scholarly journals A Genome-Wide Proteome Array Reveals a Limited Set of Immunogens in Natural Infections of Humans and White-Footed Mice with Borrelia burgdorferi

2008 ◽  
Vol 76 (8) ◽  
pp. 3374-3389 ◽  
Author(s):  
Alan G. Barbour ◽  
Algimantas Jasinskas ◽  
Matthew A. Kayala ◽  
D. Huw Davies ◽  
Allen C. Steere ◽  
...  
Keyword(s):  
Borrelia Burgdorferi ◽  
Lyme Borreliosis ◽  
Flagellar Apparatus ◽  
Reading Frame ◽  
High Ranking ◽  
Paralogous Family ◽  
Genome Wide ◽  
A Genome ◽  
Statistical Criteria

ABSTRACT Humans and other animals with Lyme borreliosis produce antibodies to a number of components of the agent Borrelia burgdorferi, but a full accounting of the immunogens during natural infections has not been achieved. Employing a protein array produced in vitro from 1,292 DNA fragments representing ∼80% of the genome, we compared the antibody reactivities of sera from patients with early or later Lyme borreliosis to the antibody reactivities of sera from controls. Overall, ∼15% of the open reading frame (ORF) products (Orfs) of B. burgdorferi in the array detectably elicited an antibody response in humans with natural infections. Among the immunogens, 103 stood out on the basis of statistical criteria. The majority of these Orfs were also immunogenic with sera obtained from naturally infected Peromyscus leucopus mice, a major reservoir. The high-ranking set included several B. burgdorferi proteins hitherto unrecognized as immunogens, as well as several proteins that have been established as antigens. The high-ranking immunogens were more likely than nonreactive Orfs to have the following characteristics: (i) plasmid-encoded rather than chromosome-encoded proteins, (ii) a predicted lipoprotein, and (iii) a member of a paralogous family of proteins, notably the Bdr and Erp proteins. The newly discovered antigens included Orfs encoded by several ORFs of the lp36 linear plasmid, such as BBK07 and BBK19, and proteins of the flagellar apparatus, such as FliL. These results indicate that the majority of deduced proteins of B. burgdorferi do not elicit antibody responses during infection and that the limited sets of immunogens are similar for two different host species.

Transcriptional enhancers: from prediction to functional assessment on a genome-wide scale

Genome ◽  
2020 ◽  
pp. 1-23
Author(s):  
Ian C. Tobias ◽  
Luis E. Abatti ◽  
Sakthi D. Moorthy ◽  
Shanelle Mullany ◽  
Tiegh Taylor ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Target Genes ◽  
Regulatory Sequences ◽  
Enhancer Activity ◽  
Reading Frame ◽  
Enhancer Prediction ◽  
Genome Wide ◽  
A Genome ◽  
Enhancer Function

Enhancers are cis-regulatory sequences located distally to target genes. These sequences consolidate developmental and environmental cues to coordinate gene expression in a tissue-specific manner. Enhancer function and tissue specificity depend on the expressed set of transcription factors, which recognize binding sites and recruit cofactors that regulate local chromatin organization and gene transcription. Unlike other genomic elements, enhancers are challenging to identify because they function independently of orientation, are often distant from their promoters, have poorly defined boundaries, and display no reading frame. In addition, there are no defined genetic or epigenetic features that are unambiguously associated with enhancer activity. Over recent years there have been developments in both empirical assays and computational methods for enhancer prediction. We review genome-wide tools, CRISPR advancements, and high-throughput screening approaches that have improved our ability to both observe and manipulate enhancers in vitro at the level of primary genetic sequences, chromatin states, and spatial interactions. We also highlight contemporary animal models and their importance to enhancer validation. Together, these experimental systems and techniques complement one another and broaden our understanding of enhancer function in development, evolution, and disease.

scholarly journals Genome-wide CRISPR Screen to Identify Genes that Suppress Transformation in the Presence of Endogenous KrasG12D

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jianguo Huang ◽  
Mark Chen ◽  
Eric S. Xu ◽  
Lixia Luo ◽  
Yan Ma ◽  
...  
Keyword(s):  
Kinase Inhibitor ◽  
Gene Mutations ◽  
Soft Agar ◽  
Reading Frame ◽  
Immunodeficient Mice ◽  
Genome Wide ◽  
A Genome ◽  
Crispr Screen

AbstractCooperating gene mutations are typically required to transform normal cells enabling growth in soft agar or in immunodeficient mice. For example, mutations in Kras and transformation-related protein 53 (Trp53) are known to transform a variety of mesenchymal and epithelial cells in vitro and in vivo. Identifying other genes that can cooperate with oncogenic Kras and substitute for Trp53 mutation has the potential to lead to new insights into mechanisms of carcinogenesis. Here, we applied a genome-wide CRISPR/Cas9 knockout screen in KrasG12D immortalized mouse embryonic fibroblasts (MEFs) to search for genes that when mutated cooperate with oncogenic Kras to induce transformation. We also tested if mutation of the identified candidate genes could cooperate with KrasG12D to generate primary sarcomas in mice. In addition to identifying the well-known tumor suppressor cyclin dependent kinase inhibitor 2A (Cdkn2a), whose alternative reading frame product p19 activates Trp53, we also identified other putative tumor suppressors, such as F-box/WD repeat-containing protein 7 (Fbxw7) and solute carrier family 9 member 3 (Slc9a3). Remarkably, the TCGA database indicates that both FBXW7 and SLC9A3 are commonly co-mutated with KRAS in human cancers. However, we found that only mutation of Trp53 or Cdkn2a, but not Fbxw7 or Slc9a3 can cooperate with KrasG12D to generate primary sarcomas in mice. These results show that mutations in oncogenic Kras and either Fbxw7 or Slc9a3 are sufficient for transformation in vitro, but not for in vivo sarcomagenesis.

scholarly journals Transcriptome analysis ofSchistosoma mansonilarval development using serial analysis of gene expression (SAGE)

Parasitology ◽  
2009 ◽  
Vol 136 (5) ◽  
pp. 469-485 ◽  
Author(s):  
A. S. TAFT ◽  
J. J. VERMEIRE ◽  
J. BERNIER ◽  
S. R. BIRKELAND ◽  
M. J. CIPRIANO ◽  
...  
Keyword(s):  
Gene Expression ◽  
Sequence Data ◽  
Subsequent Development ◽  
Differentially Expressed ◽  
Cdna Libraries ◽  
Genome Wide ◽  
A Genome ◽  
Genome Wide Expression ◽  
Cell Conditioned Medium

SUMMARYInfection of the snail,Biomphalaria glabrata, by the free-swimming miracidial stage of the human blood fluke,Schistosoma mansoni, and its subsequent development to the parasitic sporocyst stage is critical to establishment of viable infections and continued human transmission. We performed a genome-wide expression analysis of theS. mansonimiracidia and developing sporocyst using Long Serial Analysis of Gene Expression (LongSAGE). Five cDNA libraries were constructed from miracidia andin vitrocultured 6- and 20-day-old sporocysts maintained in sporocyst medium (SM) or in SM conditioned by previous cultivation with cells of theB. glabrataembryonic (Bge) cell line. We generated 21 440 SAGE tags and mapped 13 381 to theS. mansonigene predictions (v4.0e) either by estimating theoretical 3′ UTR lengths or using existing 3′ EST sequence data. Overall, 432 transcripts were found to be differentially expressed amongst all 5 libraries. In total, 172 tags were differentially expressed between miracidia and 6-day conditioned sporocysts and 152 were differentially expressed between miracidia and 6-day unconditioned sporocysts. In addition, 53 and 45 tags, respectively, were differentially expressed in 6-day and 20-day cultured sporocysts, due to the effects of exposure to Bge cell-conditioned medium.

scholarly journals The FAM171A2 gene is a key regulator of progranulin expression and modifies the risk of multiple neurodegenerative diseases

2020 ◽  
Vol 6 (43) ◽  
pp. eabb3063
Author(s):  
Wei Xu ◽  
Si-Da Han ◽  
Can Zhang ◽  
Jie-Qiong Li ◽  
Yan-Jiang Wang ◽  
...  
Keyword(s):  
Neurodegenerative Diseases ◽  
Genome Wide Association Study ◽  
In Vitro Study ◽  
Genome Wide ◽  
A Genome ◽  
Increased Risk ◽  
Pathophysiological Role ◽  
Independent Cohort

Progranulin (PGRN) is a secreted pleiotropic glycoprotein associated with the development of common neurodegenerative diseases. Understanding the pathophysiological role of PGRN may help uncover biological underpinnings. We performed a genome-wide association study to determine the genetic regulators of cerebrospinal fluid (CSF) PGRN levels. Common variants in region of FAM171A2 were associated with lower CSF PGRN levels (rs708384, P = 3.95 × 10−12). This was replicated in another independent cohort. The rs708384 was associated with increased risk of Alzheimer’s disease, Parkinson’s disease, and frontotemporal dementia and could modify the expression of the FAM171A2 gene. FAM171A2 was considerably expressed in the vascular endothelium and microglia, which are rich in PGRN. The in vitro study further confirmed that the rs708384 mutation up-regulated the expression of FAM171A2, which caused a decrease in the PGRN level. Collectively, genetic, molecular, and bioinformatic findings suggested that FAM171A2 is a key player in regulating PGRN production.

scholarly journals Complete biosynthetic pathways of ascofuranone and ascochlorin inAcremonium egyptiacum

2019 ◽  
Vol 116 (17) ◽  
pp. 8269-8274 ◽  
Author(s):  
Yasuko Araki ◽  
Takayoshi Awakawa ◽  
Motomichi Matsuzaki ◽  
Rihe Cho ◽  
Yudai Matsuda ◽  
...  
Keyword(s):  
Differential Expression Analysis ◽  
Cost Effective ◽  
Drug Candidate ◽  
P450 Monooxygenase ◽  
Genome Wide ◽  
Common Precursor ◽  
A Genome ◽  
Membrane Bound ◽  
The Cost

Ascofuranone (AF) and ascochlorin (AC) are meroterpenoids produced by various filamentous fungi, includingAcremonium egyptiacum(synonym:Acremonium sclerotigenum), and exhibit diverse physiological activities. In particular, AF is a promising drug candidate against African trypanosomiasis and a potential anticancer lead compound. These compounds are supposedly biosynthesized through farnesylation of orsellinic acid, but the details have not been established. In this study, we present all of the reactions and responsible genes for AF and AC biosyntheses inA. egyptiacum, identified by heterologous expression, in vitro reconstruction, and gene deletion experiments with the aid of a genome-wide differential expression analysis. Both pathways share the common precursor, ilicicolin A epoxide, which is processed by the membrane-bound terpene cyclase (TPC) AscF in AC biosynthesis. AF biosynthesis branches from the precursor by hydroxylation at C-16 by the P450 monooxygenase AscH, followed by cyclization by a membrane-bound TPC AscI. All genes required for AC biosynthesis (ascABCDEFG) and a transcriptional factor (ascR) form a functional gene cluster, whereas those involved in the late steps of AF biosynthesis (ascHIJ) are present in another distantly located cluster. AF is therefore a rare example of fungal secondary metabolites requiring multilocus biosynthetic clusters, which are likely to be controlled by the single regulator, AscR. Finally, we achieved the selective production of AF inA. egyptiacumby genetically blocking the AC biosynthetic pathway; further manipulation of the strain will lead to the cost-effective mass production required for the clinical use of AF.

In vitro-targeted gene identification in patients with hepatitis C using a genome-wide microarray technology

Hepatology ◽  
2008 ◽  
Vol 49 (2) ◽  
pp. 378-386 ◽  
Author(s):  
Susanne Hagist ◽  
Holger Sültmann ◽  
Gunda Millonig ◽  
Ulrike Hebling ◽  
Dörthe Kieslich ◽  
...  
Keyword(s):  
Hepatitis C ◽  
Gene Identification ◽  
Microarray Technology ◽  
Genome Wide ◽  
A Genome

scholarly journals Abnormal expression of homeobox genes and transthyretin in C9ORF72 expansion carriers

2017 ◽  
Vol 3 (4) ◽  
pp. e161 ◽  
Author(s):  
NiCole A. Finch ◽  
Xue Wang ◽  
Matthew C. Baker ◽  
Michael G. Heckman ◽  
Tania F. Gendron ◽  
...  
Keyword(s):  
Neuronal Development ◽  
Homeobox Genes ◽  
Vital Role ◽  
Chromosome 9 ◽  
Compensatory Mechanisms ◽  
Reading Frame ◽  
Genome Wide ◽  
A Genome ◽  
Brain Expression ◽  
Genome Wide Expression

Objective:We performed a genome-wide brain expression study to reveal the underpinnings of diseases linked to a repeat expansion in chromosome 9 open reading frame 72 (C9ORF72).Methods:The genome-wide expression profile was investigated in brain tissue obtained from C9ORF72 expansion carriers (n = 32), patients without this expansion (n = 30), and controls (n = 20). Using quantitative real-time PCR, findings were confirmed in our entire pathologic cohort of expansion carriers (n = 56) as well as nonexpansion carriers (n = 31) and controls (n = 20).Results:Our findings were most profound in the cerebellum, where we identified 40 differentially expressed genes, when comparing expansion carriers to patients without this expansion, including 22 genes that have a homeobox (e.g., HOX genes) and/or are located within the HOX gene cluster (top hit: homeobox A5 [HOXA5]). In addition to the upregulation of multiple homeobox genes that play a vital role in neuronal development, we noticed an upregulation of transthyretin (TTR), an extracellular protein that is thought to be involved in neuroprotection. Pathway analysis aligned with these findings and revealed enrichment for gene ontology processes involved in (anatomic) development (e.g., organ morphogenesis). Additional analyses uncovered that HOXA5 and TTR levels are associated with C9ORF72 variant 2 levels as well as with intron-containing transcript levels, and thus, disease-related changes in those transcripts may have triggered the upregulation of HOXA5 and TTR.Conclusions:In conclusion, our identification of genes involved in developmental processes and neuroprotection sheds light on potential compensatory mechanisms influencing the occurrence, presentation, and/or progression of C9ORF72-related diseases.

scholarly journals Genomic variation across a clinical Cryptococcus population linked to disease outcome

2021 ◽  
Author(s):  
Poppy Channa Sakti Sephton-Clark ◽  
Jennifer Tenor ◽  
Dena Toffaletti ◽  
Nancy Meyers ◽  
Charles Giamberardino ◽  
...  
Keyword(s):  
Patient Outcomes ◽  
Large Scale ◽  
Genome Wide Association Study ◽  
Sugar Transport ◽  
Genomic Variation ◽  
Clinical Isolates ◽  
Genome Wide ◽  
A Genome ◽  
Gwas Approach

Cryptococcus neoformans is the causative agent of cryptococcosis, a disease with poor patient outcomes, accounting for approximately 180,000 deaths each year. Patient outcomes may be impacted by the underlying genetics of the infecting isolate, however, our current understanding of how genetic diversity contributes to clinical outcomes is limited. Here, we leverage clinical, in vitro growth and genomic data for 284 C. neoformans isolates to identify clinically relevant pathogen variants within a population of clinical isolates from patients with HIV-associated cryptococcosis in Malawi. Through a genome-wide association study (GWAS) approach, we identify variants associated with fungal burden and growth rate. We also find both small and large-scale variation, including aneuploidy, associated with alternate growth phenotypes, which may impact the course of infection. Genes impacted by these variants are involved in transcriptional regulation, signal transduction, glycolysis, sugar transport, and glycosylation. When combined with clinical data, we show that growth within the CNS is reliant upon glycolysis in an animal model, and likely impacts patient mortality, as CNS burden modulates patient outcome. Additionally, we find genes with roles in sugar transport are under selection in the majority of these clinical isolates. Further, we demonstrate that two hypothetical proteins identified by GWAS impact virulence in animal models. Our approach illustrates links between genetic variation and clinically relevant phenotypes, shedding light on survival mechanisms within the CNS and pathways involved in this persistence.

scholarly journals Natural selection driven by DNA binding proteins shapes genome-wide motif statistics

2016 ◽  
Author(s):  
Long Qian ◽  
Edo Kussell
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Binding Proteins ◽  
Dna Binding Proteins ◽  
Large Collection ◽  
Functional Sites ◽  
Genome Wide ◽  
A Genome ◽  
Global Selection

AbstractEctopic DNA binding by transcription factors and other DNA binding proteins can be detrimental to cellular functions and ultimately to organismal fitness. The frequency of protein-DNA binding at non-functional sites depends on the global composition of a genome with respect to all possible short motifs, or k-mer words. To determine whether weak yet ubiquitous protein-DNA interactions could exert significant evolutionary pressures on genomes, we correlate in vitro measurements of binding strengths on all 8-mer words from a large collection of transcription factors, in several different species, against their relative genomic frequencies. Our analysis reveals a clear signal of purifying selection to reduce the large number of weak binding sites genome-wide. This evolutionary process, which we call global selection, has a detectable hallmark in that similar words experience similar evolutionary pressure, a consequence of the biophysics of protein-DNA binding. By analyzing a large collection of genomes, we show that global selection exists in all domains of life, and operates through tiny selective steps, maintaining genomic binding landscapes over long evolutionary timescales.

scholarly journals Genome-Wide Screening, Cloning, Chromosomal Assignment, and Expression of Full-Length Human Endogenous Retrovirus Type K

1999 ◽  
Vol 73 (11) ◽  
pp. 9187-9195 ◽  
Author(s):  
Ralf R. Tönjes ◽  
Frank Czauderna ◽  
Reinhard Kurth
Keyword(s):  
Human Genome ◽  
Endogenous Retrovirus ◽  
Full Length ◽  
In Vitro Translation ◽  
Type K ◽  
Genome Wide ◽  
A Genome

ABSTRACT The human genome harbors 25 to 50 proviral copies of the endogenous retrovirus type K (HERV-K), some of which code for the characteristic retroviral proteins Gag, Pol, and Env. For a genome-wide cloning approach of full-length and intact HERV-K proviruses, a human P1 gene library was screened with a gag-specific probe. Both HERV-K type 1 and 2 clones were isolated. Sixteen HERV-K type 2 proviral genomes were characterized by direct coupled in vitro transcription-in vitro translation assays to analyze the coding potential of isolatedgag, pol, and env amplicons from individual P1 clones. After determination of long terminal repeat (LTR) sequences and adjacent chromosomal integration sites by inverse PCR techniques, two HERV-K type 2 proviruses displaying long retroviral open reading frames (ORFs) were assigned to chromosomes 7 (C7) and 19 (C19) by using a human-rodent monochromosomal cell hybrid mapping panel. HERV-K(C7) shows an altered (YIDD-to-CIDD) motif in the reverse transcriptase domain. HERV-K(C19) is truncated in the 5′ LTR and harbors a defective protease gene due to a point mutation. Direct amplification of proviral structures from single chromosomes by using chromosomal flanking primers was performed by long PCR for HERV-K(C7) and HERV-K(C19) and for type 1 proviruses HERV-K10 and HERV-K18 from chromosomes 5 and 1, respectively. HERV-K18, in contrast to HERV-K10, bears no intact gag ORF and shows close homology to HERV-K/IDDMK1,222. In transfection experiments, HERV-K(C7) and HERV-K cDNA-based expression vectors yielded the proteins Gag and cORF whereas HERV-K10 vectors yielded Gag alone. The data suggest that the human genome does not contain an entire, intact proviral copy of HERV-K.

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