cheng ◽

Yan Li ◽

Santosh R. Mordekar ◽

...

Keyword(s):

Dopamine Transporter ◽

Loss Of Function ◽

Gene Encoding

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Rett-causing mutations reveal two domains critical for MeCP2 function and for toxicity in MECP2 duplication syndrome mice

eLife ◽

10.7554/elife.02676 ◽

2014 ◽

Vol 3 ◽

Cited By ~ 46

Author(s):

Laura Dean Heckman ◽

Maria H Chahrour ◽

Huda Y Zoghbi

Keyword(s):

Transcriptional Repression ◽

Neurological Disorder ◽

Dominant Negative ◽

Dominant Negative Effect ◽

Loss Of Function ◽

Gene Encoding ◽

C Terminus ◽

Mecp2 Duplication Syndrome ◽

Negative Effect ◽

Duplication Syndrome

Loss of function of the X-linked gene encoding methyl-CpG binding protein 2 (MeCP2) causes the progressive neurological disorder Rett syndrome (RTT). Conversely, duplication or triplication of Xq28 causes an equally wide-ranging progressive neurological disorder, MECP2 duplication syndrome, whose features overlap somewhat with RTT. To understand which MeCP2 functions cause toxicity in the duplication syndrome, we generated mouse models expressing endogenous Mecp2 along with a RTT-causing mutation in either the methyl-CpG binding domain (MBD) or the transcriptional repression domain (TRD). We determined that both the MBD and TRD must function for doubling MeCP2 to be toxic. Mutating the MBD reproduces the null phenotype and expressing the TRD mutant produces milder RTT phenotypes, yet both mutations are harmless when expressed with endogenous Mecp2. Surprisingly, mutating the TRD is more detrimental than deleting the entire C-terminus, indicating a dominant-negative effect on MeCP2 function, likely due to the disruption of a basic cluster.

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Identification and functional characterisation of a novel KCNJ2 mutation, Val302del, causing Andersen–Tawil syndrome

Canadian Journal of Physiology and Pharmacology ◽

10.1139/cjpp-2014-0527 ◽

2015 ◽

Vol 93 (7) ◽

pp. 569-575 ◽

Cited By ~ 2

Author(s):

Balázs Ördög ◽

Lidia Hategan ◽

Mária Kovács ◽

György Seprényi ◽

Zsófia Kohajda ◽

...

Keyword(s):

Membrane Trafficking ◽

Genetic Disorder ◽

Clinical Manifestations ◽

Clinical Importance ◽

Inhibitory Effect ◽

Periodic Paralysis ◽

Inward Rectification ◽

Loss Of Function ◽

Gene Encoding ◽

Functional Characterisation

Loss-of-function mutations of the KCNJ2 gene encoding for the inward rectifier potassium channel subunit Kir2.1 cause Andersen–Tawil Syndrome (ATS), a rare genetic disorder characterised by periodic paralysis, ventricular arrhythmias, and dysmorphic features. Clinical manifestations of the disease appear to vary greatly with the nature of mutation, therefore, functional characterisation of ATS-causing mutations is of clinical importance. In this study, we describe the identification and functional analysis of a novel KCNJ2 mutation, Val302del, identified in a patient with ATS. Heterologously expressed wild type (WT) and Val302del mutant alleles showed similar subcellular distribution of the Kir2.1 protein with high intensity labelling from the membrane region, demonstrating normal membrane trafficking of the Val302del Kir2.1 variant. Cells transfected with the WT allele displayed a robust current with strong inward rectification, while no current above background was detected in cells expressing the Val302del Kir2.1 subunit. Co-transfection of CHO cells with the WT and the Val302del Kir2.1 revealed a dose-dependent inhibitory effect of the Val302del Kir2.1 mutant subunit on WT Kir2.1 currents. These observations indicate that the WT and the Val302del mutant subunits co-assemble in the cell membrane and that the mutation affects potassium conductivity and (or) gating of the WT/Val302del heteromeric Kir2.1 channels.

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The role of the NK-homeobox gene slouch (S59) in somatic muscle patterning

Development ◽

10.1242/dev.126.20.4525 ◽

1999 ◽

Vol 126 (20) ◽

pp. 4525-4535 ◽

Cited By ~ 12

Author(s):

S. Knirr ◽

N. Azpiazu ◽

M. Frasch

Keyword(s):

Muscle Development ◽

Ectopic Expression ◽

Homeobox Gene ◽

Drosophila Embryo ◽

Loss Of Function ◽

Cell Fates ◽

Somatic Muscle ◽

Gene Encoding ◽

Body Wall Muscles ◽

Founder Cells

In the Drosophila embryo, a distinct class of myoblasts, designated as muscle founders, prefigures the mature pattern of somatic body wall muscles. Each founder cell appears to be instrumental in generating a single larval muscle with a defined identity. The NK homeobox gene S59 was the first of a growing number of proposed ‘identity genes’ that have been found to be expressed in stereotyped patterns in specific subsets of muscle founders and their progenitor cells and are thought to control their developmental fates. In the present study, we describe the effects of gain- and loss-of-function experiments with S59. We find that a null mutation in the gene encoding S59, which we have named slouch (slou), disrupts the development of all muscles that are derived from S59-expressing founder cells. The observed phenotypes upon mutation and ectopic expression of slouch include transformations of founder cell fates, thus confirming that slouch (S59) functions as an identity gene in muscle development. These fate transformations occur between sibling founder cells as well as between neighboring founders that are not lineage-related. In the latter case, we show that slouch (S59) activity is required cell-autonomously to repress the expression of ladybird (lb) homeobox genes, thereby preventing specification along the lb pathway. Together, these findings provide new insights into the regulatory interactions that establish the somatic muscle pattern.

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