scholarly journals A Novel Mutation in Cse1l Disrupts Brain and Eye Development with Specific Effects on Pax6 Expression

2021 ◽  
Vol 9 (3) ◽  
pp. 27
Author(s):  
Lauren E. Blizzard ◽  
Chelsea Menke ◽  
Shaili D. Patel ◽  
Ronald R. Waclaw ◽  
Salil A. Lachke ◽  
...  
Keyword(s):  
Neural Development ◽  
Null Allele ◽  
Eye Development ◽  
Novel Mutation ◽  
Forward Genetics ◽  
Mammalian Development ◽  
Variants Of Unknown Significance ◽  
New Genes ◽  
Conditional Allele ◽  
Sequencing Studies

Forward genetics in the mouse continues to be a useful and unbiased approach to identifying new genes and alleles with previously unappreciated roles in mammalian development and disease. Here, we report a new mouse allele of Cse1l that was recovered from an ENU mutagenesis screen. Embryos homozygous for the anteater allele of Cse1l display a number of variable phenotypes, with craniofacial and ocular malformations being the most obvious. We provide evidence that Cse1l is the causal gene through complementation with a novel null allele of Cse1l generated by CRISPR-Cas9 editing. While the variability in the anteater phenotype was high enough to preclude a detailed molecular analysis, we demonstrate a very penetrant reduction in Pax6 levels in the developing eye along with significant ocular developmental phenotypes. The eye gene discovery tool iSyTE shows Cse1l to be significantly expressed in the lens from early eye development stages in embryos through adulthood. Cse1l has not previously been shown to be required for organogenesis as homozygosity for a null allele results in very early lethality. Future detailed studies of Cse1l function in craniofacial and neural development will be best served with a conditional allele to circumvent the variable phenotypes we report here. We suggest that human next-generation (whole genome or exome) sequencing studies yielding variants of unknown significance in CSE1L could consider these findings as part of variant analysis.

scholarly journals Abnormalities of the Genitourinary Tract in Female Mice Lacking GATA5

2000 ◽  
Vol 20 (14) ◽  
pp. 5256-5260 ◽  
Author(s):  
Jeffery D. Molkentin ◽  
Kevin M. Tymitz ◽  
James A. Richardson ◽  
Eric N. Olson
Keyword(s):  
Cell Fate ◽  
Null Allele ◽  
Cell Fate Specification ◽  
Mammalian Development ◽  
Gata Factor ◽  
Genitourinary System ◽  
Gata Factors ◽  
Developing Heart

ABSTRACT Members of the GATA family of transcription factors play important roles in cell fate specification, differentiation, and morphogenesis during mammalian development. GATA5, the only one of the six vertebrate GATA factor genes not yet inactivated in mice, is expressed in a pattern that overlaps with but is distinct from that of other GATA factor genes. During mouse embryogenesis, GATA5 is expressed first in the developing heart and subsequently in the lung, vasculature, and genitourinary system. To investigate the function of GATA5 in vivo, we created mice homozygous for a GATA5 null allele. Homozygous mutants were viable and fertile, but females exhibited pronounced genitourinary abnormalities that included vaginal and uterine defects and hypospadias. In contrast, the genitourinary system was unaffected in male GATA5 mutants. These results reveal a specific role of GATA5 in development of the female genitourinary system and suggest that other GATA factors may have functions overlapping those of GATA5 in other tissues.

scholarly journals Case Report: Infantile Urticaria as a Herald of Neonatal Onset Multisystem Inflammatory Disease With a Novel Mutation in NLRP3

2021 ◽  
Vol 12 ◽  
Author(s):  
Anna E. Patrick ◽  
Eden M. Lyons ◽  
Lisa Ishii ◽  
Alan S. Boyd ◽  
Joseph M. Choi ◽  
...  
Keyword(s):  
Clinical Symptoms ◽  
Early Recognition ◽  
Novel Mutation ◽  
Genetic Diagnosis ◽  
Inflammatory Disorder ◽  
Laboratory Findings ◽  
Genetic Sequencing ◽  
Variants Of Unknown Significance ◽  
Unknown Significance ◽  
The Impact

Neonatal multisystem onset inflammatory disorder (NOMID) is a severe autoinflammatory syndrome that can have an initial presentation as infantile urticaria. Thus, an immediate recognition of the clinical symptoms is essential for obtaining a genetic diagnosis and initiation of early therapies to prevent morbidity and mortality. Herein, we describe a neonate presenting with urticaria and systemic inflammation within hours after birth who developed arthropathy and neurologic findings. Pathologic evaluation of the skin revealed an infiltration of lymphocytes, eosinophils, and scattered neutrophils. Genetic analysis identified a novel heterozygous germline variant of unknown significance in the NLRP3 gene, causing the missense mutation M408T. Variants of unknown significance are common in genetic sequencing studies and are diagnostically challenging. Functional studies of the M408T variant demonstrated enhanced formation and activity of the NLRP3 inflammasome, with increased cleavage of the inflammatory cytokine IL-1β. Upon initiation of IL-1 pathway blockade, the infant had a robust response and improvement in clinical and laboratory findings. Our experimental data support that this novel variant in NLRP3 is causal for this infant’s diagnosis of NOMID. Rapid assessment of infantile urticaria with biopsy and genetic diagnosis led to early recognition and targeted anti-cytokine therapy. This observation expands the NOMID-causing variants in NLRP3 and underscores the role of genetic sequencing in rapidly identifying and treating autoinflammatory disease in infants. In addition, these findings highlight the importance of establishing the functional impact of variants of unknown significance, and the impact this knowledge may have on therapeutic decision making.

scholarly journals Small molecule inhibitors and a kinase-dead expressing mouse model demonstrate that the kinase activity of Chk1 is essential for mouse embryos and cancer cells

2020 ◽  
Author(s):  
Somsundar V Muralidharan ◽  
Lisa M Nilsson ◽  
Mattias F Lindberg ◽  
Jonas A Nilsson
Keyword(s):  
T Cells ◽  
Mouse Model ◽  
Cancer Cells ◽  
Null Allele ◽  
Protein Levels ◽  
Conditional Allele ◽  
Car T ◽  
High Level ◽  
Atr Kinase ◽  
Chk1 Kinase

AbstractChk1 kinase is downstream of the ATR kinase in the sensing of improper replication. Previous cell culture studies have demonstrated that Chk1 is essential for replication and Chk1 inhibitors are efficacious against tumors with high-level replication stress such as Myc-induced lymphoma cells. Treatment with Chk1 inhibitors also combines well with certain chemotherapeutic drugs and effects associates with induction of DNA damage and reduction of Chk1 protein levels. Most studies of Chk1 function has relied on the use of inhibitors. Whether or not a mouse or cancer cells could survive if a kinase-dead form of Chk1 is expressed has not been investigated before. Here we generate a mouse model that expresses a kinase-dead (D130A) allele in the mouse germline. We find that this mouse is overtly normal and does not have problems with erythropoiesis with ageing as previously been shown for a mouse expressing one null allele. However, similar to a null allele, homozygous kinase-dead mice cannot be generated and timed pregnancies of heterozygous mice suggest lethality of homozygous blastocysts at around the time of implantation. By breeding the kinase-dead Chk1 mouse with a conditional allele we are able to demonstrate that expression of only one kinase-dead allele, but no wildtype allele, of Chek1 is lethal for Myc-induced cancer cells. Finally, treatment of melanoma cells with tumor-infiltrating T cells or CAR-T cells is effective even if Chk1 is inhibited, suggesting that Chk1 inhibitors can be safely administered in patients where immunotherapy is an essential component of the arsenal against cancer.

scholarly journals Molecular Characterization of Chikungunya Virus Isolates From Two Localized Outbreaks During 2014-2019 in Kerala, India

2021 ◽  
Author(s):  
Anukumar Balakrishnan ◽  
Asia Devi Thounaojam ◽  
Aishwarya Babu ◽  
Jijo Koshy ◽  
Nikhil T L ◽  
...  
Keyword(s):  
South African ◽  
Chikungunya Virus ◽  
Novel Mutation ◽  
Indian Subcontinent ◽  
Virus Transmission ◽  
Amino Acid Position ◽  
East Central ◽  
Sequencing Studies ◽  
Central South

Abstract After the 2005-2009 chikungunya epidemic, intermittent outbreaks were reported in many parts of India. The outbreaks were caused by either locally circulating strains or imported viruses. Virus transmission route can be traced by complete genome sequencing studies. We investigated two outbreaks in the year 2014 and 2019 in Kerala, India. The chikungunya virus (CHIKV) was isolated from the samples and whole genome was sequenced for a 2014 isolate and a 2019 isolate. The phylogenetic tree revealed that the isolates formed a separate group with 2019 isolate from Pune, Maharashtra and belonged to the East/ Central/ South African (ECSA) genotype, Indian subcontinent sub lineage of Indian Ocean Lineage (IOL). A novel mutation at amino acid position 76 of E2 gene was observed in the group. The phylogenetic results suggest that the outbreaks might have caused by a virus, which has been circulating in India since 2014. Furthermore a detailed study is necessary to find out the evolution of CHIKV in India.

scholarly journals BMP7 functions predominantly as a heterodimer with BMP2 or BMP4 during mammalian embryogenesis

2019 ◽  
Author(s):  
Hyung-Seok Kim ◽  
Judith Neugebauer ◽  
Autumn McKnite ◽  
Anup Tilak ◽  
Jan L. Christian
Keyword(s):  
Body Wall ◽  
Null Allele ◽  
Congenital Defects ◽  
Mammalian Development ◽  
Proteolytic Activation ◽  
Embryonic Lethal ◽  
Mammalian Embryogenesis ◽  
Compound Heterozygotes

AbstractBMP7/BMP2 or BMP7/BMP4 heterodimers are more active than homodimers in vitro, but it is not known whether these heterodimers signal in vivo. To test this, we generated knock in mice carrying a mutation (Bmp7R-GFlag) that prevents proteolytic activation of the dimerized BMP7 precursor protein. This mutation eliminates the function of BMP7 homodimers and all other BMPs that normally heterodimerize with BMP7. While Bmp7 null homozygotes are live born, Bmp7R-GFlag homozygotes are embryonic lethal and have broadly reduced BMP activity. Furthermore, compound heterozygotes carrying the Bmp7R-G allele together with a null allele of Bmp2 or Bmp4 die during embryogenesis with defects in ventral body wall closure and/or the heart. Co-immunoprecipitation assays confirm that endogenous BMP4/7 heterodimers exist. Thus, BMP7 functions predominantly as a heterodimer with BMP2 or BMP4 during mammalian development, which may explain why mutations in either Bmp4 or Bmp7 lead to a similar spectrum of congenital defects in humans.

Three Kinships with ALAS2 P520L Mutations, Two in Association with Severe Iron Overload, and One with Sideroblastic Anemia and Severe Iron Overload.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3723-3723
Author(s):  
Pauline L. Lee ◽  
James C. Barton ◽  
Sreenivas V. Rao ◽  
Ronald T. Acton ◽  
Brian K. Adler ◽  
...  
Keyword(s):  
Iron Overload ◽  
Novel Mutation ◽  
White Male ◽  
Missense Mutations ◽  
Sideroblastic Anemia ◽  
Aminolevulinate Synthase ◽  
C282y Homozygote ◽  
Sequencing Studies ◽  
Unknown Gene ◽  
Distinctive Phenotype

Abstract Aminolevulinate synthase 2 (ALAS2) is an erythroid-expressed gene located on chromosome Xp11.21. Mutations in ALAS2 have been shown to be associated with sideroblastic anemia. We have identified a novel mutation in ALAS2, P520L, in three kinships. The P520L mutation was not found in 316 white male control subjects. The proline in this position is highly conserved across species from humans to zebrafish. In the C kinship, the P520L mutation was first identified in a white man who presented with severe iron overload at an early age and was a HFE C282Y homozygote. Genetic analyses of members of the C kinship suggest that the presence of P520L alone in hemizygous males, or simple heterozygosity in females, is not associated with anemia or iron overload. Females heterozygous for both HFE C282Y and ALAS2 P520L also had normal ferritin levels. Only subjects homozygous for HFE C282Y and hemizygous or heterozygous for ALAS2 P520L had severe iron overload. Sequencing studies revealed that the propositus did not have missense mutations in HAMP, HJV, FPN1, ABC7, IL6, or RAG1. In the H kinship, the propositus was a white woman with severe iron overload who was heterozygous for ALAS2 P520L; she had a wildtype HFE genotype. The pedigree of the H kinship identified 5 additional females who were heterozygous for the P520L ALAS2 mutation. Only the propositus had iron overload and none of the subjects with P520L had sideroblastic anemia. The propositus did not have missense mutations in FPN1, HAMP, HJV, TFR2, B2M, IRP2, ABC7, or SFT. We speculate that the propositus in the H kinship has a mutation in a currently unknown gene that contributes to her severe iron overload. In the S kinship, the P520L mutation was identified in a white man with sideroblastic anemia and severe iron overload. The patient had a second mutation in ALAS2, R560H, a mutation previously described in two brothers with sideroblastic anemia of variable penetrance (Blood100:4236–4238, 2002). It is not possible to determine the effect of the P520L mutation on the penetrance of the R560H-associated sideroblastic anemia and iron overload. We conclude that the ALAS2 P520L occurs at a very low allele frequency in the white population. The present observations also suggest that there is no distinctive phenotype associated with the P520L mutation alone, but that P520L may act as a modifier of iron overload in the presence of HFE homozygosity, other missense mutations of ALAS2, or mutations of uncharacterized iron regulatory genes.

scholarly journals EBV-Negative Monomorphic B-Cell Posttransplant Lymphoproliferative Disorder with Marked Morphologic Pleomorphism and Pathogenic Mutations in ASXL1, BCOR, CDKN2A, NF1, and TP53

2017 ◽  
Vol 2017 ◽  
pp. 1-8 ◽  
Author(s):  
Agata M. Bogusz
Keyword(s):  
B Cell ◽  
B Cell Lymphoma ◽  
Genetic Alterations ◽  
Chromosome 17 ◽  
Malignant Cells ◽  
Variants Of Unknown Significance ◽  
Barr Virus ◽  
Pathogenic Mutations ◽  
Sequencing Studies ◽  
Epstein Barr

Posttransplant lymphoproliferative disorders (PTLDs) are a diverse group of lymphoid or plasmacytic proliferations frequently driven by Epstein-Barr virus (EBV). EBV-negative PTLDs appear to represent a distinct entity. This report describes an unusual case of a 33-year-old woman that developed a monomorphic EBV-negative PTLD consistent with diffuse large B-cell lymphoma (DLBCL) 13 years after heart-lung transplant. Histological examination revealed marked pleomorphism of the malignant cells including nodular areas reminiscent of classical Hodgkin lymphoma (cHL) with abundant large, bizarre Hodgkin-like cells. By immunostaining, the malignant cells were immunoreactive for CD45, CD20, CD79a, PAX5, BCL6, MUM1, and p53 and negative for CD15, CD30, latent membrane protein 1 (LMP1), and EBV-encoded RNA (EBER). Flow cytometry demonstrated lambda light chain restricted CD5 and CD10 negative B-cells. Fluorescence in situ hybridization studies (FISH) were negative for cMYC, BCL2, and BCL6 rearrangements but showed deletion of TP53 and monosomy of chromosome 17. Next-generation sequencing studies (NGS) revealed numerous genetic alterations including 6 pathogenic mutations in ASXL1, BCOR, CDKN2A, NF1, and TP53(x2) genes and 30 variants of unknown significance (VOUS) in ABL1, ASXL1, ATM, BCOR, BCORL1, BRNIP3, CDH2, CDKN2A, DNMT3A, ETV6, EZH2, FBXW7, KIT, NF1, RUNX1, SETPB1, SF1, SMC1A, STAG2, TET2, TP53, and U2AF2.

scholarly journals BMP7 functions predominantly as a heterodimer with BMP2 or BMP4 during mammalian embryogenesis

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Hyung-Seok Kim ◽  
Judith Neugebauer ◽  
Autumn McKnite ◽  
Anup Tilak ◽  
Jan L Christian
Keyword(s):  
Body Wall ◽  
Null Allele ◽  
Congenital Defects ◽  
Mammalian Development ◽  
Proteolytic Activation ◽  
Embryonic Lethal ◽  
Mammalian Embryogenesis ◽  
Compound Heterozygotes

BMP7/BMP2 or BMP7/BMP4 heterodimers are more active than homodimers in vitro, but it is not known whether these heterodimers signal in vivo. To test this, we generated knock in mice carrying a mutation (Bmp7R-GFlag) that prevents proteolytic activation of the dimerized BMP7 precursor protein. This mutation eliminates the function of BMP7 homodimers and all other BMPs that normally heterodimerize with BMP7. While Bmp7 null homozygotes are live born, Bmp7R-GFlag homozygotes are embryonic lethal and have broadly reduced BMP activity. Furthermore, compound heterozygotes carrying the Bmp7R-G allele together with a null allele of Bmp2 or Bmp4 die during embryogenesis with defects in ventral body wall closure and/or the heart. Co-immunoprecipitation assays confirm that endogenous BMP4/7 heterodimers exist. Thus, BMP7 functions predominantly as a heterodimer with BMP2 or BMP4 during mammalian development, which may explain why mutations in either Bmp4 or Bmp7 lead to a similar spectrum of congenital defects in humans.

scholarly journals Genetic and molecular characterization of suppressors of SIR4 mutations in Saccharomyces cerevisiae.

Genetics ◽  
1989 ◽  
Vol 122 (1) ◽  
pp. 29-46 ◽  
Author(s):  
R Schnell ◽  
L D'Ari ◽  
M Foss ◽  
D Goodman ◽  
J Rine
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Null Allele ◽  
Loss Of Function ◽  
Induced Mutations ◽  
New Genes ◽  
Suppressor Mutations ◽  
Silencer Elements ◽  
Compensatory Mutations ◽  
The Stability

Abstract In order to learn more about other proteins that may be involved in repression of HML and HMR in Saccharomyces cerevisiae, extragenic suppressor mutations were identified that could restore repression in cells defective in SIR4, a gene required for function of the silencer elements flanking HML and HMR. These suppressor mutations, which define at least three new genes, SAN1, SAN2 and SAN3, arose at the frequency expected for loss-of-function mutations following mutagenesis. All san mutations were recessive. Suppression by san1 was allele-nonspecific, since san1 could suppress two very different alleles of SIR4, and was locus-specific since san1 was unable to suppress a SIR3 mutation or a variety of mutations conferring auxotrophies. The SAN1 gene was cloned, sequenced, and used to construct a null allele. The null allele had the same phenotype as the EMS-induced mutations and exhibited no pleiotropies of its own. Thus, the SAN1 gene was not essential. SAN1-mediated suppression was neither due to compensatory mutations in interacting proteins, nor to translational missense suppression. SAN1 may act posttranslationally to control the stability or activity of the SIR4 protein.

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