scholarly journals Antibody-mediated depletion of protein variant expression in living cells (Protein interference)

2017 ◽  
Author(s):  
Vijay K. Ulaganathan ◽  
Axel Ullrich
Keyword(s):  
Protein Sequence ◽  
Sequence Variants ◽  
Intracellular Protein ◽  
Protein Variant ◽  
Protein Coding ◽  
Significant Development ◽  
Human Genomes ◽  
Reference Protein ◽  
Allele Specific ◽  
Protein Variants

AbstractA significant development in the field of human biology is the revelation of millions of unannotated protein sequence variants emerging from the several human genotyping and genome sequencing initiatives. This presents unique opportunities as well as confounding challenges in our understanding of how molecular signalling outcomes vary among individuals in the general population. As a result the conventional ‘one drug fits all’ lines of approach in the drug discovery process is becoming obsolete. However, an innovative genotype-specific approach targeting protein sequence variants instead of a reference protein target is currently lacking. In this short communication we report a remarkable observation of antibody-mediated knockdown of intracellular protein expression. This suggests allele-specific inhibition of protein-variant expression can be achieved by intracellular delivery of lipid conjugated linear epitope-specific monoclonal antibodies. The results presented here demonstrate novel opportunities for interrogating the protein coding variations in the human genomes and new therapeutic strategies for the inhibition of pathogenic protein variants in a genotype-centric manner.

Identification of Two Tobacco streak virus Capsid Protein Variants Associated With Leaf Mottle and Necrosis Symptoms on Astilbe

2013 ◽  
Vol 14 (1) ◽  
pp. 50 ◽  
Author(s):  
John R. Fisher
Keyword(s):  
Capsid Protein ◽  
Protein Sequence ◽  
Tobacco Streak Virus ◽  
Streak Virus ◽  
Sequence Variants ◽  
Virus Capsid ◽  
Capsid Protein Sequence ◽  
Protein Variants ◽  
Virus Capsid Protein

In this brief we report the identification of two Tobacco streak virus capsid protein sequence variants associated with leaf mottle and necrosis symptoms on Astilbe × arendsii (false spirea). To our knowledge this is the first confirmed report of TSV infecting Astilbe. Accepted for publication 23 April 2013. Published 29 July 2013.

scholarly journals Using diverse U.S. beef cattle genomes to identify missense mutations in EPAS1, a gene associated with pulmonary hypertension

F1000Research ◽  
2016 ◽  
Vol 5 ◽  
pp. 2003 ◽  
Author(s):  
Michael P. Heaton ◽  
Timothy P.L. Smith ◽  
Jacky K. Carnahan ◽  
Veronica Basnayake ◽  
Jiansheng Qiu ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Beef Cattle ◽  
In Silico ◽  
Read Depth ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Missense Mutations ◽  
Protein Variant ◽  
Flight Mass Spectrometry ◽  
Protein Variants

The availability of whole genome sequence (WGS) data has made it possible to discover protein variantsin silico. However, existing bovine WGS databases do not show data in a form conducive to protein variant analysis, and tend to under represent the breadth of genetic diversity in global beef cattle. Thus, our first aim was to use 96 beef sires, sharing minimal pedigree relationships, to create a searchable and publicly viewable set of mapped genomes relevant for 19 popular breeds of U.S. cattle. Our second aim was to identify protein variants encoded by the bovine endothelial PAS domain-containing protein 1 gene (EPAS1), a gene associated with pulmonary hypertension in Angus cattle. The identity and quality of genomic sequences were verified by comparing WGS genotypes to those derived from other methods. The average read depth, genotype scoring rate, and genotype accuracy exceeded 14, 99%, and 99%, respectively. The 96 genomes were used to discover four amino acid variants encoded byEPAS1(E270Q, P362L, A671G, and L701F) and confirm two variants previously associated with disease (A606T and G610S). The sixEPAS1missense mutations were verified with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry assays, and their frequencies were estimated in a separate collection of 1154 U.S. cattle representing 46 breeds. A rooted phylogenetic tree of eight polypeptide sequences provided a framework for evaluating the likely order of mutations and potential impact ofEPAS1alleles on the adaptive response to chronic hypoxia in U.S. cattle. This public, whole genome resource facilitatesin silicoidentification of protein variants in diverse types of U.S. beef cattle, and provides a means of translating WGS data into a practical biological and evolutionary context for generating and testing hypotheses.

Physiological Dysfunctions Associated with Mutations of the Imprinted Gnas Locus

Physiology ◽  
2008 ◽  
Vol 23 (4) ◽  
pp. 221-229 ◽  
Author(s):  
Stefan Krechowec ◽  
Antonius Plagge
Keyword(s):  
Signal Transduction ◽  
Mouse Models ◽  
Genomic Organization ◽  
Receptor Activation ◽  
Deficient Mouse ◽  
Protein Coding ◽  
Alternative Protein ◽  
Stimulatory G Protein ◽  
Protein Variants ◽  
Complex Arrangement

The ubiquitous Gαs-subunit of the trimeric, stimulatory G-protein plays a central role in receptor-mediated signal transduction, coupling receptor activation with the production of cAMP. The Gαs-encoding locus Gnas is now known to consist of a complex arrangement of several protein-coding and noncoding transcripts. We provide an overview of its genomic organization, its regulation by genomic imprinting, and a summary of the physiological roles of the alternative protein variants Gαs and XLαs as determined from deficient mouse models.

scholarly journals Understanding the Early Evolutionary Stages of a Tandem Drosophilamelanogaster-Specific Gene Family: A Structural and Functional Population Study

2020 ◽  
Vol 37 (9) ◽  
pp. 2584-2600 ◽  
Author(s):  
Bryan D Clifton ◽  
Jamie Jimenez ◽  
Ashlyn Kimura ◽  
Zeinab Chahine ◽  
Pablo Librado ◽  
...  
Keyword(s):  
Gene Family ◽  
Sequence Similarity ◽  
Gene Families ◽  
Read Depth ◽  
Specific Gene ◽  
Protein Variant ◽  
Protein Coding ◽  
Expression Levels ◽  
Number Variation ◽  
Reference Quality

Abstract Gene families underlie genetic innovation and phenotypic diversification. However, our understanding of the early genomic and functional evolution of tandemly arranged gene families remains incomplete as paralog sequence similarity hinders their accurate characterization. The Drosophila melanogaster-specific gene family Sdic is tandemly repeated and impacts sperm competition. We scrutinized Sdic in 20 geographically diverse populations using reference-quality genome assemblies, read-depth methodologies, and qPCR, finding that ∼90% of the individuals harbor 3–7 copies as well as evidence of population differentiation. In strains with reliable gene annotations, copy number variation (CNV) and differential transposable element insertions distinguish one structurally distinct version of the Sdic region per strain. All 31 annotated copies featured protein-coding potential and, based on the protein variant encoded, were categorized into 13 paratypes differing in their 3′ ends, with 3–5 paratypes coexisting in any strain examined. Despite widespread gene conversion, the only copy present in all strains has functionally diverged at both coding and regulatory levels under positive selection. Contrary to artificial tandem duplications of the Sdic region that resulted in increased male expression, CNV in cosmopolitan strains did not correlate with expression levels, likely as a result of differential genome modifier composition. Duplicating the region did not enhance sperm competitiveness, suggesting a fitness cost at high expression levels or a plateau effect. Beyond facilitating a minimally optimal expression level, Sdic CNV acts as a catalyst of protein and regulatory diversity, showcasing a possible evolutionary path recently formed tandem multigene families can follow toward long-term consolidation in eukaryotic genomes.

scholarly journals Evolution of Human Brain Size-Associated NOTCH2NL Genes Proceeds toward Reduced Protein Levels

2020 ◽  
Vol 37 (9) ◽  
pp. 2531-2548
Author(s):  
Gerrald A Lodewijk ◽  
Diana P Fernandes ◽  
Iraklis Vretzakis ◽  
Jeanne E Savage ◽  
Frank M J Jacobs
Keyword(s):  
Human Brain ◽  
Brain Size ◽  
Modern Human ◽  
Copy Number Variations ◽  
Optimal Dosage ◽  
Protein Coding ◽  
Protein Levels ◽  
Human Genomes ◽  
Coding Variants ◽  
Human Specific

Abstract Ever since the availability of genomes from Neanderthals, Denisovans, and ancient humans, the field of evolutionary genomics has been searching for protein-coding variants that may hold clues to how our species evolved over the last ∼600,000 years. In this study, we identify such variants in the human-specific NOTCH2NL gene family, which were recently identified as possible contributors to the evolutionary expansion of the human brain. We find evidence for the existence of unique protein-coding NOTCH2NL variants in Neanderthals and Denisovans which could affect their ability to activate Notch signaling. Furthermore, in the Neanderthal and Denisovan genomes, we find unusual NOTCH2NL configurations, not found in any of the modern human genomes analyzed. Finally, genetic analysis of archaic and modern humans reveals ongoing adaptive evolution of modern human NOTCH2NL genes, identifying three structural variants acting complementary to drive our genome to produce a lower dosage of NOTCH2NL protein. Because copy-number variations of the 1q21.1 locus, encompassing NOTCH2NL genes, are associated with severe neurological disorders, this seemingly contradicting drive toward low levels of NOTCH2NL protein indicates that the optimal dosage of NOTCH2NL may have not yet been settled in the human population.

scholarly journals Posttranslational Modification of the 20S Proteasomal Proteins of the Archaeon Haloferax volcanii

2006 ◽  
Vol 188 (21) ◽  
pp. 7521-7530 ◽  
Author(s):  
Matthew A. Humbard ◽  
Stanley M. Stevens ◽  
Julie A. Maupin-Furlow
Keyword(s):  
Posttranslational Modification ◽  
Protein Sequence ◽  
Phosphorylation Site ◽  
Haloferax Volcanii ◽  
20S Proteasome ◽  
Β Subunit ◽  
Halophilic Archaeon ◽  
Intracellular Protein ◽  
Flight Mass Spectrometry ◽  
Intracellular Protein Degradation

ABSTRACT 20S proteasomes are large, multicatalytic proteases that play an important role in intracellular protein degradation. The barrel-like architecture of 20S proteasomes, formed by the stacking of four heptameric protein rings, is highly conserved from archaea to eukaryotes. The outer two rings are composed of α-type subunits, and the inner two rings are composed of β-type subunits. The halophilic archaeon Haloferax volcanii synthesizes two different α-type proteins, α1 and α2, and one β-type protein that assemble into at least two 20S proteasome subtypes. In this study, we demonstrate that all three of these 20S proteasomal proteins (α1, α2, and β) are modified either post- or cotranslationally. Using electrospray ionization quadrupole time-of-flight mass spectrometry, a phosphorylation site of the β subunit was identified at Ser129 of the deduced protein sequence. In addition, α1 and α2 contained N-terminal acetyl groups. These findings represent the first evidence of acetylation and phosphorylation of archaeal proteasomes and are one of the limited examples of post- and/or cotranslational modification of proteins in this unusual group of organisms.

scholarly journals Yeast Models Of Phosphomannomutase 2 Deficiency, A Congenital Disorder Of Glycosylation

2018 ◽  
Author(s):  
Jessica P. Lao ◽  
Nina DiPrimio ◽  
Madeleine Prangley ◽  
Feba S. Sam ◽  
Joshua D. Mast ◽  
...  
Keyword(s):  
Congenital Disorder ◽  
Yeast Cells ◽  
Model Organisms ◽  
Missense Mutations ◽  
Loss Of Function ◽  
Protein Variant ◽  
Congenital Disorder Of Glycosylation ◽  
Experimental Drugs ◽  
Protein Variants ◽  
Phosphomannomutase 2

AbstractPhosphomannomutase 2 Deficiency (PMM2-CDG) is the most common monogenic congenital disorder of glycosylation (CDG) affecting at least 800 patients globally. PMM2 orthologs are present in model organisms, including the budding yeast Saccharomyces cerevisiae gene SEC53. Here we describe conserved genotype-phenotype relationships across yeast and human patients between five PMM2 loss-of-function missense mutations and their orthologous SEC53 mutations. These alleles range in severity from folding defective (hypomorph) to dimerization defective (severe hypomorph) to catalytic dead (null). We included the first and second most common missense mutations – R141H, F119L respectively– and the most common compound heterozygote genotype – PMM2R141H/F119L – observed in PMM2-CDG patients. Each mutation described is expressed in haploid as well as homozygous and heterozygous diploid yeast cells at varying protein expression levels as either SEC53 protein variants or PMM2 protein variants. We developed a 384-well-plate, growth-based assay for use in a screen of the 2,560-compound Microsource Spectrum library of approved drugs, experimental drugs, tool compounds and natural products. We identified three compounds that suppress growth defects of SEC53 variants, F126L and V238M, based on the biochemical defect of the allele, protein abundance or ploidy. The rare PMM2 E139K protein variant is fully functional in yeast cells, suggesting that its pathogenicity in humans is due to the underlying DNA mutation that results in skipping of exon 5 and a nonfunctional truncated protein. Together, these results demonstrate that yeast models can be used to characterize known and novel PMM2 patient alleles in quantitative growth and enzymatic activity assays, and used as patient avatars for PMM2-CDG drug screens yielding compounds that could be rapidly cross-validated in zebrafish, rodent and human organoid models.

scholarly journals HICL table can manipulate all proteins in human complete proteome

2016 ◽  
Author(s):  
Zhenhua Xie
Keyword(s):  
Amino Acid ◽  
Protein Sequence ◽  
Family Members ◽  
Distribution Patterns ◽  
Acid Properties ◽  
Amino Acid Properties ◽  
Isoelectric Points ◽  
Physicochemical Features ◽  
Protein Variants ◽  
Universal Protein

AbstractBackgroundThe data of human complete proteome in the databases of Universal Protein Resource (UniProt) or National Center for Biotechnology Information(NCBI) were disorderly organized and hardly handled by an ordinary biologist.ResultsThe HICL table enable an ordinary biologist efficiently to handle the human complete proteome with 67911 entries, to get an overview on the distribution of the physicochemical features of all proteins in the human complete proteome, to perceive the details of the distribution patterns of the physicochemical features in some protein family members and protein variants, to find some particular proteins.Moreover, two discoveries were made via the HICL table: (1) The amino aicds(Asp,Glu) have symmetrical trend of the distributions versus pI, but the amino aicds(Arg, Lys) have local asymmetrical trend of the distributions versus pI in human complete proteome. (2) Protein sequence, besides amino acid properties, can in theory influence the modal distribution of protein isoelectric points.ConclusionI has created the HICL table as a robust tool for orderly managing 67911 proteins in human complete proteome by their physicochemical features, the names and sequences. Any proteins with the particular physicochemical features can be screened out from the human complete proteome via the HICL table. In addition, the unbalanced distribution of the amino aicds(Arg, Lys) in high pI proteins of human complete proteome and the effect of protein sequence on modal distribution of protein isoelectric points have been discovered through the HICL table.

scholarly journals AlleleAnalyzer: a tool for personalized and allele-specific sgRNA design

2018 ◽  
Author(s):  
Kathleen C. Keough ◽  
Svetlana Lyalina ◽  
Michael P. Olvera ◽  
Sean Whalen ◽  
Bruce R. Conklin ◽  
...  
Keyword(s):  
Genome Editing ◽  
Reference Genome ◽  
Individual Genome ◽  
Experimental Performance ◽  
Human Genomes ◽  
Large Populations ◽  
Allele Specific ◽  
Sgrna Design ◽  
Single Base Pair ◽  
The Individual

AbstractThe CRISPR/Cas system is a highly specific genome editing tool capable of distinguishing alleles differing by even a single base pair. However, current tools only design sgRNAs for a reference genome, not taking into account individual variants which may generate, remove, or modify CRISPR/Cas sgRNA sites. This may cause mismatches between designed sgRNAs and the individual genome they are intended to target, leading to decreased experimental performance. Here we describe AlleleAnalyzer, a tool for designing personalized and allele-specific sgRNAs for genome editing. We leverage >2,500 human genomes to identify optimized pairs of sgRNAs that can be used for human therapeutic editing in large populations in the future.

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