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Clinica Chimica Acta; International... Aug 2023Spinal muscular atrophy (SMA) is an autosomal recessive inherited neuromuscular condition caused by biallelic mutations in the survival of motor neuron 1 (SMN1) gene. A...
BACKGROUND AND AIMS
Spinal muscular atrophy (SMA) is an autosomal recessive inherited neuromuscular condition caused by biallelic mutations in the survival of motor neuron 1 (SMN1) gene. A homozygous deletion of the SMN1 gene accounts for approximately 95-98% of SMA patients. A highly homologous gene survival motor neuron 2 (SMN2) can partially compensate for SMN1 deletion, and its copy number is associated with disease severity. Population-based carrier screening by simultaneous quantification of SMN1 and SMN2 copy numbers is the best method to prevent SMA.
MATERIALS AND METHODS
In this study, a total of 516 samples were re-tested for the SMN1 copy number by using quantitative polymerase chain reaction (qPCR), multiplex ligation probe amplification (MLPA), droplet digital PCR (ddPCR), high-resolution melting (HRM) analysis, and PCR-based capillary electrophoresis (PCR/CE) simultaneously. Then, the performance of these methods was compared by using MLPA results as the reference.
RESULTS
The results of qPCR, ddPCR, HRM, and PCR/CE in detecting heterozygous deletion of SMN1 exon 7 and the results of ddPCR, HRM, and PCR/CE in detecting ≥2 copies of SMN1 exon7 are totally consistent with those of MLPA. The sensitivity and specificity of qPCR for detection of 2 copies of SMN1 exon 7 were 99.7% and 98.8%, respectively. The sensitivity and specificity of qPCR for detection of >2 copies of SMN1 exon 7 were 96.3% and 99.8%, respectively. Compared with the MLPA results, the sensitivity and specificity of qPCR and HRM for detection of heterozygous deletion of SMN1 exon 8 were 100% and 100%, respectively. They were 99.4% and 100%, respectively for detection of 2 copies, and 100% and 100%, respectively for detection of >2 copies. The results of PCR/CE in detecting SMN1 exon 8 were consistent with those of MLPA.
CONCLUSION
All these four methods show excellent performance in detecting heterozygous deletion of SMN1 exon 7. All PCR/CE results are totally concordant with those of MLPA. As the most cost-effective method, qPCR also shows high sensitivity and specificity in detecting SMN1. Taken together, our study provides useful information to select appropriate methods for SMA carrier screening.
Topics: Humans; Homozygote; Sequence Deletion; Muscular Atrophy, Spinal; Polymerase Chain Reaction; Exons; Survival of Motor Neuron 1 Protein
PubMed: 37479010
DOI: 10.1016/j.cca.2023.117496 -
Neurology India Jan 2024Chromosomal deletion and duplication syndromes can lead to intellectual disability, autism, microcephaly, and poor growth. Usually manifestations of duplication...
Chromosomal deletion and duplication syndromes can lead to intellectual disability, autism, microcephaly, and poor growth. Usually manifestations of duplication syndromes are milder than that of the deletion syndromes. With the availability of tests for analysis of copy number variants, it is possible to identify the deletion and duplication syndromes with greater ease. We report 32 cases of chromosomal duplication syndromes, identified in children presenting with developmental delay, intellectual disability, or microcephaly and/or additional features, at a tertiary care center on karyotyping or microarray analysis. Seven were isolated duplications, and one child had an additional smaller pathogenic deletion. Thus, duplication syndromes can have milder presentations with spectrum of dysmorphism, behavioral problems, and intellectual disability, but it is possible to diagnose easily with latest emerging high-throughput technologies.
Topics: Child; Humans; Chromosome Duplication; Microcephaly; Intellectual Disability; Research; Chromosome Deletion; Syndrome
PubMed: 38443013
DOI: 10.4103/ni.ni_400_21 -
BMC Ecology and Evolution Sep 2023No phenotypic trait evolves independently of all other traits, but the cause of trait-trait coevolution is poorly understood. While the coevolution could arise simply...
No phenotypic trait evolves independently of all other traits, but the cause of trait-trait coevolution is poorly understood. While the coevolution could arise simply from pleiotropic mutations that simultaneously affect the traits concerned, it could also result from multivariate natural selection favoring certain trait relationships. To gain a general mechanistic understanding of trait-trait coevolution, we examine the evolution of 220 cell morphology traits across 16 natural strains of the yeast Saccharomyces cerevisiae and the evolution of 24 wing morphology traits across 110 fly species of the family Drosophilidae, along with the variations of these traits among gene deletion or mutation accumulation lines (a.k.a. mutants). For numerous trait pairs, the phenotypic correlation among evolutionary lineages differs significantly from that among mutants. Specifically, we find hundreds of cases where the evolutionary correlation between traits is strengthened or reversed relative to the mutational correlation, which, according to our population genetic simulation, is likely caused by multivariate selection. Furthermore, we detect selection for enhanced modularity of the yeast traits analyzed. Together, these results demonstrate that trait-trait coevolution is shaped by natural selection and suggest that the pleiotropic structure of mutation is not optimal. Because the morphological traits analyzed here are chosen largely because of their measurability and thereby are not expected to be biased with regard to natural selection, our conclusion is likely general.
Topics: Animals; Saccharomyces cerevisiae; Computer Simulation; Drosophilidae; Gene Deletion; Mutation
PubMed: 37700252
DOI: 10.1186/s12862-023-02164-4 -
Blood Advances Jul 2023TP53 aberrations, including mutations and deletion of 17p13, are important adverse prognostic markers in chronic lymphocytic leukemia (CLL) but are less studied in high...
TP53 aberrations, including mutations and deletion of 17p13, are important adverse prognostic markers in chronic lymphocytic leukemia (CLL) but are less studied in high count monoclonal B-cell lymphocytosis (HCMBL), an asymptomatic pre-malignant stage of CLL. Here we estimated the prevalence and impact of TP53 aberrations in 1,230 newly diagnosed treatment-naïve individuals (849 CLL, 381 HCMBL). We defined TP53 state as: wild-type (no TP53 mutations and normal 17p), single-hit (del(17p) or one TP53 mutation), or multi-hit (TP53 mutation and del(17p), TP53 mutation and loss of heterozygosity, or multiple TP53 mutations). Cox regression was used to estimate hazard ratios (HR) and 95% confidence intervals (CI) for time to first treatment and overall survival by TP53 state. We found 64 (7.5%) CLL patients and 17 (4.5%) HCMBL individuals had TP53 mutations with variant allele fraction >10%. Del(17p) was present in 58 (6.8%) of CLL and 11 (2.9%) of HCMBL cases. Most individuals had wild-type (N=1,128, 91.7%) TP53 state, followed by multi-hit (N=55, 4.5%) and then single-hit (N=47, 3.8%) TP53 state. The risk of shorter time to therapy and death increased with the number of TP53 abnormalities. Compared to wild-type patients, multi-hit patients had 3-fold and single-hit patients had 1.5-fold increased risk of requiring therapy. Multi-hit patients also had 2.9-fold increased risk of death compared to wild-type. These results remained stable after accounting for other known poor prognostic factors. Both TP53 mutations and del(17p) may provide important prognostic information for HCMBL and CLL that would be missed if only one were measured.
Topics: Humans; Leukemia, Lymphocytic, Chronic, B-Cell; Tumor Suppressor Protein p53; Prognosis; Mutation; Chromosome Deletion
PubMed: 36877634
DOI: 10.1182/bloodadvances.2022009040 -
American Journal of Medical Genetics.... Nov 20233q29 deletion syndrome (3q29del) is a rare genomic disorder caused by a 1.6 Mb deletion (hg19, chr3:195725000-197350000). 3q29del is associated with neurodevelopmental...
3q29 deletion syndrome (3q29del) is a rare genomic disorder caused by a 1.6 Mb deletion (hg19, chr3:195725000-197350000). 3q29del is associated with neurodevelopmental and psychiatric phenotypes, including an astonishing >40-fold increased risk for schizophrenia, but medical phenotypes are less well-described. We used the online 3q29 registry of 206 individuals (3q29deletion.org) to recruit 57 individuals with 3q29del (56.14% male) and requested information about musculoskeletal phenotypes with a custom questionnaire. 85.96% of participants with 3q29del reported at least one musculoskeletal phenotype. Congenital anomalies were most common (70.18%), with pes planus (40.35%), pectus excavatum (22.81%), and pectus carinatum (5.26%) significantly elevated relative to the pediatric general population. 49.12% of participants reported fatigue after 30 min or less of activity. Bone fractures (8.77%) were significantly elevated relative to the pediatric general population. Participants commonly report receiving medical care for musculoskeletal complaints (71.93%), indicating that these phenotypes impact quality of life for individuals with 3q29del. This is the most comprehensive description of musculoskeletal phenotypes in 3q29del to date, suggests ideas for clinical evaluation, and expands our understanding of the phenotypic spectrum of this syndrome.
Topics: Humans; Child; Male; Female; Developmental Disabilities; Chromosome Deletion; Quality of Life; Intellectual Disability; Phenotype; Syndrome
PubMed: 37691301
DOI: 10.1002/ajmg.a.63384 -
Journal of Virology Sep 2023Porcine epidemic diarrhea virus (PEDV) leads to enormous economic losses for the pork industry. However, the commercial vaccines failed to fully protect against the...
Porcine epidemic diarrhea virus (PEDV) leads to enormous economic losses for the pork industry. However, the commercial vaccines failed to fully protect against the epidemic strains. Previously, the rCH/SX/2016-S strain with the entire E protein and the rCH/SX/2015 strain with the deletion of 7-amino-acid (7-aa) at positions 23-29 in E protein were constructed and rescued. The pathogenicity assay indicated that rCH/SX/2015 is an attenuated strain, but rCH/SX/2016-S belongs to the virulent strains. Then, the recombination PEDV (rPEDV-E)strain with a 7-aa deletion in the E protein was generated, using the highly virulent rCH/SX/2016-S strain (rPEDV-E) as the backbone. Compared with the rPEDV-E strain, the release and infectivity of the rPEDV-E strain were significantly reduced , but stronger interferon (IFN) responses were triggered both and . The pathogenicity assay showed that the parental strain resulted in severe diarrhea (100%) and death (100%) in all piglets. Compared with the parental strain group, rPEDV-E caused lower mortality (33%) and diminished fecal PEDV RNA shedding. At 21 days, all surviving pigs were challenged orally with rPEDV-E. No pigs died in the two groups. Compared with the mock group, significantly delayed and milder diarrhea and reduced fecal PEDV RNA shedding were detected in the rPEDV-E group. In conclusion, the deletion of a 7-aa fragment in the E protein (E) attenuated PEDV but retained its immunogenicity, which can offer new ideas for the design of live attenuated vaccines and provide new insights into the attenuated mechanism of PEDV. IMPORTANCE Porcine epidemic diarrhea virus (PEDV) causes high mortality in neonatal piglets and remains a large challenge to the pork industry. Unfortunately, no safe and effective vaccines are available yet. The pathogenesis and molecular basis of the attenuation of PEDV remain unclear, which seriously hinders the development of PEDV vaccines. This study found that the rPEDV carrying E mutation in the E protein induced significantly higher IFN responses than the parental virus, partially attenuated, and remained immunogenic in piglets. For the first time, PEDV E was verified as an IFN antagonist in the infection context and identified as a virulence factor of PEDV. Our data also suggested that E mutation can be a good target for the development of live attenuated vaccines for PEDV and also provide new perspectives for the attenuated mechanism of PEDV.
Topics: Animals; Coronavirus Infections; Interferons; Porcine epidemic diarrhea virus; RNA; Swine; Swine Diseases; Vaccines, Attenuated; Sequence Deletion; Viral Envelope Proteins
PubMed: 37681956
DOI: 10.1128/jvi.00847-23 -
Science Advances Dec 2023Large-scale chromosomal aberrations are prevalent in human cancer, but their function remains poorly understood. We established chromosome-engineered hepatocellular...
Large-scale chromosomal aberrations are prevalent in human cancer, but their function remains poorly understood. We established chromosome-engineered hepatocellular carcinoma cell lines using CRISPR-Cas9 genome editing. A 33-mega-base pair region on chromosome 8p (chr8p) was heterozygously deleted, mimicking a frequently observed chromosomal deletion. Using this isogenic model system, we delineated the functional consequences of chr8p loss and its impact on metastatic behavior and patient survival. We found that metastasis-associated genes on chr8p act in concert to induce an aggressive and invasive phenotype characteristic for chr8p-deleted tumors. Genome-wide CRISPR-Cas9 viability screening in isogenic chr8p-deleted cells served as a powerful tool to find previously unidentified synthetic lethal targets and vulnerabilities accompanying patient-specific chromosomal alterations. Using this target identification strategy, we showed that chr8p deletion sensitizes tumor cells to targeting of the reactive oxygen sanitizing enzyme Nudix hydrolase 17. Thus, chromosomal engineering allowed for the identification of novel synthetic lethalities specific to chr8p loss of heterozygosity.
Topics: Humans; Synthetic Lethal Mutations; Liver Neoplasms; Chromosome Deletion; Chromosome Aberrations; Chromosomes; CRISPR-Cas Systems
PubMed: 38134284
DOI: 10.1126/sciadv.adh1442 -
Translational Pediatrics Aug 2023Primary ciliary dyskinesia (PCD) is a clinically heterogeneous group of autosomal or, less frequently, X-chromosomal recessive inheritance syndrome of motile cilia...
BACKGROUND
Primary ciliary dyskinesia (PCD) is a clinically heterogeneous group of autosomal or, less frequently, X-chromosomal recessive inheritance syndrome of motile cilia dysfunction characterized by neonatal respiratory distress, oto-sino-pulmonary disease, infertility and situs inversus. Recently, type 43 PCD (CILD43, OMIM#618699) was established by autosomal-dominant loss-of-function mutations identified in Forkhead box J1 (). However, the functional validation of mutations in humans and mice has not been fully performed. Here we studied a three-generation family with heterotaxy and proband with complex congenital heart disease (CHD).
METHODS
We performed whole-exome sequencing to investigate the causative variant of this family and generated gene knock-in mice carrying the human equivalent mutation by homologous recombination. Then, microscopy analysis was used to characterize the phenotype and ciliary ultrastructure of the model. Effects of the variant on heart anomaly were preliminarily explored through transcriptome sequencing.
RESULTS
A novel heterozygous deletion variant (c.1129delC/p.Leu377Trpfs*76) of was discovered that exerts a dominant-negative effect (DNE) . Notably, both homozygous () and heterozygous () mice developed situs inversus, hydrocephalus and showed a disruption of trachea cilia structure, whereas these abnormalities were only observed in previously reported , not mice. Thus, a more severe phenotype and higher expressivity of our mouse model further indicated the DNE of this mutation. Meanwhile, several cardiomyopathy-related genes were differentially expressed in the homozygous knock-in mouse hearts, pointing to a probable function in cardiac pathology.
CONCLUSIONS
Overall, our study results showed that c.1129delC mutation in was regarded as the cause of situs inversus in this family and this mutant showed a capacity of DNE over wild-type FOXJ1, causing more serious consequences than the allelic deletion of .
PubMed: 37692537
DOI: 10.21037/tp-23-27 -
Clinical Pediatrics Dec 2023Homozygous deletion of can lead to isolated nephronophthisis (NPHP) and syndromic disorders. However, the phenotype of scalp tumor and hydroureteronephrosis in NPHP...
Homozygous deletion of can lead to isolated nephronophthisis (NPHP) and syndromic disorders. However, the phenotype of scalp tumor and hydroureteronephrosis in NPHP patients with homozygous deletion of has not been reported. Clinical data, laboratory results, and genetic testing of 4 NPHP patients were collected. Examination of their eyes, heart, and urinary tract and of their hepatobiliary, skeletal, and central nervous systems was evaluated. Isolated NPHP was observed in 1 case, and syndromic disorders were observed in the other 3 patients. Their syndromic disorders showed NPHP combined with central nervous system defects, eye involvement, scalp tumor, arachnoid cyst, or hydroureteronephrosis. Large homozygous deletions covering the whole gene locus were identified in all 4 patients. We report a novel phenotype of scalp tumor and hydroureteronephrosis in NPHP patients with homozygous deletion of , paving an avenue for further research on -associated deformity in the skin and the urinary system.
Topics: Humans; Homozygote; Kidney Failure, Chronic; Membrane Proteins; Cytoskeletal Proteins; Scalp; Sequence Deletion; Adaptor Proteins, Signal Transducing; Kidney Diseases, Cystic; Neoplasms
PubMed: 36942623
DOI: 10.1177/00099228231162416 -
Nature Communications Sep 2023Molecular screens comparing different disease states to identify candidate genes rely on the availability of fast, reliable and multiplexable systems to interrogate...
Molecular screens comparing different disease states to identify candidate genes rely on the availability of fast, reliable and multiplexable systems to interrogate genes of interest. CRISPR/Cas9-based reverse genetics is a promising method to eventually achieve this. However, such methods are sorely lacking for multi-nucleated muscle fibers, since highly efficient nuclei editing is a requisite to robustly inactive candidate genes. Here, we couple Cre-mediated skeletal muscle fiber-specific Cas9 expression with myotropic adeno-associated virus-mediated sgRNA delivery to establish a system for highly effective somatic gene deletions in mice. Using well-characterized genes, we show that local or systemic inactivation of these genes copy the phenotype of traditional gene-knockout mouse models. Thus, this proof-of-principle study establishes a method to unravel the function of individual genes or entire signaling pathways in adult skeletal muscle fibers without the cumbersome requirement of generating knockout mice.
Topics: Mice; Animals; CRISPR-Cas Systems; Gene Editing; Gene Deletion; RNA, Guide, CRISPR-Cas Systems; Mice, Knockout; Muscle Fibers, Skeletal
PubMed: 37777530
DOI: 10.1038/s41467-023-41769-7