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1.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 37(12): 1371-1375, 2020 Dec 10.
Artigo em Chinês | MEDLINE | ID: mdl-33306825

RESUMO

OBJECTIVE: To explore the genetic basis for an infant with neonatal diabetes (NDM) and multiple malformations. METHODS: Genetic variants were detected by next generation sequencing (NGS). Suspected variant was verified by Sanger sequencing. RESULTS: A de novo heterozygous variant, c.1454_1455del(p.K485Rfs), was detected in exon 5 of the GATA6 gene. The variant was undetected in his parents and unreported previously. Bioinformatic analysis predicted the variant to be pathogenic. CONCLUSION: The heterozygous variant of c.1454_1455del(p.K485Rfs) of the GATA6 gene probably underlies the disease in this child. Genetic testing can facilitate diagnosis and genetic counseling for NDM.


Assuntos
Anormalidades Múltiplas , Diabetes Mellitus , Deleção de Sequência , Adulto , Diabetes Mellitus/genética , Feminino , Testes Genéticos , Heterozigoto , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Recém-Nascido , Masculino , Deleção de Sequência/genética
2.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 37(12): 1387-1390, 2020 Dec 10.
Artigo em Chinês | MEDLINE | ID: mdl-33306829

RESUMO

OBJECTIVE: To carry out prenatal diagnose for a fetus with ultrasonography abnormalities using multiple genetic techniques. METHODS: Routine G-banding chromosomal analysis and single nucleotide polymorphism array (SNP-array) were applied in conjunction for the prenatal diagnosis of the fetus. The result was confirmed by fluorescence in situ hybridization (FISH). RESULTS: SNP-array detected that the fetus has carried a hemizygous 5.1 Mb deletion at 22q13.31q13.33, which is associated with Phelan-McDermid syndrome, and a hemizygous 4.5 Mb deletion at 21q21.1q21.2. FISH analysis of the fetus and its parents suggested that both deletions were de novo in origin. CONCLUSION: The hemizygous deletions on 21q21.1q21.2 and 22q13.31q13.33 probably underlay the abnormal phenotype of the fetus. Genetic analysis can provide crucial information for the prenatal diagnosis and genetic counseling.


Assuntos
Diagnóstico Pré-Natal , Deleção de Sequência , Deleção Cromossômica , Transtornos Cromossômicos/genética , Cromossomos Humanos Par 21/genética , Cromossomos Humanos Par 22/genética , Feminino , Feto , Humanos , Hibridização in Situ Fluorescente , Masculino , Polimorfismo de Nucleotídeo Único , Gravidez , Deleção de Sequência/genética
3.
Sci Rep ; 10(1): 14108, 2020 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-32839485

RESUMO

Selection pressures exerted on Staphylococcus aureus by host factors during infection may lead to the emergence of regulatory phenotypes better adapted to the infection site. Traits convenient for persistence may be fixed by mutation thus turning these mutants into microevolution endpoints. The feasibility that stable, non-encapsulated S. aureus mutants can regain expression of key virulence factors for survival in the bloodstream was investigated. S. aureus agr mutant HU-14 (IS256 insertion in agrC) from a patient with chronic osteomyelitis was passed through the bloodstream using a bacteriemia mouse model and derivative P3.1 was obtained. Although IS256 remained inserted in agrC, P3.1 regained production of capsular polysaccharide type 5 (CP5) and staphyloxanthin. Furthermore, P3.1 expressed higher levels of asp23/SigB when compared with parental strain HU-14. Strain P3.1 displayed decreased osteoclastogenesis capacity, thus indicating decreased adaptability to bone compared with strain HU-14 and exhibited a trend to be more virulent than parental strain HU-14. Strain P3.1 exhibited the loss of one IS256 copy, which was originally located in the HU-14 noncoding region between dnaG (DNA primase) and rpoD (sigA). This loss may be associated with the observed phenotype change but the mechanism remains unknown. In conclusion, S. aureus organisms that escape the infected bone may recover the expression of key virulence factors through a rapid microevolution pathway involving SigB regulation of key virulence factors.


Assuntos
Cápsulas Bacterianas/metabolismo , Proteínas de Bactérias/genética , Staphylococcus aureus/genética , Transativadores/genética , Xantofilas/metabolismo , Adulto , Animais , Antibacterianos/farmacologia , Bacteriemia/microbiologia , Cápsulas Bacterianas/genética , Modelos Animais de Doenças , Farmacorresistência Bacteriana Múltipla/genética , Regulação Bacteriana da Expressão Gênica/genética , Humanos , Masculino , Camundongos , Osteomielite/microbiologia , Deleção de Sequência/genética , Staphylococcus aureus/isolamento & purificação , Staphylococcus aureus/patogenicidade , Virulência/genética , Fatores de Virulência/genética
4.
Sci Rep ; 10(1): 13757, 2020 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-32792587

RESUMO

Nighttime surges in melatonin levels activate melatonin receptors, which synchronize cellular activities with the natural light/dark cycle. Melatonin receptors are expressed in several cell types in the retina, including the photon-sensitive rods and cones. Previous studies suggest that long-term photoreceptor survival and retinal health is in part reliant on melatonin orchestration of circadian homeostatic activities. This scenario would accordingly envisage that disruption of melatonin receptor signaling is detrimental to photoreceptor health. Using in vivo CRISPR/Cas9 genomic editing, we discovered that a small deletion mutation of the Mel1a melatonin receptor (mtnr1a) gene causes a loss of rod photoreceptors in retinas of developing Xenopus tropicalis heterozygous, but not homozygous mutant tadpoles. Cones were relatively spared from degeneration, and the rod loss phenotype was not obvious after metamorphosis. Localization of Mel1a receptor protein appeared to be about the same in wild type and mutant retinas, suggesting that the mutant protein is expressed at some level in mutant retinal cells. The severe impact on early rod photoreceptor viability may signify a previously underestimated critical role in circadian influences on long-term retinal health and preservation of sight. These data offer evidence that disturbance of homeostatic, circadian signaling, conveyed through a mutated melatonin receptor, is incompatible with rod photoreceptor survival.


Assuntos
Receptores de Melatonina/genética , Degeneração Retiniana/genética , Células Fotorreceptoras Retinianas Bastonetes/patologia , Proteínas de Xenopus/genética , Xenopus/genética , Sequência de Aminoácidos , Animais , Animais Geneticamente Modificados , Sistemas CRISPR-Cas , Ritmo Circadiano/genética , Mutação da Fase de Leitura/genética , Melatonina/metabolismo , Células Fotorreceptoras Retinianas Cones/patologia , Deleção de Sequência/genética , Transdução de Sinais/genética
5.
Medicine (Baltimore) ; 99(22): e20413, 2020 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-32481435

RESUMO

BACKGROUND: Autosomal recessive polycystic kidney disease (ARPKD) is an autosomal recessive hepatorenal fibrocystic syndrome. The majority of ARPKD patients progress to end-stage renal disease. Precise molecular diagnosis of ARPKD has proven valuable for understanding its mechanism and selecting optimal therapy. METHODS: A Chinese family with ARPKD was recruited in current study. The clinical characteristics of ARPKD patient were collected from medical records and the potential responsible genes were studied by the whole exome sequencing (WES). Candidate pathogenic variants were validated by Sanger sequencing. RESULTS: Both renal manifestation and hepatobiliary phenotype were observed. WES revealed compound heterozygous mutations of polycystic kidney and hepatic disease 1 genes, NM_138694: c.751G>T, (p.Asp251Tyr) and c.3998_4004delACCTGAA (p.Asn1333Thr fs × 13), which were confirmed by Sanger sequencing. Moreover, the mutations in the proband and its affected sib were co-segregated with the phenotype. CONCLUSIONS: The novel mutation in polycystic kidney and hepatic disease 1 gene identified by WES might be molecular pathogenic basis of this disorder.


Assuntos
Rim Policístico Autossômico Recessivo/genética , Sequenciamento Completo do Exoma , Proteínas Adaptadoras de Transdução de Sinal/genética , Adulto , Criança , China , Feminino , Predisposição Genética para Doença/genética , Humanos , Mutação de Sentido Incorreto/genética , Receptores de Superfície Celular/genética , Deleção de Sequência/genética , Sequenciamento Completo do Exoma/métodos
6.
Cytogenet Genome Res ; 160(5): 255-263, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32544910

RESUMO

Fusions of the Runt-related transcription factor 1 (RUNX1) with different partner genes have been associated with various hematological disorders. Interestingly, the C-terminally truncated form of RUNX1 and RUNX1 fusion proteins are similarly considered important contributors to leukemogenesis. Here, we describe a 59-year-old male patient who was initially diagnosed with acute myeloid leukemia, inv(16)(p13;q22)/CBFB-MYH11 (FAB classification M4Eo). He achieved complete remission and negative CBFB-MYH11 status with daunorubicin/cytarabine combination chemotherapy but relapsed 3 years later. Cytogenetic analysis of relapsed leukemia cells revealed CBFB-MYH11 negativity and complex chromosomal abnormalities without inv(16)(p13;q22). RNA-seq identified the glutamate receptor, ionotropic, kinase 2 (GRIK2) gene on 6q16 as a novel fusion partner for RUNX1 in this case. Specifically, the fusion of RUNX1 to the GRIK2 antisense strand (RUNX1-GRIK2as) generated multiple missplicing transcripts. Because extremely low levels of wild-type GRIK2 were detected in leukemia cells, RUNX1-GRIK2as was thought to drive the pathogenesis associated with the RUNX1-GRIK2 fusion. The truncated RUNX1 generated from RUNX1-GRIK2as induced the expression of the granulocyte colony-stimulating factor (G-CSF) receptor on 32D myeloid leukemia cells and enhanced proliferation in response to G-CSF. In summary, the RUNX1-GRIK2as fusion emphasizes the importance of aberrantly truncated RUNX1 in leukemogenesis.


Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/genética , DNA Antissenso/genética , Fusão Gênica/genética , Fator Estimulador de Colônias de Granulócitos/farmacologia , Leucemia Mieloide Aguda/genética , Receptores de Ácido Caínico/genética , Deleção de Sequência/genética , Translocação Genética/genética , Proliferação de Células/efeitos dos fármacos , Humanos , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , RNA Mensageiro/análise , RNA Mensageiro/genética , Receptores de Fator Estimulador de Colônias de Granulócitos/biossíntese , Receptores de Fator Estimulador de Colônias de Granulócitos/metabolismo
7.
Hum Genet ; 139(11): 1417-1427, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32488466

RESUMO

An inverted duplication with a terminal deletion (inv-dup-del) is one of the complex constitutional structural rearrangements that can occur in a chromosome. Although breakages of dicentric chromosome have been suggested, the precise mechanism of this is yet to be fully understood. In our present study, we investigated the genomic structure of 10 inv-dup-del cases to elucidate this mechanism. Two recurrent 8p inv-dup-del cases harbored a large copy-number-neutral region between the duplication and deletion in common. Although the other non-recurrent cases did not appear to have this copy-number-neutral region, refined sequencing analysis identified that they contained a small intervening region at the junction between the inverted and non-inverted segment. The size of this small intervening region ranged from 1741 to 3728 bp. Combined with a presence of microhomology at the junction, a resolution of the replication fork stalling through template switching within the same replication fork is suggested. We further observed two cases with mosaicism of the dicentric chromosome and various structural rearrangements related to the dicentric chromosome. Refined analysis allowed us to identify different breakpoints on the same chromosome in the same case, implicating multiple rounds of U-type formation and its breakage. From these results, we propose that a replication-based mechanism generates unstable dicentric chromosomes and that their breakage leads to the formation of inv-dup-dels and other related derivative chromosomes.


Assuntos
Transtornos Cromossômicos/genética , Inversão Cromossômica/genética , Cromossomos/genética , Duplicação Gênica/genética , Deleção de Sequência/genética , Deleção Cromossômica , Replicação do DNA/genética , Humanos , Mosaicismo
9.
Mol Genet Genomics ; 295(4): 923-931, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32246227

RESUMO

Amelogenesis imperfecta type IG (AI1G) is caused by mutations in FAM20A. Genotypic and phenotypic features of AI1G are diverse and their full spectra remain to be characterized. The aim of this study was to identify and summarize variants in FAM20A in a broad population of patients with AI1G. We identified a Thai female (Pt-1) and a Saudi male (Pt-2) affected with AI1G. Both had hypoplastic enamel, gingival hyperplasia, and intrapulpal calcification. Pt-1 also had rapidly progressive embedding of unerupted teeth, early eruption of permanent teeth, and spontaneous dental infection. Uniquely, Pt-2 had all permanent teeth erupted which was uncommon in AI1G patients. Whole exome sequencing (WES) identified that Pt-1 was heterozygous for FAM20A, c.758A > G (p.Tyr253Cys), inherited from her father. The mutation on maternal allele was not detected by WES. Pt-2 possessed compound heterozygous mutations, c.1248dupG (p.Phe417Valfs*7); c.1081C > T (p.Arg361Cys) in FAM20A. Array comparative genomic hybridization (aCGH), cDNA sequencing, and whole genome sequencing successfully identified 7531 bp deletion on Pt-1's maternal allele. This was the largest FAM20A deletion ever found. A review of all 70 patients from 50 independent families with AI1G (including two families in this study) showed that the penetrance of hypoplastic enamel and gingival hyperplasia was complete. Unerupted permanent teeth were found in all 70 patients except Pt-2. Exons 1 and 11 were mutation-prone. Most mutations were frameshift. Certain variants showed founder effect. To conclude, this study reviews and expands phenotypic and genotypic spectra of AI1G. A large deletion missed by WES can be detected by WGS. Hypoplastic enamel, gingival hyperplasia, and unerupted permanent teeth prompt genetic testing of FAM20A. Screening of nephrocalcinosis, early removal of embedded teeth, and monitoring of dental infection are recommended.


Assuntos
Amelogênese Imperfeita/genética , Proteínas do Esmalte Dentário/genética , Nefrocalcinose/genética , Deleção de Sequência/genética , Adolescente , Adulto , Amelogênese Imperfeita/patologia , Criança , Hibridização Genômica Comparativa , Éxons/genética , Feminino , Mutação da Fase de Leitura/genética , Genótipo , Heterozigoto , Homozigoto , Humanos , Masculino , Mutação/genética , Nefrocalcinose/patologia , Linhagem , Fenótipo
10.
J Biol Chem ; 295(20): 6926-6935, 2020 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-32249210

RESUMO

Mouse hepatitis virus (MHV; murine coronavirus) causes meningoencephalitis, myelitis, and optic neuritis followed by axonal loss and demyelination. This murine virus is used as a common model to study acute and chronic virus-induced demyelination in the central nervous system. Studies with recombinant MHV strains that differ in the gene encoding the spike protein have demonstrated that the spike has a role in MHV pathogenesis and retrograde axonal transport. Fusion peptides (FPs) in the spike protein play a key role in MHV pathogenesis. In a previous study of the effect of deleting a single proline residue in the FP of a demyelinating MHV strain, we found that two central, consecutive prolines are important for cell-cell fusion and pathogenesis. The dihedral fluctuation of the FP was shown to be repressed whenever two consecutive prolines were present, in contrast to the presence of a single proline in the chain. Using this proline-deleted MHV strain, here we investigated whether intracranial injection of this strain can induce optic neuritis by retrograde axonal transport from the brain to the retina through the optic nerve. We observed that the proline-deleted recombinant MHV strain is restricted to the optic nerve, is unable to translocate to the retina, and causes only minimal demyelination and no neuronal death. We conclude that an intact proline dyad in the FP of the recombinant demyelinating MHV strain plays a crucial role in translocation of the virus through axons and subsequent neurodegeneration.


Assuntos
Transporte Axonal/genética , Vírus da Hepatite Murina/genética , Glicoproteína da Espícula de Coronavírus/metabolismo , Animais , Transporte Axonal/fisiologia , Axônios/metabolismo , Axônios/virologia , Encéfalo/metabolismo , Infecções por Coronavirus/patologia , Doenças Desmielinizantes/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Vírus da Hepatite Murina/metabolismo , Nervo Óptico/metabolismo , Nervo Óptico/virologia , Peptídeos/metabolismo , Prolina/metabolismo , Deleção de Sequência/genética , Glicoproteína da Espícula de Coronavírus/genética , Proteínas do Envelope Viral/metabolismo
11.
BMC Med Genet ; 21(1): 77, 2020 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-32293312

RESUMO

BACKGROUND: Myopathy with extrapyramidal signs (MPXPS) is an autosomal recessive mitochondrial disorder which is caused by mutation in mitochondrial calcium uptake 1 (MICU1) gene located on chromosome 10q22.1. Next Generation Sequencing (NGS) technology is the most effective method for identification of pathogenic variants with the ability to overcome some limitations which Sanger sequencing may encountered. There are few reports on this rare disease around the world and here in this study we first revealed genetic identification of two affected individuals in an Iranian family with a novel mutation. CASE PRESENTATION: The proband was a 5-year-old girl from consanguenous parents. She was first clinically suspicious of affected with limb-girdle muscular dystrophy (LGMD). Muscle biopsy studies and autozygosity mapping, using four short tandem repeat (STR) markers linked to 6 genes of the most prevalent forms of LGMD, ruled out calpainopathy, dysferlinopathy, and sarcoglycanopathis. DNA sample of the proband was sent for NGS. Whole exome sequencing (WES) revealed a novel mutation c.1295delA in exon 13 of MICU1 gene. This homozygous deletion creates a frameshift and a premature stop codon downstream of canonical EF4 calcium binding motif of MICU1. According to the American College of Medical Genetics and Genomics (ACMG) guidline for sequence interpretation, this variant was a pathogenic one. Sanger sequencing in all family members confirmed the results of the WES. CONCLUSIONS: This study was the first report of MPXPS in Iranian population which also revealed a novel mutation in the MICU1 gene.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Proteínas de Transporte de Cátions/genética , Repetições de Microssatélites/genética , Proteínas de Transporte da Membrana Mitocondrial/genética , Doenças Musculares/genética , Distrofia Muscular do Cíngulo dos Membros/genética , Pré-Escolar , Exoma/genética , Éxons/genética , Tratos Extrapiramidais/metabolismo , Tratos Extrapiramidais/patologia , Feminino , Mutação da Fase de Leitura/genética , Sequenciamento de Nucleotídeos em Larga Escala , Homozigoto , Humanos , Irã (Geográfico)/epidemiologia , Doenças Musculares/patologia , Distrofia Muscular do Cíngulo dos Membros/patologia , Linhagem , Deleção de Sequência/genética , Sequenciamento Completo do Exoma
12.
BMC Med Genet ; 21(1): 84, 2020 04 19.
Artigo em Inglês | MEDLINE | ID: mdl-32306954

RESUMO

BACKGROUND: Nephronophthisis (NPHP) is a rare autosomal recessive inherited disorder with high heterogeneity. The majority of NPHP patients progress to end-stage renal disease (ESRD) within the first three decades of life. As an inherited disorder with highly genetic heterogeneity and clinical presentations, NPHP still poses a challenging task for nephrologists without special training to make a well-judged decision on its precise diagnosis, let alone its mechanism and optimal therapy. CASE PRESENTATION: A Chinese family with NPHP was recruited in current study. The clinical characteristics (including findings from renal biopsy) of NPHP patients were collected from medical records and the potential responsible genes were explored by the whole exome sequencing (WES). A homozygous deletion of NPHP1 (1-20 exons) was found in both affected patients, which was further confirmed by quantitative PCR. CONCLUSIONS: Homozygous full gene deletion of the NPHP1 gene was identified in a Chinese family with NPHP, which was the molecular pathogenic basis of this disorder. Furthermore, identification of the pathogenic genes for those affected patients can help to have a full knowledge on NPHP's molecular mechanism and precise treatment.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas do Citoesqueleto/genética , Predisposição Genética para Doença , Doenças Renais Císticas/congênito , Falência Renal Crônica/genética , Adulto , Éxons/genética , Feminino , Deleção de Genes , Homozigoto , Humanos , Doenças Renais Císticas/genética , Doenças Renais Císticas/fisiopatologia , Falência Renal Crônica/fisiopatologia , Masculino , Linhagem , Deleção de Sequência/genética , Sequenciamento Completo do Exoma
13.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 37(5): 559-562, 2020 May 10.
Artigo em Chinês | MEDLINE | ID: mdl-32335886

RESUMO

OBJECTIVE: To carry out genetic analysis for a family with a fetus manifesting bilateral polycystic renal dysplasia and oligohydramnios at 16+ gestational week and a previous history for fetal renal anomaly. METHODS: Ultrasound scan was carried out to detect the morphological changes. Following genetic counselling, the parents had decided to terminate the pregnancy. Fetal kidneys were subjected to histological examination. Target capture and next generation sequencing (NGS) was applied to the abortus to detect potential variants. The results were verified by Sanger sequencing. RESULTS: Histological examination of fetal kidneys revealed cystic changes without cortex, medulla or normal renal structure. NGS has identified a heterozygous c.100+1G>A variant and deletion of exon 3 of the INVS gene, which were respectively inherited from the mother and father. CONCLUSION: Through NGS and Sanger sequencing, the fetus was diagnosed with type II nephronophthisis (NPHP2). Above result can provide guidance for further pregnancy and enforce understanding of clinical features and genetic etiologies for NPHP.


Assuntos
Rim Policístico Autossômico Dominante , Fatores de Transcrição , Ultrassonografia , Feminino , Feto , Testes Genéticos , Heterozigoto , Humanos , Mutação , Rim Policístico Autossômico Dominante/diagnóstico por imagem , Rim Policístico Autossômico Dominante/genética , Gravidez , Deleção de Sequência/genética , Fatores de Transcrição/genética
14.
Int J Mol Sci ; 21(6)2020 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-32183336

RESUMO

The continuous search for novel enzyme backbones and the engineering of already well studied enzymes for biotechnological applications has become an increasing challenge, especially by the increasing potential diversity space provided by directed enzyme evolution approaches and the demands of experimental data generated by rational design of enzymes. In this work, we propose a semi-rational mutational strategy focused on introducing diversity in structurally variable regions in enzymes. The identified sequences are subjected to a progressive deletion of two amino acids and the joining residues are subjected to saturation mutagenesis using NNK degenerate codons. This strategy offers a novel library diversity approach while simultaneously decreasing enzyme size in the variable regions. In this way, we intend to identify and reduce variable regions found in enzymes, probably resulting from neutral drift evolution, and simultaneously studying the functional effect of said regions. This strategy was applied to Bacillus. subtilis lipase A (BSLA), by selecting and deleting six variable enzyme regions (named regions 1 to 6) by the deletion of two amino acids and additionally randomizing the joining amino acid residues. After screening, no active variants were found in libraries 1% and 4%, 15% active variants were found in libraries 2% and 3%, and 25% for libraries 5 and 6 (n = 3000 per library, activity detected using tributyrin agar plates). Active variants were assessed for activity in microtiter plate assay (pNP-butyrate), thermal stability, substrate preference (pNP-butyrate, -palmitate), and compared to wildtype BSLA. From these analyses, variant P5F3 (F41L-ΔW42-ΔD43-K44P), from library 3 was identified, showing increased activity towards longer chain p-nitrophenyl fatty acid esters, when compared to BSLA. This study allowed to propose the targeted region 3 (positions 40-46) as a potential modulator for substrate specificity (fatty acid chain length) in BSLA, which can be further studied to increase its substrate spectrum and selectivity. Additionally, this variant showed a decreased thermal resistance but interestingly, higher isopropanol and Triton X-100 resistance. This deletion-randomization strategy could help to expand and explore sequence diversity, even in already well studied and characterized enzyme backbones such as BSLA. In addition, this strategy can contribute to investigate and identify important non-conserved regions in classic and novel enzymes, as well as generating novel biocatalysts with increased performance in specific processes, such as enzyme immobilization.


Assuntos
Bacillus subtilis/genética , Ácidos Graxos/metabolismo , Engenharia de Proteínas/métodos , Esterol Esterase/genética , Aminoácidos/genética , Bacillus subtilis/enzimologia , Bacillus subtilis/metabolismo , Sítios de Ligação , Biblioteca Gênica , Hidrólise , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Mutagênese Sítio-Dirigida , Conformação Proteica , Deleção de Sequência/genética , Esterol Esterase/metabolismo , Especificidade por Substrato/genética
15.
Am J Med Genet A ; 182(5): 1230-1235, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32022420

RESUMO

We describe a patient with palatal abnormalities-cleft palate and bifid uvula; distinctive facial features-long and triangular face, large ears and nose, thin lips and dental crowding; musculoskeletal abnormalities-severe scoliosis, joint laxity, long digits, flat feet, decreased muscle mass, and diminished muscle strength; and cardiac features-a dilatated ascending aorta at the level of Valsalva sinuses and a patent foramen ovale. Sequence analysis and deletion/duplication testing for a panel of genes involved in connective tissue disorders revealed the presence of a novel homozygous deletion of exons 2-7 in TGFB3 gene. Heterozygous pathogenic mutations in TGFB3 have been associated with Loeys-Dietz syndrome 5 (LDS5) and Arrhythmogenic Right Ventricular Dysplasia type 1. Here, we report the first case of a homozygous TGFB3 variant associated with a severe LDS5 and Marfan-like presentation.


Assuntos
Displasia Arritmogênica Ventricular Direita/genética , Síndrome de Loeys-Dietz/genética , Síndrome de Marfan/genética , Fator de Crescimento Transformador beta3/genética , Displasia Arritmogênica Ventricular Direita/diagnóstico por imagem , Displasia Arritmogênica Ventricular Direita/fisiopatologia , Criança , Pré-Escolar , Éxons/genética , Deleção de Genes , Predisposição Genética para Doença , Homozigoto , Humanos , Lactente , Síndrome de Loeys-Dietz/diagnóstico por imagem , Síndrome de Loeys-Dietz/fisiopatologia , Masculino , Síndrome de Marfan/diagnóstico por imagem , Síndrome de Marfan/fisiopatologia , Mutação/genética , Deleção de Sequência/genética
16.
PLoS One ; 15(2): e0228925, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32053675

RESUMO

Adenocarcinoma is the most common type of non-small cell lung cancer. Some causative genomic alterations in epidermal growth factor receptor (EGFR), including deletions in exon 19 (E19 dels) and a point mutation in E21, are known to have favourable prognoses due to sensitivity to tyrosine kinase inhibitors; however, the prognoses of other uncommon mutations are unclear. This study analysed the clinical significance of EGFR mutation types in lung adenocarcinoma. We retrospectively reviewed 1,020 subjects (mean age: 66.8 years, female: 41.7%) who were diagnosed with advanced lung adenocarcinoma, had EGFR mutation data, and did not undergo surgery from five medical institutes between 2010 and 2016. Subjects were classified according to EGFR mutation status, particularly for exon-specific mutations. EGFR positivity was defined as the presence of mutation and EGFR negativity was defined as wild-type EGFR. EGFR positivity was 38.0%, with the incidence of mutations in E18, E19, E20, and E21 was 3.6%, 51.0%, 3.4%, and 42.0%, respectively. The EGFR positive group survived significantly longer than the negative group (p<0.001), and there was a significant difference in survival among the four EGFR mutation sites (p = 0.003); E19 dels were the only significant factor that lowered mortality (HR: 0.678, p = 0.002), while an E21 mutation was the prognostic factor associated with the most increased mortality (HR: 1.365, p = 0.015). Amongst EGFR positive subjects, the proportion of E19 dels in TKI-responders was significantly higher and that of E21 mutations significantly lower, compared with non-responders. In TKI treatment, mutations in E18 and E20 were not worse factors than the E21 L858R mutation. In conclusion, the presence of EGFR mutations in advanced lung adenocarcinoma can predict a good prognosis; E19 dels prospect to have a better prognosis than other mutations, while an E21 mutation is expected to increase mortality.


Assuntos
Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/mortalidade , Adenocarcinoma de Pulmão/metabolismo , Idoso , Grupo com Ancestrais do Continente Asiático , Carcinoma Pulmonar de Células não Pequenas/patologia , Receptores ErbB/genética , Receptores ErbB/metabolismo , Éxons/genética , Feminino , Humanos , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Mutação/genética , Mutação Puntual/genética , Prognóstico , República da Coreia , Estudos Retrospectivos , Deleção de Sequência/genética
17.
BMC Med Genet ; 21(1): 20, 2020 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-32005174

RESUMO

BACKGROUND: X-linked ichthyosis (XLI; OMIM# 308100) is a recessive keratinization disorder characterized by the presence of dark brown, polygonal, adherent scales on different parts of the body surface. It almost exclusively affects males and the estimated prevalence ranges from 1:2000-6000 in males worldwide. Extracutaneous manifestations are frequent including corneal opacities, cryptorchidism, neuropsychiatric symptoms or others. Up to 90% of XLI cases are caused by recurrent hemizygous microdeletion encompassing entire STS gene on chromosome Xp22.3, while only a minority of patients shows partial deletions or loss of function point mutations in STS. Larger deletions also involving contiguous genes are identified in syndromic patients. METHODS: Here, we report clinical and genetic findings of a large Pakistani family having 16 affected individuals including 2 females with XLI. Molecular karyotyping and direct DNA sequencing of coding region of the STS gene was performed. RESULTS: The clinical manifestations in affected individuals involved generalized dryness and scaling of the skin with polygonal, dark scales of the skin on scalp, trunk, limbs, and neck while sparing face, palms and soles. There were no associated extra-cutaneous features such as short stature, cryptorchidism, photophobia, corneal opacities, male baldness, and behavioral, cognitive, or neurological phenotypes including intellectual disability, autism or attention deficit hyperactivity disorder. Molecular karyotyping was normal and no copy number variation was found. Sanger sequencing identified a novel hemizygous nonsense mutation (c.287G > A; p.W96*), in exon 4 of STS gene in all affected male individuals. In addition, two XLI affected females in the family were found to be homozygous for the identified variant. CONCLUSIONS: This study is useful for understanding the genetic basis of XLI in the patients studied, for extending the known mutational spectrum of STS, diagnosis of female carriers and for further application of mutation screening in the genetic counseling of this family.


Assuntos
Triagem de Portadores Genéticos , Ictiose Ligada ao Cromossomo X/genética , Pele/metabolismo , Esteril-Sulfatase/genética , Adolescente , Adulto , Códon sem Sentido/genética , Variações do Número de Cópias de DNA/genética , Feminino , Heterozigoto , Homozigoto , Humanos , Ictiose Ligada ao Cromossomo X/fisiopatologia , Masculino , Pessoa de Meia-Idade , Paquistão/epidemiologia , Fenótipo , Deleção de Sequência/genética , Pele/patologia , Adulto Jovem
18.
Cancer Sci ; 111(4): 1180-1192, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32080953

RESUMO

The onset of malignant mesothelioma (MM) is linked to exposure to asbestos fibers. Asbestos fibers are classified as serpentine (chrysotile) or amphibole, which includes the crocidolite, amosite, anthophyllite, tremolite, and actinolite types. Although few studies have been undertaken, anthophyllite has been shown to be associated with mesothelioma, and tremolite, a contaminant in talc and chrysotile, is a risk factor for carcinogenicity. Here, after characterizing the length and width of these fibers by scanning electron microscopy, we explored the cytotoxicity induced by tremolite and anthophyllite in cells from an immortalized human mesothelial cell line (MeT5A), murine macrophages (RAW264.7), and in a rat model. Tremolite and short anthophyllite fibers were phagocytosed and localized to vacuoles, whereas the long anthophyllite fibers were caught on the pseudopod of the MeT5A and Raw 264.7 cells, according to transmission electron microscopy. The results from a 2-day time-lapse study revealed that tremolite was engulfed and damaged the MeT5A and RAW264.7 cells, but anthophyllite was not cytotoxic to these cells. Intraperitoneal injection of tremolite in rats induced diffuse serosal thickening, whereas anthophyllite formed focal fibrosis and granulomas on peritoneal serosal surfaces. Furthermore, the loss of Cdkn2a/2b, which are the most frequently lost foci in human MM, were observed in 8 cases of rat MM (homozygous deletion [5/8] and loss of heterozygosity [3/8]) by array-based comparative genomic hybridization techniques. These results indicate that tremolite initiates mesothelial injury and persistently frustrates phagocytes, causing subsequent peritoneal fibrosis and MM. The possible mechanisms of carcinogenicity based on fiber diameter/length are discussed.


Assuntos
Asbestos/toxicidade , Inibidor de Quinase Dependente de Ciclina p15/genética , Inibidor p16 de Quinase Dependente de Ciclina/genética , Neoplasias Pulmonares/genética , Mesotelioma/genética , Animais , Amiantos Anfibólicos/toxicidade , Asbesto Crocidolita/toxicidade , Asbestos Serpentinas/toxicidade , Hibridização Genômica Comparativa , Homozigoto , Humanos , Neoplasias Pulmonares/induzido quimicamente , Neoplasias Pulmonares/patologia , Mesotelioma/induzido quimicamente , Mesotelioma/patologia , Ratos , Fatores de Risco , Deleção de Sequência/genética
19.
Infect Genet Evol ; 81: 104260, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32092483

RESUMO

COVID-19 is a viral respiratory illness caused by a new coronavirus called SARS-CoV-2. The World Health Organization declared the SARS-CoV-2 outbreak a global public health emergency. We performed genetic analyses of eighty-six complete or near-complete genomes of SARS-CoV-2 and revealed many mutations and deletions on coding and non-coding regions. These observations provided evidence of the genetic diversity and rapid evolution of this novel coronavirus.


Assuntos
Betacoronavirus/classificação , Betacoronavirus/genética , Evolução Molecular , Variação Genética , Genoma Viral/genética , Mutação , Deleção de Sequência/genética
20.
Biochem Biophys Res Commun ; 524(3): 689-695, 2020 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-32033747

RESUMO

Dehydration-responsive element binding (DREB) transcription factors activate the expression of downstream functional genes in combination with a dehydration-responsive element (DRE), and thereby improve the resistance of plants to abiotic stresses such as heat. However, the upstream regulatory mechanism of DREB genes under heat is unclear. A DREBA4 subfamily transcription factor (SlDREBA4), which is heat-responsive and improves heat resistance, was isolated from Solanum lycopersicum 'Microtom'. In this study, promoter truncation experiments were performed to verify changes in ß-glucuronidase (GUS) enzyme activity and GUS gene expression levels in transgenic plants with different lengths of promoter fragments under heat and to identify specific regions in the promoter that respond to heat. Our results showed that the GUS reporter gene was constitutively expressed in tissues of the full-length promoter transgenic 'Microtom' plants, with higher expression in conducting tissues of root, stem, and leaf, as well as sepals of flowers and fruits. Under heat treatment, GUS enzyme activity and GUS gene expression levels in tissues of the full-length promoter transgenic plants increased. Promoter deletion analysis identified two positive regulatory regions (-1095 to -730 bp and -162 to -38 bp) responsible for the promoter's response to heat. These results indicated that the heat shock element (HSE) and MYC recognition sequences may cooperate in heat-induced activation of SlDREBA4 promoter.


Assuntos
Regulação da Expressão Gênica de Plantas , Temperatura Alta , Lycopersicon esculentum/genética , Proteínas de Plantas/genética , Regiões Promotoras Genéticas , Sequência de Bases , Glucuronidase/metabolismo , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Deleção de Sequência/genética
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