RESUMO
Blunt snout bream (Megalobrama amblycephala) is sensitive to hypoxia environment. Hypoxia-inducible factor (HIF) is the most critical factor in the HIF pathway, which strictly regulates the hypoxia stress process of fish. In this study, we found six hifα genes in blunt snout bream that demonstrated different expressions under hypoxia conditions. In HEK293T cells, all six hifαs were detected to activate the HRE region by luciferase reporter assay. More importantly, we identified two linkage-disequilibrium SNP sites at exon 203 and 752 of the hif2αb gene in blunt snout bream. Haplotype II (A203A752) and its homozygous diplotype II (A203A203A752A752) appeared frequently in a selected strain of blunt snout bream with hypoxia tolerance. Diplotype II has a lower oxygen tension threshold for loss of equilibrium (LOEcrit) over a similar range of temperatures. Moreover, its erythrocyte number increased significantly (p < 0.05) than those in diplotype I and diplotype III strains at 48 h of hypoxia. The enzymes related with hypoxia tolerant traits, i.e., reduced glutathione, superoxide dismutase, and catalase, were also significantly (p < 0.05) induced in diplotype II than in diplotype I or III. In addition, the expression of epo in the liver of diplotype II was significantly (p < 0.01) higher than that in the diplotype I or III strains at 48 h of hypoxia. Taken together, our results found that the hypoxia-tolerant-related diplotype II of hif2αb has the potential to be used as a molecular marker in future genetic breeding of hypoxia-tolerant strain.
Assuntos
Cyprinidae , Cipriniformes , Animais , Humanos , Cyprinidae/metabolismo , Células HEK293 , Cipriniformes/metabolismo , Mutação , Hipóxia/genética , Hipóxia/metabolismo , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismoRESUMO
N-ethyl-N-nitrosourea (ENU) selection is a useful technique to generate new mutations that may cause some functional changes in the gene. Through our previous genomic bulked segregant analysis (BSA), one single nucleotide polymorphism (SNP) at the 3' UTR of Toll interacting protein gene (TOLLIP982T>C) was identified in grass carp (Ctenopharyngodon idella) subjected to ENU-induced mutagenesis. We found that the overexpression of cid-miR-nov-1043 mimics significantly suppressed the luciferase activity of the TOLLIP 3' UTR, but TOLLIP982T>C mutation at the target site can decrease the binding affinity between the miRNA cid-miR-nov-1043 and TOLLIP 3' UTR, reducing the inhibition of TOLLIP mRNA transcription in grass carp subjected to ENU-induced mutagenesis. More importantly, we demonstrated that TOLLIP mRNA transcription levels in the gills, liver, kidney and the isolate white cells of the mutant grass carp were significantly (p < 0.01) higher than those in the corresponding tissues from the wild-type grass carp following infection with Grass Carp Reovirus (GCRV) for seven days, while the downstream gene of TOLLIP transforming growth factor ß-activated kinase 1 (TAK1) and TAK1-binding protein 1 (TAB1), were higher expressed in wild-type grass carp. As a negative regulator in the pro-inflammatory pathway of NF-κB, TOLLIP inhibits the excessive inflammation in ENU grass carp after GCRV infection. Consistent with the TOLLIP expression, histopathological results demonstrated more severe inflammation in wild-type grass carp, compared to the TOLLIP982T>C mutant grass carp on the seventh day. Severe inflammation will lead to thoroughly infiltration of chloride and inflammatory cells in the gill filaments. This seriously hindered the exchange of oxygen, which ultimately disrupted blood circulation. Meanwhile, the survival rate of the mutant grass carp was significantly (p < 0.01) higher than that of the wild-type grass carp, indicating that the TOLLIP982T>C mutants showed strong anti-viral abilities. Our results revealed that an SNP in the TOLLIP 3' UTR may contribute to the suppression of serve inflammation subjected to ENU-induced mutagenesis following GCRV infection, which may be helpful for future resistant breeding development of grass carp.
Assuntos
Carpas , Doenças dos Peixes , Peptídeos e Proteínas de Sinalização Intracelular/genética , MicroRNAs , Polimorfismo de Nucleotídeo Único , Infecções por Reoviridae , Regiões 3' não Traduzidas , Animais , Carpas/genética , Carpas/virologia , Etilnitrosoureia , Doenças dos Peixes/genética , Doenças dos Peixes/virologia , Proteínas de Peixes/genética , Inflamação , MicroRNAs/genética , Mutagênese , Reoviridae , Infecções por Reoviridae/genética , Infecções por Reoviridae/veterináriaRESUMO
Although CYP2C19 is minor human liver enzyme, it is responsible for the metabolism of many clinically important drugs. In this work, CYP2C19 wild type and its SNP mutants (R132Q and W120R) were prepared using over-expression system in E. coli, purified by column chromatography and their biological activities were compared. The enzyme activity toward certain drugs (amitriptyline, imipramine, lansoprazole and omeprazole) was investigated. Resonance Raman and UV-VIS spectroscopies revealed a minimal effect of SNP mutations on the heme structure. However, the mutation greatly affected the drug metabolism activities of CYP2C19. The degree of these effects was dependent on both the mutation and the chemical structure of the substrate. Surprisingly, the affected amino acid residue is located remotely from the heme center. Therefore, the direct effect of the mutation on the metabolic center is excluded. Alternatively, the significant impairment in the drug metabolism of these mutants could be attributed to a decrease in the electron flow to the iron center. Accordingly, understanding the effect of SNPs of CYP2C19, and the extents in which they participate in the drug metabolism, are important pillars that can enhance the therapeutic drugs efficacy and improve the patient outcomes toward the development of patient's tailored medicine.
Assuntos
Citocromo P-450 CYP2C19/metabolismo , Escherichia coli , Humanos , Omeprazol/metabolismo , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Myocardial fibrosis (MF) is characterized by the excessive deposition of extracellular matrix within the heart, often following a cardiovascular insult. SHARPIN, a protein implicated in fibrosis, has emerged as a potential therapeutic target. This study aimed to elucidate the molecular mechanisms of SHARPIN in MF and to investigate the influence of its single nucleotide polymorphism (SNP), rs117299156, on myocardial infarction (MI) patients. A mouse model of Angiotensin II (AngII)-induced MF was established in SHARPIN heterozygous (SHARPIN+/-) and wild-type mice. Adult mouse cardiac fibroblasts (CFs) were isolated and subjected to adenovirus-encapsulated SHARPIN short hairpin RNA (shRNA) infection. Transcriptomic analysis was performed on CFs from SHARPIN+/- and wild-type (WT) mice, complemented by single-cell sequencing data from human cardiac tissues. Additionally, the association between the rs117299156 mutation and cardiovascular events in MI patients was assessed. Our findings indicate that SHARPIN is predominantly expressed in CFs and is upregulated in fibrotic myocardium. Partial knockdown of SHARPIN in murine hearts mitigated AngII-induced cardiac dysfunction and MF. Furthermore, reduced SHARPIN expression in CFs attenuated TGF-ß1-induced collagen synthesis, cell proliferation, and myofibroblast transformation. Notably, MI patients carrying the rs117299156_C allele exhibited a reduced incidence of stroke events compared to those without the mutation.
RESUMO
BACKGROUND: Phalaenopsis is an important ornamental plant that has great economic value in the world flower market as one of the most popular flower resources. OBJECTIVE: To investigate the flower colour formation of Phalaenopsis at the transcription level, the genes involved in flower color formation were identified from RNA-seq in this study. METHODS: In this study, white and purple petals of Phalaenopsis were collected and analyzed to obtained (1) differential expression genes (DEGs) between white and purple flower color and (2) the association between single nucleotide polymorphisms (SNP) mutations and DEGs at the transcriptome level. RESULTS: The results indicated that a total of 1,175 DEGs were identified, and 718 and 457 of them were up- and down-regulated genes, respectively. Gene Ontology and pathway enrichment showed that the biosynthesis of the secondary metabolites pathway was key to color formation, and the expression of 12 crucial genes (C4H, CCoAOMT, F3'H, UA3'5'GT, PAL, 4CL, CCR, CAD, CALDH, bglx, SGTase, and E1.11.17) that are involved in the regulation of flower color in Phalaenopsis. CONCLUSION: This study reported the association between the SNP mutations and DEGs for color formation at RNA level, and provides a new insight to further investigate the gene expression and its relationship with genetic variants from RNA-seq data in other species.
Assuntos
Orchidaceae , Orchidaceae/genética , Cor , Polimorfismo de Nucleotídeo Único , Perfilação da Expressão Gênica , Flores/genética , Flores/metabolismoRESUMO
BACKGROUND: Emergence of new variant of SARS-CoV-2, namely omicron, has posed a global concern because of its high rate of transmissibility and mutations in its genome. Researchers worldwide are trying to understand the evolution and emergence of such variants to understand the mutational cascade events. METHODS: We have considered all omicron genomes (n = 302 genomes) available till 2nd December 2021 in the public repository of GISAID along with representatives of variants of concern (VOC), i.e., alpha, beta, gamma, delta, and omicron; variant of interest (VOI) mu and lambda; and variant under monitoring (VUM). Whole genome-based phylogeny and mutational analysis were performed to understand the evolution of SARS CoV-2 leading to emergence of omicron variant. RESULTS: Whole genome-based phylogeny depicted two phylogroups (PG-I and PG-II) forming variant specific clades except for gamma and VUM GH. Mutational analysis detected 18,261 mutations in the omicron variant, majority of which were non-synonymous mutations in spike (A67, T547K, D614G, H655Y, N679K, P681H, D796Y, N856K, Q954H), followed by RNA dependent RNA polymerase (rdrp) (A1892T, I189V, P314L, K38R, T492I, V57V), ORF6 (M19M) and nucleocapsid protein (RG203KR). CONCLUSION: Delta and omicron have evolutionary diverged into distinct phylogroups and do not share a common ancestry. While, omicron shares common ancestry with VOI lambda and its evolution is mainly derived by the non-synonymous mutations.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , Mutação , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/metabolismoRESUMO
Antimalarial drug resistance has emerged as a major threat to global malaria control efforts, particularly in the Greater Mekong Subregion (GMS). In this study, we analyzed the polymorphism and prevalence of molecular markers associated with resistance to first-line antimalarial drugs, such as artemisinin, chloroquine, and pyrimethamine, using blood samples collected from malaria patients in the China-Myanmar border region of the GMS from 2008 to 2017, including 225 cases of Plasmodium falciparum and 194 cases of Plasmodium vivax. In artemisinin resistance, only the C580Y mutation with low frequency was detected in pfk13, and no highly frequent stable mutation was found in pvk12. In chloroquine resistance, the frequency of K76T mutation in pfcrt was always high, and the frequency of double mutations in pvmdr1 of P. vivax has been steadily increasing every year. In pyrimidine resistance, pfdhfr and pvdhfr had relatively more complex mutant types associated with drug resistance sites, and the overall mutation rate was still high. Therefore, artemisinin-based combination therapies are still suitable for use as the first choice of antimalarial strategy in the China-Myanmar border region in the future.