Dominant-negative loss of function arises from a second, more frequent variant within the SAND domain of autoimmune regulator ( AIRE )

AbstractAutosomal dominant hyper-IgE syndrome caused by dominant-negative loss-of-function mutations in signal transducer and activator of transcription factor 3 (STAT3) (STAT3-HIES) is a rare primary immunodeficiency with multisystem pathology. The quality of life in patients with STAT3-HIES is determined by not only the progressive, life-limiting pulmonary disease, but also significant skin disease including recurrent infections and abscesses requiring surgery. Our early report indicated that hematopoietic stem cell transplantation might not be effective in patients with STAT3-HIES, although a few subsequent reports have reported successful outcomes. We update on progress of our patient now with over 18 years of follow-up and report on an additional seven cases, all of whom have survived despite demonstrating significant disease-related pathology prior to transplant. We conclude that effective cure of the immunological aspects of the disease and stabilization of even severe lung involvement may be achieved by allogeneic hematopoietic stem cell transplantation. Recurrent skin infections and abscesses may be abolished. Donor TH17 cells may produce comparable levels of IL17A to healthy controls. The future challenge will be to determine which patients should best be offered this treatment and at what point in their disease history.

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A Modified Screening System for Loss-of-Function and Dominant Negative Alleles of Essential MCMV Genes

PLoS ONE ◽

10.1371/journal.pone.0094918 ◽

2014 ◽

Vol 9 (4) ◽

pp. e94918

Author(s):

Madlen Pogoda ◽

Jens B. Bosse ◽

Karl-Klaus Conzelmann ◽

Ulrich H. Koszinowski ◽

Zsolt Ruzsics

Keyword(s):

Dominant Negative ◽

Screening System ◽

Loss Of Function

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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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Cation channel regulation by COOH-terminal cytoplasmic tail of polycystin-1: mutational and functional analysis

Physiological Genomics ◽

10.1152/physiolgenomics.00092.2001 ◽

2002 ◽

Vol 8 (2) ◽

pp. 87-98 ◽

Cited By ~ 15

Author(s):

David H. Vandorpe ◽

Sabine Wilhelm ◽

Lianwei Jiang ◽

Oxana Ibraghimov-Beskrovnaya ◽

Marina N. Chernova ◽

...

Keyword(s):

Fusion Protein ◽

Point Mutations ◽

Coiled Coil ◽

Cytoplasmic Tail ◽

Surface Expression ◽

Dominant Negative ◽

Cation Channel ◽

Loss Of Function ◽

Cation Current ◽

Autosomal Dominant Polycystic Kidney

Polycystin-1 (PKD1) mutations account for ∼85% of autosomal dominant polycystic kidney disease (ADPKD). We have shown previously that oocyte surface expression of a transmembrane fusion protein encoding part of the cytoplasmic COOH terminus of PKD1 increases activity of a Ca2+-permeable cation channel. We show here that human ADPKD mutations incorporated into this fusion protein attenuated or abolished encoded cation currents. Point mutations and truncations showed that cation current expression requires integrity of a region encompassing the putative coiled coil domain of the PKD1 cytoplasmic tail. Whereas these loss-of-function mutants did not exhibit dominant negative phenotypes, coexpression of a fusion protein expressing the interacting COOH-terminal cytoplasmic tail of PKD2 did suppress cation current. Liganding of the ectodomain of the PKD1 fusion protein moderately activated cation current. The divalent cation permeability and pharmacological profile of the current has been extended. Inducible expression of the PKD1 fusion in EcR-293 cells was also associated with activation of cation channels and increased Ca2+ entry.

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Caenorhabditis elegans SUR-5, a Novel but Conserved Protein, Negatively Regulates LET-60 Ras Activity during Vulval Induction

Molecular and Cellular Biology ◽

10.1128/mcb.18.8.4556 ◽

1998 ◽

Vol 18 (8) ◽

pp. 4556-4564 ◽

Cited By ~ 88

Author(s):

Trent Gu ◽

Satoshi Orita ◽

Min Han

Keyword(s):

Caenorhabditis Elegans ◽

Sequence Similarity ◽

Signal Transduction Pathway ◽

Specific Aspect ◽

Dominant Negative ◽

Loss Of Function ◽

Negative Function ◽

Ras Activation ◽

Dominant Negative Mutations ◽

Novel Protein

ABSTRACT The let-60 ras gene acts in a signal transduction pathway to control vulval differentiation in Caenorhabditis elegans. By screening suppressors of a dominant negativelet-60 ras allele, we isolated three loss-of-function mutations in the sur-5 gene which appear to act as negative regulators of let-60 ras during vulval induction.sur-5 mutations do not cause an obvious mutant phenotype of their own, and they appear to specifically suppress only one of the two groups of let-60 ras dominant negative mutations, suggesting that the gene may be involved in a specific aspect of Ras activation. Consistent with its negative function, overexpressing sur-5 from an extragenic array partially suppresses the Multivulva phenotype of an activatedlet-60 ras mutation and causes synergistic phenotypes with a lin-45 raf mutation. We have clonedsur-5 and shown that it encodes a novel protein. We have also identified a potential mammalian SUR-5 homolog that is about 35% identical to the worm protein. SUR-5 also has some sequence similarity to acetyl coenzyme A synthetases and is predicted to contain ATP/GTP and AMP binding sites. Our results suggest thatsur-5 gene function may be conserved through evolution.

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Identification of a Rare and Potential Pathogenic MC4R Variant in a Brazilian Patient With Adulthood-Onset Severe Obesity

Frontiers in Genetics ◽

10.3389/fgene.2020.608840 ◽

2020 ◽

Vol 11 ◽

Author(s):

Kaio Cezar Rodrigues Salum ◽

Guilherme Orofino de Souza ◽

Gabriella de Medeiros Abreu ◽

Mário Campos Junior ◽

Fabiana Barzotto Kohlrausch ◽

...

Keyword(s):

Body Weight ◽

Severe Obesity ◽

Rare Variants ◽

Dominant Negative ◽

Dominant Negative Effect ◽

Loss Of Function ◽

Coding Region ◽

Childhood Onset ◽

Brazilian Patient ◽

Onset Group

BackgroundThe melanocortinergic pathway orchestrates the energy homeostasis and impairments in this system often lead to an increase in body weight. Rare variants in the melanocortin 4 receptor (MC4R) gene resulting in partial or complete loss of function have been described with autosomal co-dominant inheritance. These mutations are the most common cause of non-syndromic monogenic obesity. In this context, this study aimed to sequence the MC4R gene in a Brazilian cohort of adults with severe obesity.MethodsThis study included 163 unrelated probands with Body Mass Index (BMI) ≥ 35 kg/m2, stratified into three groups, according to the period of obesity onset. From the total sample, 25 patients were enrolled in the childhood-onset group (0–11 years), 19 patients in the adolescence/youth-onset group (12–21 years), and 119 patients in the adult-onset group (>21 years). Blood pressure, anthropometric and biochemical characteristics were obtained, and the MC4R coding region of each subject’s DNA was assessed using automated Sanger sequencing.ResultsSignificant anthropometric differences between the groups were observed. Higher body weight and BMI medians were found in patients with childhood-onset or adolescence/youth-onset when compared to the adulthood-onset obesity group. A total of five mutations were identified, including four missense variants: p.Ser36Thr, p.Val103Ile, p.Ala175Thr, and p.Ile251Leu. Additionally, we observed one synonymous variant (p.Ile198=). The p.Ala175Thr variant was identified in a female case with severe obesity and adulthood-onset. This variant was previously described as a partial loss-of-function mutation, in which the minor allele poses dominant-negative effect, probably resulting in reduced cAMP activity.ConclusionThis study showed a prevalence of common and rare variants in a cohort of Brazilian adults with severe obesity and candidates to bariatric surgery. We have identified a rare potentially pathogenic MC4R variant in a Brazilian patient with severe and adulthood-onset obesity.

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Impaired cytoplasmic domain interactions cause co-assembly defect and loss of function in the p.Glu293Lys KNCJ2 variant isolated from an Andersen–Tawil syndrome patient

Cardiovascular Research ◽

10.1093/cvr/cvaa249 ◽

2020 ◽

Author(s):

Szilvia Déri ◽

János Borbás ◽

Teodóra Hartai ◽

Lidia Hategan ◽

Beáta Csányi ◽

...

Keyword(s):

Salt Bridge ◽

Cytoplasmic Domain ◽

Inward Rectifier ◽

Dominant Negative ◽

Dominant Negative Effect ◽

Causative Role ◽

Loss Of Function ◽

Subunit Interactions ◽

Negative Effect

Abstract Aims Subunit interactions at the cytoplasmic domain interface (CD-I) have recently been shown to control gating in inward rectifier potassium channels. Here we report the novel KCNJ2 variant p.Glu293Lys that has been found in a patient with Andersen–Tawil syndrome type 1 (ATS1), causing amino acid substitution at the CD-I of the inward rectifier potassium channel subunit Kir2.1. Neither has the role of Glu293 in gating control been investigated nor has a pathogenic variant been described at this position. This study aimed to assess the involvement of Glu293 in CD-I subunit interactions and to establish the pathogenic role of the p.Glu293Lys variant in ATS1. Methods and results The p.Glu293Lys variant produced no current in homomeric form and showed dominant-negative effect over wild-type (WT) subunits. Immunocytochemical labelling showed the p.Glu293Lys subunits to distribute in the subsarcolemmal space. Salt bridge prediction indicated the presence of an intersubunit salt bridge network at the CD-I of Kir2.1, with the involvement of Glu293. Subunit interactions were studied by the NanoLuc® Binary Technology (NanoBiT) split reporter assay. Reporter constructs carrying NanoBiT tags on the intracellular termini produced no bioluminescent signal above background with the p.Glu293Lys variant in homomeric configuration and significantly reduced signals in cells co-expressing WT and p.Glu293Lys subunits simultaneously. Extracellularly presented reporter tags, however, generated comparable bioluminescent signals with heteromeric WT and p.Glu293Lys subunits and with homomeric WT channels. Conclusions Loss of function and dominant-negative effect confirm the causative role of p.Glu293Lys in ATS1. Co-assembly of Kir2.1 subunits is impaired in homomeric channels consisting of p.Glu293Lys subunits and is partially rescued in heteromeric complexes of WT and p.Glu293Lys Kir2.1 variants. These data point to an important role of Glu293 in mediating subunit assembly, as well as in gating of Kir2.1 channels.

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Acute Myeloid Leukemia with Deletion 9q Is Associated with CEBPA Loss-of-Function Mutations.

Blood ◽

10.1182/blood.v104.11.2896.2896 ◽

2004 ◽

Vol 104 (11) ◽

pp. 2896-2896

Author(s):

Stefan Frohling ◽

Richard F. Schlenk ◽

Jurgen Krauter ◽

Arnold Ganser ◽

Christian Thiede ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Mutational Analysis ◽

Good Prognosis ◽

Dominant Negative ◽

Binding Potential ◽

Complex Karyotype ◽

Loss Of Function ◽

Cebpa Mutations ◽

Acute Myeloid

Abstract The transcription factor CCAAT/enhancer binding protein alpha (C/EBPalpha) plays an important role in the development of mature neutrophils. Two common types of mutations in the CEBPA gene encoding C/EBPalpha have been identified in approximately 10% of adults with acute myeloid leukemia (AML): N-terminal, dominant-negative frameshift mutations that result in loss of C/EBPalpha function and C-terminal in-frame mutations that result in C/EBPalpha proteins with decreased DNA-binding potential. Previous studies concluded that mutant CEBPA predicts favorable outcome in younger AML patients with intermediate-risk karyotypes or normal cytogenetics. To assess the prevalence of CEBPA mutations in specific cytogenetic subgroups, mutational analysis of the CEBPA gene was performed in 125 AML patients. No CEBPA mutations were detected in 77 patients with t(8;21), inv(16)/t(16;16), t(15;17), or balanced translocations with breakpoints in band 11q23. In 58 patients with various non-complex karyotypic abnormalities, CEBPA mutations were present in 8 (14%). Surprisingly, 5 of the 8 patients with del(9q) as the sole aberration or in combination with a single additional abnormality other than t(8;21) had CEBPA mutations associated with loss of C/EBPalpha function. Consequently, 41 additional del(9q) cases were analyzed; 9 had CEBPA loss-of-function mutations. The overall prevalence of CEBPA loss-of-function mutations in cases with del(9q) within a non-complex karyotype was 41% (14 of 34 patients), whereas none of the patients who had a del(9q) within a complex karyotype (n = 7), in combination with a t(8;21) (n = 10), or together with a t(15;17) (n = 1) demonstrated mutant CEBPA. Analysis of associated mutations indicated that alterations of the FLT3, MLL, and NRAS genes are not common cooperating events in the pathogenesis of del(9q) AML with inactivating CEBPA mutations. This is the first study to show that AML with del(9q) is strongly associated with CEBPA loss-of-function mutations. The coincidence of del(9q) with inactivating CEBPA mutations and the fact that del(9q) is a common secondary cytogenetic abnormality in t(8;21)-positive AML, that is characterized by specific down-regulation of CEBPA, raise the possibility that loss of a critical segment of 9q, most likely in 9q22, and disruption of C/EBPalpha function cooperate in the pathogenesis of these leukemias. Further refinement of the commonly deleted segment of 9q using high-resolution techniques is underway to identify the critical gene(s) involved and their role in normal hematopoiesis and leukemogenesis. A collaborative intergroup study has been initiated to define whether the relatively good prognosis associated with del(9q) is related to the presence of a CEBPA mutation.

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Expression of P73 Splice Variants Correlates to Resistance to DNA Damaging Drugs in Childhood T-Lineage Acute Lymphoblastic Leukemia.

Blood ◽

10.1182/blood.v104.11.995.995 ◽

2004 ◽

Vol 104 (11) ◽

pp. 995-995

Author(s):

Marrit Meier ◽

Monique L. Den Boer ◽

Jules P.P. Meijerink ◽

Monique Passier ◽

Elisabeth R. Van Wering ◽

...

Keyword(s):

Acute Lymphoblastic Leukemia ◽

Mononuclear Cells ◽

Splice Variants ◽

Lymphoblastic Leukemia ◽

Mrna Level ◽

Dominant Negative ◽

Leukemic Cells ◽

Mrna Levels ◽

Transactivation Domain ◽

Loss Of Function

Abstract Children with T-lineage Acute Lymphoblastic Leukemia (T-ALL) have a higher relapse-risk and are in-vitro more resistant to therapeutic drugs compared to ALL patients with a precursor-B phenotype. Cellular resistance to anti-cancer agents has previously shown to be associated with failure of P53 family member signaling by abrogation of P53 function due to loss-of-function mutations or dominant-negative inhibition by isoforms of P73 lacking (part of) the N-terminal transactivation domain (P73ΔEX2, P73ΔEX2/3, ΔN-P73 and ΔN’-P73). Since p53 mutations are not commonly found in T-ALL, we investigated the expression levels of p73 splice variants in relation to drug resistance in children with T-ALL. Splice variants were quantitatively measured at the mRNA level in leukemic cells of 55 T-ALL patients and mononuclear cells of 12 non-leukemic controls. TA-p73 (transactivation competent), p73Δex2, p73Δex2/3, ΔN-p73 and ΔN’-p73 were all found to be present at a relatively higher mRNA level in T-ALL patients than controls (P < 0.05 for all), suggesting that expression of the TP73 gene is deregulated in T-ALL. Resistance of T-ALL cells to the DNA damaging drug daunorubicin correlated with mRNA levels of the dominant-negative variants of p73, i.e. ΔN-p73 and ΔN’-p73 (Rs = 0.38, P = 0.03). In contrast, expression of none of the variants, including ΔN-p73 and ΔN’-p73, was related to resistance of T-ALL cells to non-DNA damaging drugs (prednisolone, vincristine and L-asparaginase). In conclusion, high expression of ΔN-p73 and ΔN’-p73 variants possibly contributes to resistance to DNA damaging drugs in childhood T-ALL.

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