Effects Of Gene Mutations Research Articles

Ribosomal computing: implementation of the computational method

BackgroundSeveral computational and mathematical models of protein synthesis have been explored to accomplish the quantitative analysis of protein synthesis components and polysome structure. The effect of gene sequence (coding and non-coding region) in protein synthesis, mutation in gene sequence, and functional model of ribosome needs to be explored to investigate the relationship among protein synthesis components further. Ribosomal computing is implemented by imitating the functional property of protein synthesis.ResultIn the proposed work, a general framework of ribosomal computing is demonstrated by developing a computational model to present the relationship between biological details of protein synthesis and computing principles. Here, mathematical abstractions are chosen carefully without probing into intricate chemical details of the micro-operations of protein synthesis for ease of understanding. This model demonstrates the cause and effect of ribosome stalling during protein synthesis and the relationship between functional protein and gene sequence. Moreover, it also reveals the computing nature of ribosome molecules and other protein synthesis components. The effect of gene mutation on protein synthesis is also explored in this model.ConclusionThe computational model for ribosomal computing is implemented in this work. The proposed model demonstrates the relationship among gene sequences and protein synthesis components. This model also helps to implement a simulation environment (a simulator) for generating protein chains from gene sequences and can spot the problem during protein synthesis. Thus, this simulator can identify a disease that can happen due to a protein synthesis problem and suggest precautions for it.

ABO*A2.08 Subtype Allele Identification and Protein Structure Analysis in Newborns

To study the serological characteristics of ABO*A2.08 subtype and explore its genetic molecular mechanism. ABO blood group identification was performed on proband and her family members by routine serological methods. ABO genotyping and sequence analysis were performed by polymerase chain reaction-sequence specific primer (PCR-SSP), and direct sequencing of PCR products from exons 6 and 7 of ABO gene were directly sequenced and analyzed. The effect of gene mutation in A2.08 subtype on structural stability of GTA protein was investigated by homologous protein conserved analysis, 3D molecular modeling and protein stability prediction. The proband's serological test results showed subtype Ax, and ABO genotyping confirmed that the proband's genotype was ABO*A207/08. Gene sequencing of the proband's father confirmed the characteristic variation of c.539G>C in the 7th exon of ABO gene, leading to the replacement of polypeptide chain p.Arg180Pro (R180P). 3D protein molecular modeling and analysis suggested that the number of hydrogen bonds of local amino acids in the protein structure was changed after the mutation, and protein stability prediction showed that the mutation had a great influence on the protein structure stability. The mutation of the 7th exon c.539G>C of ABO gene leads to the substitution of polypeptide chain amino acid, which affects the structural stability of GTA protein and leads to the change of enzyme activity, resulting in the A2.08 phenotype. The mutated gene can be stably inherited.

Effect of altered production and storage of dopamine on development and behavior in C. elegans.

The nematode, Caenorhabditis elegans (C. elegans), is an advantageous model for studying developmental toxicology due to its well-defined developmental stages and homology to humans. It has been established that across species, dopaminergic neurons are highly vulnerable to neurotoxicant exposure, resulting in developmental neuronal dysfunction and age-induced degeneration. C. elegans, with genetic perturbations in dopamine system proteins, can provide insight into the mechanisms of dopaminergic neurotoxicants. In this study, we present a comprehensive analysis on the effect of gene mutations in dopamine-related proteins on body size, development, and behavior in C. elegans. We studied C. elegans that lack the ability to sequester dopamine (OK411) and that overproduce dopamine (UA57) and a novel strain (MBIA) generated by the genetic crossing of OK411 and UA57, which both lack the ability to sequester dopamine into vesicles and, additionally, endogenously overproduce dopamine. The MBIA strain was generated to address the hypothesis that an endogenous increase in the production of dopamine can rescue deficits caused by a lack of vesicular dopamine sequestration. These strains were analyzed for body size, developmental stage, reproduction, egg laying, motor behaviors, and neuronal health utilizing multiple methods. Our results further implicate proper dopamine synthesis and sequestration in the regulation of C. elegans body size, development through larval stages into gravid adulthood, and motor functioning. Furthermore, our analyses demonstrate that body size in terms of length is distinct from the developmental stage as fully developed gravid adult C. elegans with disruptions in the dopamine system have decreased body lengths. Thus, body size should not be used as a proxy for the developmental stage when designing experiments. Our results provide additional evidence that the dopamine system impacts the development, growth, and reproduction in C. elegans. Furthermore, our data suggest that endogenously increasing the production of dopamine mitigates deficits in C. elegans lacking the ability to package dopamine into synaptic vesicles. The novel strain, MBIA, and novel analyses of development and reproduction presented here can be utilized in developmental neurotoxicity experiments.

Insulin regulates human pancreatic endocrine cell differentiation in vitro

ObjectiveThe consequences of mutations in genes associated with monogenic forms of diabetes on human pancreas development cannot be studied in a time-resolved fashion in vivo. More specifically, if recessive mutations in the insulin gene influence human pancreatic endocrine lineage formation is still an unresolved question. MethodsTo model the extremely reduced insulin levels in patients with recessive insulin gene mutations, we generated a novel knock-in H2B-Cherry reporter human induced pluripotent stem cell (iPSC) line expressing no insulin upon differentiation to stem cell-derived (SC-) β cells in vitro. Differentiation of iPSCs into the pancreatic and endocrine lineage, combined with immunostaining, Western blotting and proteomics analysis phenotypically characterized the insulin gene deficiency in SC-islets. Furthermore, we leveraged FACS analysis and confocal microscopy to explore the impact of insulin shortage on human endocrine cell induction, composition, differentiation and proliferation. ResultsInterestingly, insulin-deficient SC-islets exhibited low insulin receptor (IR) signaling when stimulated with glucose but displayed increased IR sensitivity upon treatment with exogenous insulin. Furthermore, insulin shortage did not alter neurogenin-3 (NGN3)-mediated endocrine lineage induction. Nevertheless, lack of insulin skewed the SC-islet cell composition with an increased number in SC-β cell formation at the expense of SC-α cells. Finally, insulin deficiency reduced the rate of SC-β cell proliferation but had no impact on the expansion of SC-α cells. ConclusionsUsing iPSC disease modelling, we provide first evidence of insulin function in human pancreatic endocrine lineage formation. These findings help to better understand the phenotypic impact of recessive insulin gene mutations during pancreas development and shed light on insulin gene function beside its physiological role in blood glucose regulation.

Mechanistic Insights into S-Depalmitolyse Activity of Cln5 Protein Linked to Neurodegeneration and Batten Disease: A QM/MM Study.

Ceroid lipofuscinosis neuronal protein 5 (Cln5) is encoded by the CLN5 gene. The genetic variants of this gene are associated with the CLN5 form of Batten disease. Recently, the first crystal structure of Cln5 was reported. Cln5 shows cysteine palmitoyl thioesterase S-depalmitoylation activity, which was explored via fluorescent emission spectroscopy utilizing the fluorescent probe DDP-5. In this work, the mechanism of the reaction between Cln5 and DDP-5 was studied computationally by applying a QM/MM methodology at the ωB97X-D/6-31G(d,p):AMBER level. The results of our study clearly demonstrate the critical role of the catalytic triad Cys280-His166-Glu183 in S-depalmitoylation activity. This is evidenced through a comparison of the pathways catalyzed by the Cys280-His166-Glu183 triad and those with only Cys280 involved. The computed reaction barriers are in agreement with the catalytic efficiency. The calculated Gibb's free-energy profile suggests that S-depalmitoylation is a rate-limiting step compared to the preceding S-palmitoylation, with barriers of 26.1 and 25.3 kcal/mol, respectively. The energetics were complemented by monitoring the fluctuations in the electron density distribution through NBO charges and bond strength alterations via local mode stretching force constants during the catalytic pathways. This comprehensive protocol led to a more holistic picture of the reaction mechanism at the atomic level. It forms the foundation for future studies on the effects of gene mutations on both the S-palmitoylation and S-depalmitoylation steps, providing valuable data for the further development of enzyme replacement therapy, which is currently the only FDA-approved therapy for childhood neurodegenerative diseases, including Batten disease.

Association between fatty acid metabolism gene mutations and Mycobacterium tuberculosis transmission revealed by whole genome sequencing

BackgroundFatty acid metabolism greatly promotes the virulence and pathogenicity of Mycobacterium tuberculosis (M.tb). However, the regulatory mechanism of fatty acid metabolism in M.tb remains to be elucidated, and limited evidence about the effects of gene mutations in fatty acid metabolism on the transmission of M.tb was reported.ResultsOverall, a total of 3193 M.tb isolates were included in the study, of which 1596 (50%) were genomic clustered isolates. Most of the tuberculosis isolates belonged to lineage2(n = 2744,85.93%), followed by lineage4(n = 439,13.75%) and lineage3(n = 10,0.31%).Regression results showed that the mutations of gca (136,605, 317G > C, Arg106Pro; OR, 22.144; 95% CI, 2.591-189.272), ogt(1,477,346, 286G > C ,Gly96Arg; OR, 3.893; 95%CI, 1.432–10.583), and rpsA (1,834,776, 1235 C > T, Ala412Val; OR, 3.674; 95% CI, 1.217–11.091) were significantly associated with clustering; mutations in gca and rpsA were also significantly associated with clustering of lineage2. Mutation in arsA(3,001,498, 885 C > G, Thr295Thr; OR, 6.278; 95% CI, 2.508–15.711) was significantly associated with cross-regional clusters. We also found that 20 mutation sites were positively correlated with cluster size, while 11 fatty acid mutation sites were negatively correlated with cluster size.ConclusionOur research results suggested that mutations in genes related to fatty acid metabolism were related to the transmission of M.tb. This research could help in the future control of the transmission of M.tb.

Clinical Outcomes Following an Initial Stereotactic Radiosurgery Course for Brain Metastases from Melanoma

Brain metastases (BM) are common in melanoma patients. The effect of gene mutations is not well characterized since first-line metastatic therapy has shifted from chemotherapy (CHT) to molecularly targeted therapies (TT) and immunotherapy (IO). We report outcomes of melanoma BM patients stratified by molecular subtype and pre-stereotactic radiosurgery (SRS) systemic therapy. We identified all patients completing an initial SRS course for BM at two institutions between 1/2015 and 12/2020. Patients who had prior WBRT and/or resection were eligible. Demographic and clinical parameters were collected, along with melanoma tumor molecular characteristics. Intracranial progression (ICP) was defined as any radiographic distant and/or in-field progression per multidisciplinary consensus. Overall survival (OS) and freedom from ICP (FFICP) were estimated via the Kaplan Meier method. From a total of 1383 SRS BM patients, we identified 118 (8.5%) with melanoma. Median follow up was 8.7 months, median age 64 years (IQR 51-72), 81% had cutaneous origin, and 55% had a KPS of 90-100. Molecular subtypes included BRAF (45%), NRAS (9.3%), and c-KIT (3.4%). Overall, 61% received IO prior to SRS, while 25% and 9.3% received TT and CHT prior to SRS respectively. 60% of patients harboring a mutation received IO as first line therapy, 10% received TT, and 30% received both TT and IO prior to SRS. BRAFmut patients more likely to have received TT prior to SRS (43% vs 9.2%, p<0.001) compared to BRAFwt patients. Median OS was 9.7 months (95% CI 7.8-13) and was not significantly different from non-melanoma patients (p = 0.6). Median FFICP was worse for melanoma patients (5.9 mos, 95% CI 3.5-8.5) than non-melanoma patients (8.96 mos, 95% CI 8.2-9.7, p = 0.009). A total of 72 ICP events occurred, with 56 (77.8%) distant ICP cases, 3 (4.2%) in-field ICP, and 13 (18%) ICP events that were radionecrosis (RN) only. RN was associated with the presence of a targetable mutation (18% vs 2%, p = 0.006) and receipt of TT pre-SRS (36% vs 9.8%, p = 0.001). BRAFmut patients had significantly worse FFICP (3.8 mos, 95% CI 3.0-6.8) compared to BRAFwt patients (8.5 mos, 95% CI 5.8-30.2, p = 0.006), although median OS was not significantly different (9.6 mos, 95% CI 6.9-16 vs 10.7 mos, 95% CI 6.7-15.5, p = 0.8). NRASmut was associated with better FFICP (29 mos, 95% CI 2.94-NA, p = 0.02). In this modern, multi-institutional cohort of SRS patients, melanoma BM patients had worse FFICP compared to non-melanoma BM patients, and BRAFmut patients had worse FFICP than BRAFwt patients. RN was associated with mutational status and receipt of TT pre-SRS. OS did not vary significantly across groups. This analysis may help inform systemic therapy decisions and future genomic studies for patients with BMs from melanoma.

Impact of somatic mutations and transcriptomic alterations on cancer aneuploidy.

Aneuploidy has been recognized as one of hallmark of tumorigenesis since the early 20th century. Recent developments in structural variation analysis in the human genome have revealed the diversity of aneuploidy in cancer. However, the effects of gene mutation and expression in tumors on aneuploidy remain poorly understood. Here, we performed whole exome analysis of over 5,000 Japanese cancer cases and investigated the impact of somatic mutations and gene expression alterations on aneuploidy. First, we evaluated tumor content and genomic alterations that could influence aneuploidy. Next, we compared the aneuploidy frequency in 18 cancer types and observed that TP53 mutations were associated with the aneuploidy on specific chromosomes in colorectal and gastric cancers. Finally, we used expression analysis to isolate pathways involved in aneuploidy accumulation from tumors without TP53 mutations. Chromosomal instability and cell cycle aberration were associated with aneuploidy in TP53 wild-type tumors, and 26 commonly upregulated genes were identified in aneuploidy-high solid tumors without TP53 mutations. Among them, two cancer-related genes (CENPA and PBK) were involved in aneuploidy. Our integrated analysis revealed that both TP53 mutations and transcriptomic alterations independent of somatic mutations affect aneuploidy accumulation. Our findings will facilitate further understanding of diverse aneuploidies in the tumorigenesis.

Identification of potential susceptibility genes in patients with primary Sjögren’s syndrome-associated pulmonary arterial hypertension through whole exome sequencing

BackgroundPulmonary arterial hypertension (PAH) is a rare complication of primary Sjögren’s syndrome (pSS). Several genes have proven to be associated with pSS and PAH. However, there is no study specifically addressing the genetic susceptibility in pSS combined with PAH.MethodsThirty-four unrelated patients with pSS-PAH were recruited from April 2019 to July 2021 at Peking Union Medical College Hospital. Demographic and clinical data were recorded in detail, and peripheral blood samples were collected for whole-exome sequencing (WES). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed to predict the functional effect of mutant genes. Genetic variants identified by WES were confirmed by polymerase chain reaction (PCR)-Sanger sequencing.ResultsWe totally identified 141 pathogenic variant loci of 129 genes in these 34 pSS-PAH patients, using WES analysis. Patients with a family history of rheumatic diseases are more likely to carry FLG mutations or carry gene variations related to the biosynthesis of the amino acids pathway (p < 0.05). According to Sanger sequencing confirmation and pathogenicity validation, we totally identified five candidate pathogenic variants including FLG c.12064A > T, BCR c.3275_3278dupCCGG, GIGYF2 c.3463C > A, ITK c.1741C > T, and SLC26A4 c.919-2A > G.ConclusionOur findings provide preliminary data of exome sequencing to identify susceptibility loci for pSS-PAH and enriched our understanding of the genetic etiology for pSS-PAH. The candidate pathogenic genes may be the potential genetic markers for early warning of this disease.

Albinism in the common shrew Sorex araneus

AbstractAlbinism is a hypopigmentation disorder which occurs as a consequence of mutations in genes involved in melanin biosynthesis. It is recorded in many mammalian species but is generally considered to be rare in natural populations. Here, we describe a fully albino individual of the common shrew Sorex araneus found in south-western Poland. We also present previous findings of albino specimens of this species in Europe. Since the 1960s, there have only been three confirmed records of wholly albino individuals of common shrews in mainland Europe; however, albinism has been recorded more frequently in this species from Britain. In other shrew species, wholly albino individuals are also recorded very rarely.

Study on BW.12 Subtype Caused by c.278C>T Mutation in Exon 6 of ABO Gene

To investigate the effect of ABO gene α-1,3-D galactosyl transferase mutation on B antigen expression and its molecular mechanism. The proband and their family members were identified by routine serological methods, and ABO genotyping and sequence analysis were performed by polymerase chain reaction-sequence specificity (PCR-SSP) and direct sequencing of PCR products from exon 1-7 of ABO gene. The 3D structural simulation of mutant proteins was performed by bioinformatics software. The effect of gene mutation on protein structural stability was analyzed. The proband and his family members were subtype B. ABO genotyping indicated that the proband's genotype was Bw12/O. Gene sequencing results confirmed the presence of ABO*BW.12 characteristic variation c.278C>T in the 6th exon of allele B, leading to the replacement of polypeptide chain p.Pro93Leu. The 3D structure simulation analysis of the protein showed that the hydrogen bonds and water molecules connected to the protein changed after amino acid substitution. The family investigation found that the grandfather, father, uncle and brother of the proband all carried the same ABO*BW.12 allele. The mutation of the 6th exon c.278C>T of ABO gene led to the substitution of polypeptide chain amino acids, which affected the stability of α-1,3-D galactosyl transferase protein, resulting in the change of enzyme activity, and the Bw.12 phenotype, which can be stably inherited.

Milk Exosomes Select Mutations That Decrease the Toxicity of Clostridioides difficile

ObjectivesBackground: Exosomes are natural nanoparticles that play an essential role in cell-to-cell communication. Exosomes are present in all body fluids and their concentration is particularly high in milk. We previously found that bovine milk exosomes (BMEs) select polymorphisms and mutations in gut bacteria, potentially affecting the synthesis of microbial metabolites. Clostridioides difficile (C. difficile) is the primary cause of infectious diarrhea in the United States and it is unknown whether BMEs impact its pathogenicity through the selection of mutations. Objective: Assess the effect of chronic BME exposure on C. difficile growth and pathogenicity. MethodsC. difficile was cultured in BME-supplemented and -free media under continuous culture conditions for 21 days. Strains were isolated from both conditions. Doubling times and maximal density were assessed by monitoring optical density (OD600). Toxicity was assessed by Vero cell rounding and qRT-PCR of toxin genes (tcdA and tcdB) mRNA. Whole-genome sequencing was used to identify mutations in C. difficile strains. Effects of gene mutations on growth and toxicity were determined by overexpression of the mutants in C. difficile. Mann-Whitney test was used for statistical analysis; P < 0.05 was considered significant. ResultsBMEs selected for mutations that reduced doubling time by 15% (P < 0.05), decreased tcdA and tcdB production by 92% and 93%, respectively, (P < 0.05) and relative toxin activity reduced by 74% (p = 0.055). BME-selected strains contained a 25-bp deletion in the gene encoding the glucose-specific PTS transporter subunit IIBCA (ptsG). As a result, the translated ptsG (ptsG-indel) protein is truncated by 49 amino acids with 5 mismatches compared with wild-type. Overexpression of ptsG-indel in a wild-type strain of C. difficile increased maximum OD600 by 34% (P < 0.05), lowered relative toxin activity by 88% (P < 0.001), and reduced tcdB expression by 94% (P < 0.05) compared to C. difficile that overexpress wild-type ptsG. ConclusionsBMEs decreased the pathogenicity of C. difficile. Decreased pathogenicity was likely due to mutations in ptsG selected by growth in the presence of BMEs. Funding SourcesNIH P20GM104320, NIFA 2020–67,017-30,834, USDA Hatch and W-4002. J.Z is a consultant for PureTech Health, Inc.

Predictive and prognostic value of gene mutations in myelodysplastic syndrome treated with hypomethylating agents: a meta-analysis

Although the effect of gene mutations on overall response rate (ORR) and overall survival (OS) in myelodysplastic syndrome (MDS) treated with hypomethylating agents (HMAs) has been explored, the effect is still controversial. We performed this meta-analysis to investigate the effect. The pooled odds ratio (OR) and 95% confidence interval (CI) for ORR and the pooled hazard ratio (HR) and 95%CI for OS were chosen to estimate the effect. The pooled OR of TET2 was 0.73 (95%CI: 0.59–0.91, p = 0.005) and the pooled OR of ASXL1 was 1.38 (95%CI: 1.12–1.71, p = 0.003). As for prognosis, the pooled HR of RUNX1 was 1.45 (95%CI: 1.15–1.85, p = 0.002). The pooled HR of TP53 was 2.30 (95%CI: 1.83–2.90, p < 0.001) and the pooled HR of U2AF1 was 1.41 (95%CI: 1.15–1.74, p = 0.001). There was no statistical difference shown in other genes. Therefore, TET2 mutation and ASXL1 wild-type were the predictor of better response to HMAs. Mutations of TP53, RUNX1, and U2AF1 were associated with poor prognosis in MDS.

Validation of a Novel Microfluidic Platform by Imaging Diet Uptake in Zebrafish (Danio rerio) Larvae.

Jesse Orell*, Dakota Parnell*, Chandler Russell*, and James Walters, Ph. D*. *Dept of Applied Science and Mathematics, Bluefield State College, Bluefield, WV, 24701. Validation of a Novel Microfluidic Platform by Imaging Diet Uptake in Zebrafish (Danio rerio) Larvae. &#x0D; A functional microfluidic imaging platform allows the study of several conditions such as the effect of diets on metabolism/digestion, drug interactions, and effects of gene mutations on metabolic processes. Extended durations of in vivo larval zebrafish microscopy are lacking, especially in regard to metabolism. We aim to extend the duration of in vivo larval zebrafish microscopy by using a novel microfluidic imaging platform capable of exchanging fluid (entry and exit flow), supplying oxygen, and supplying diets/drugs. Zebrafish (Danio rerio) serve as good research models for microscopy and disease related research. Zebrafish are optically transparent in larval stages and share 70% of genes with humans and 84% of which shared genes are known to be disease related. We first describe the optimal feed time to compare free feeding larval dietary consumption to larvae fed in the microfluidic platform, we imaged seven days post-fertilization (DPF) larvae fed at 1-, 3-, 5-, 7-, and 9-hour durations (10 larvae/timepoint, 3 replicates) with a Zeiss Discovery V8 microscope. We used a diet of 5% egg yolk spiked with Fluosphere polystyrene beads (580/605)™ to track the amount of diet consumed. We found the 5-hour timepoint was the best one to measure because fluorescent mean gray value quantified by NIH ImageJ had minor variation, and all larvae had fed. Future experiments will determine if seven DPF larvae will consume food equally in the microfluidic imaging platform environment. If seven DPF larvae feed equally in the microfluidic imaging platform it suggests an improved environment for imaging zebrafish larvae.

Comparison between H63D and G71D gene mutation effects on iron overload in Iraqi patients with β-thalassemia major: A case–control study

Comparison between H63D and G71D gene mutation effects on iron overload in Iraqi patients with β-thalassemia major: A case–control study

The Effect of Gene Mutations on Metastasis and Overall Survival in Metastatic and Nonmetastatic Colon Cancers.

Objective:It is known that many genes are associated with colon cancer. We aimed to investigate the effect of gene mutations on metastasis and overall survival in metastatic and non metastatic colon cancers. Methods:A total of 50 patients with metastatic (n=25) and non metastatic (n=25) diagnosed with colon cancer between 2010 and 2018 were included in the study. APC, MUTYH, RAD50, MEN1, ATM, PALB2, NSH2, BRCA1, BRCA2, MLH1, BRIP1, TP53, PTEN, BARD1, MSH6, PMS2, NBN, and FAM175A gene mutations were evaluated using the next generation sequencing method. The effect of gene mutations on metastasis and overall survival were evaluated. Results:The mean age of patients with colon cancer without distant metastasis was 48.64±14.72 years and for patients with distance metases was 56.68±11.65. The mean survival time of colon cancer patients with distant organ metastasis after the metastasis date was 104.36±58.59 weeks. The presence of APC, MUTYH, and TP53 genetic mutations was observed with a higher rate in metastatic colon cancer (p<0.05). Conclusion:We showed that APC, MUTYH, and TP53 mutations are associated with distant organ metastasis.

Identification of driver genes based on gene mutational effects and network centrality

BackgroundAs one of the deadliest diseases in the world, cancer is driven by a few somatic mutations that disrupt the normal growth of cells, and leads to abnormal proliferation and tumor development. The vast majority of somatic mutations did not affect the occurrence and development of cancer; thus, identifying the mutations responsible for tumor occurrence and development is one of the main targets of current cancer treatments.ResultsTo effectively identify driver genes, we adopted a semi-local centrality measure and gene mutation effect function to assess the effect of gene mutations on changes in gene expression patterns. Firstly, we calculated the mutation score for each gene. Secondly, we identified differentially expressed genes (DEGs) in the cohort by comparing the expression profiles of tumor samples and normal samples, and then constructed a local network for each mutation gene using DEGs and mutant genes according to the protein–protein interaction network. Finally, we calculated the score of each mutant gene according to the objective function. The top-ranking mutant genes were selected as driver genes. We name the proposed method as mutations effect and network centrality.ConclusionsFour types of cancer data in The Cancer Genome Atlas were tested. The experimental data proved that our method was superior to the existing network-centric method, as it was able to quickly and easily identify driver genes and rare driver factors.

A novel missense mutation of CRYBA1 in a northern Chinese family with inherited coronary cataract with blue punctate opacities.

To demonstrate the underlying genetic defect that contribute to inherited cataract in a northern Chinese pedigree. The study recruited a family pedigree with a diagnosis of bilateral coronary cataract with blue punctate opacities. Fourteen family members and 100 healthy volunteers were enrolled. DNA sample of the proband in this family were analyzed by high-throughput sequencing, which was then demonstrated by Sanger sequencing in the remained people in the family and 100 controls. The functional effect of mutant genes was investigated via bioinformatics analysis, including Polymorphism Phenotyping version2 (PolyPhen-2), Protein Variation Effect Analyzer (PROVEAN v1.1.3) Scale-Invariant Feature Transform (SIFT), and Mutation Taster. In this three-generation family, a novel heterozygous mutation was found in the kinase domain of CRYBA1 gene (c.340C > T, p.R114C), which was only detected in patients in the family with inherited cataract and were not detected in the remained people in the family nor in normal people. The pathogenic effect of the mutation was verified via bioinformatics analysis. Our study presented the molecular experiments to confirm that a novel missense mutation of c.340 C > T located in exon 4 of CRYBA1 gene results in a bilateral coronary cataract with blue punctate opacities, which enriches the mutation spectrum of CRYBA1 gene in inherited cataract and deepens the understanding of the pathogenesis of inherited cataract.

Modelling human blastocysts by reprogramming fibroblasts into iBlastoids.

Human pluripotent and trophoblast stem cells have been essential alternatives to blastocysts for understanding early human development1-4. However, these simple culture systems lack the complexity to adequately model the spatiotemporal cellular and molecular dynamics that occur during early embryonic development. Here we describe the reprogramming of fibroblasts into in vitro three-dimensional models of the human blastocyst, termed iBlastoids. Characterization of iBlastoids shows that they model the overall architecture of blastocysts, presenting an inner cell mass-like structure, with epiblast- and primitive endoderm-like cells, a blastocoel-like cavity and a trophectoderm-like outer layer of cells. Single-cell transcriptomics further confirmed the presence of epiblast-, primitive endoderm-, and trophectoderm-like cells. Moreover, iBlastoids can give rise to pluripotent and trophoblast stem cells and are capable of modelling, in vitro, several aspects of the early stage of implantation. In summary, we have developed a scalable and tractable system to model human blastocyst biology; we envision that this will facilitate the study of early human development and the effects of gene mutations and toxins during early embryogenesis, as well as aiding in the development of new therapies associated with in vitro fertilization.

In-depth analysis of multi- and extensively drug-resistant mycobacterium tuberculosis isolates in the capital of iran

Aims and objectives: The emergence and spread of multidrug resistant (MDR) Mycobacterium tuberculosis (M. tuberculosis) strains is a critical global health problem. In order to shorten the course of treatment and its effectiveness, it is essential to gain an in-depth insight into the genetic diversity, drug resistance mechanisms and host-pathogen interactions of drug resistant M. tuberculosis. Methods: Between 2014 and 2018, 606 MTBC strains were isolated from 13,892 suspected pulmonary tuberculosis (TB) patients in Tehran, Iran, including 16 MDR-TB cases. A combination of phenotypic (conventional drug susceptibility testing, MIC, Verapamil effectiveness) and genotypic methods (whole-genome sequencing and qPCR) was employed for the identification of additional drug resistances, the role of efflux pumps and strain-to-strain genetic distances as a marker for recent transmission events. Finally, the consequences of mutations in genes encoding the beta subunit of RNA polymerase (rpoB) for host-pathogen interactions was investigated by means of THP-1-derived macrophages infection and Cytokines and Chemokines Real-Time RT2 Profiler PCR Array. Results: MDR and extensively drug-resistant (XDR) TB cases were almost exclusively infected by lineage 2/Beijing strains (14/16, P &lt; 0.001). Out of the 16 MDR/pre-XDR/XDR isolates, 6 (37.5%) and 3 (18.8%) isolates demonstrated a significant decrease in rifampicin (RIF) MIC and isoniazid (INH) MIC due to the Verapamil exposure (64 μg/mL), respectively. More importantly, it was found that the administration of Verapamil profoundly decreased the MIC of Bedaquiline (BDQ) among most of the studied clinical isolates. Moreover, the efflux pump genes of Rv2938, Rv2936, Rv1145, Rv1146, Rv933, Rv1250, Rv876, Rv2333, Rv2459, Rv849, and Rv1819 were overexpressed in the presence of anti-TB drugs, showing the contribution of these efflux pumps to the overall resistance phenotype. Finally, the macrophages immune responses induced in response to infection with drug-resistant M. tuberculosis (RMR strain) may not be substantially different from drug susceptible M. tuberculosis (H37Rv strain), but the severity of the responses was different in some cases specially IL-6, IL-8, CNTF, IL-13, TNFSF10 and CD40LG in response to RMR and H37Rv strains. In addition, RMR strain showing significantly higher intracellular bacillary growth compared to H37Rv strain. Conclusions: The crucial finding is that almost all of our MDR/pre-XDR/XDR strains are of the Beijing lineage (14/16) and more than half of them are probably from recent transmission (8/14). Our results clearly showed that efflux systems, besides spontaneous mutations, play a role in the development of INH/RIF resistance. In addition, Verapamil was very successful at the phenotypic level specially in response to BDQ. Finally, the results initially demonstrate that the M. tuberculosis carrying the rpoB-S450L can modulate macrophage responses to mediate bacterial survival.