scholarly journals Quantitative Proteomics Comparison of Total Expressed Proteomes of Anisakis simplex Sensu Stricto, A. pegreffii, and Their Hybrid Genotype

Genes ◽  
2020 ◽  
Vol 11 (8) ◽  
pp. 913
Author(s):  
Susana C. Arcos ◽  
Lee Robertson ◽  
Sergio Ciordia ◽  
Isabel Sánchez-Alonso ◽  
Mercedes Careche ◽  
...  
Keyword(s):  
Quantitative Proteomics ◽  
Protein A ◽  
Principal Component ◽  
Sensu Stricto ◽  
Anisakis Simplex ◽  
Hybrid Genotype ◽  
Collagen Protein ◽  
Discrete Character ◽  
Lipase A ◽  
Venom Allergen

The total proteomes of Anisakis simplex s.s., A. pegreffii and their hybrid genotype have been compared by quantitative proteomics (iTRAQ approach), which considers the level of expressed proteins. Comparison was made by means of two independent experiments considering four biological replicates of A. simplex and two each for A. pegreffii and hybrid between both species. A total of 1811 and 1976 proteins have been respectively identified in the experiments using public databases. One hundred ninety-six proteins were found significantly differentially expressed, and their relationships with the nematodes’ biological replicates were estimated by a multidimensional statistical approach. Results of pairwise Log2 ratio comparisons among them were statistically treated and supported in order to convert them into discrete character states. Principal component analysis (PCA) confirms the validity of the method. This comparison selected thirty seven proteins as discriminant taxonomic biomarkers among A. simplex, A. pegreffii and their hybrid genotype; 19 of these biomarkers, encoded by ten loci, are specific allergens of Anisakis (Ani s7, Ani s8, Ani s12, and Ani s14) and other (Ancylostoma secreted) is a common nematodes venom allergen. The rest of the markers comprise four unknown or non-characterized proteins; five different proteins (leucine) related to innate immunity, four proteolytic proteins (metalloendopeptidases), a lipase, a mitochondrial translocase protein, a neurotransmitter, a thyroxine transporter, and a structural collagen protein. The proposed methodology (proteomics and statistical) solidly characterize a set of proteins that are susceptible to take advantage of the new targeted proteomics.

scholarly journals Mechanism of Dynamic Binding of Replication Protein A to ssDNA

2020 ◽  
Author(s):  
Anupam Mondal ◽  
Arnab Bhattacherjee
Keyword(s):  
Dna Repair ◽  
Dynamic Equilibrium ◽  
Diffusion Mechanism ◽  
Protein A ◽  
Principal Component ◽  
Replication Protein A ◽  
Replication Protein ◽  
Coarse Grained ◽  
Dna Processing

AbstractReplication protein A (RPA) serves as hub protein inside eukaryotic cells, where it coordinates crucial DNA metabolic processes and activates the DNA-damage response system. A characteristic feature of its action is to associate with ssDNA intermediates before handing over them to downstream proteins. The length of ssDNA intermediates differs for different pathways. This means RPA must have mechanisms for selective processing of ssDNA intermediates based on their length, the knowledge of which is fundamental to elucidate when and how DNA repair and replication processes are symphonized. By employing extensive molecular simulations, we investigated the mechanism of binding of RPA to ssDNA of different lengths. We show that the binding involves dynamic equilibrium with a stable intermediate, the population of which increases with the length of ssDNA. The vital underlying factors are decoded through collective variable principal component analysis. It suggests a differently orchestrated set of interactions that define the action of RPA based on the sizes of ssDNA intermediates. We further estimated the association kinetics and probed the diffusion mechanism of RPA to ssDNA. RPA diffuses on short ssDNA through progressive ‘bulge’ formation. With long ssDNA, we observed a conformational change in ssDNA coupled with its binding to RPA in a cooperative fashion. Our analysis explains how the ‘short-lived,’ long ssDNA intermediates are processed quickly in vivo. The study thus reveals the molecular basis of several recent experimental observations related to RPA binding to ssDNA and provides novel insights into the RPA functioning in DNA repair and replication.Significance StatementDespite ssDNA be the common intermediate to all pathways involving RPA, how does the latter function differently in the DNA processing events such as DNA repair, replication, and recombination just based on the length of ssDNA intermediates remains unknown. The major hindrance is the difficulty in capturing the transient interactions between the molecules. Even attempts to crystallize RPA complexes with 32nt and 62nt ssDNA have yielded a resolved structure of only 25nt ssDNA wrapped with RPA. Here, we used a state-of-the-art coarse-grained protein-ssDNA model to unravel the detailed mechanism of binding of RPA to ssDNA. Our study illustrates the molecular origin of variations in RPA action during various DNA processing events depending on the length of ssDNA intermediates.

scholarly journals A Systematic Analysis of the Alpine Saxifrage Complex (Saxifragaceae) in the Canadian Arctic Islands Using Morphology and Chloroplast DNA Data

2004 ◽  
Vol 118 (3) ◽  
pp. 326 ◽  
Author(s):  
Caroline Healy ◽  
Lynn J. Gillespie
Keyword(s):  
Morphological Variability ◽  
Canadian Arctic ◽  
Principal Component ◽  
Distinct Species ◽  
Morphological Characters ◽  
Sympatric Species ◽  
Sensu Stricto ◽  
Morphological Data ◽  
Systematic Analysis ◽  
Site Analysis

The Saxifraga nivalis complex displays significant ecological, morphological and cytological variation. Most European studies suggest that the S. nivalis complex comprises two distinct species: Saxifraga nivalis sensu stricto and Saxifraga tenuis. However, the presence of intermediate morphotypes, inconsistencies in chromosomal counts and variability in morphological keys and descriptions have led to different taxonomic interpretations of the complex in North America. This study investigated the systematics of Canadian Arctic Island members of this complex from 157 specimens using 23 morphological characters. Principal component analysis of the morphological data revealed two adjacent clusters, corresponding to the two taxa and consistent with a close morphological similarity and the presence of hybrids. A preliminary restriction site analysis of five non-coding regions of the chloroplast genome, trnH-trnK, trnT-trnF, trnF-trnV, trnV-rbcL and rbcL-ORF106, was conducted using 21 restriction endonucleases. This analysis indicated a length difference between the trnT-trnF region of S. nivalis and that of S. tenuis, but no difference in restriction sites for any of the assayed regions. These results confirm that in the Canadian Arctic, the S. nivalis complex consists of two closely related, largely sympatric species, with notable morphological variability, and possible hybrids.

scholarly journals The Selection of Reliable Reference Genes for RT-qPCR Analysis of Anisakis simplex Sensu Stricto Gene Expression from Different Developmental Stages

2020 ◽  
Vol 65 (4) ◽  
pp. 837-842 ◽  
Author(s):  
Elżbieta Łopieńska-Biernat ◽  
Robert Stryiński ◽  
Łukasz Paukszto ◽  
Jan Paweł Jastrzębski ◽  
Karol Makowczenko
Keyword(s):  
Gene Expression ◽  
Reference Genes ◽  
Parasitic Nematode ◽  
Developmental Stages ◽  
Receptor Gene ◽  
Elongation Factor ◽  
Sensu Stricto ◽  
Nuclear Hormone Receptor ◽  
Anisakis Simplex ◽  
Complex Life Cycle

Abstract Background Anisakis simplex s. s. is a parasitic nematode with a complex life cycle in which humans can become accidental hosts by consuming raw or not fully cooked fish containing L3 larvae. The growing popularity of raw fish dishes has contributed to an increase in the incidence of anisakiasis, which has spurred scientific efforts to develop new methods for diagnosing and treating the disease and also to investigate the gene expression at different developmental stages of this parasite. The identification of reference genes suitable for the normalization of RT-qPCR data has not been studied with respect to A. simplex s. s. Methods In the present study, eight candidate reference genes were analyzed in A. simplex s. s. at two different developmental stages: L3 and L4. The expression stability of these genes was assessed by geNorm and NormFinder softwares. Results In general, our results identified translation elongation factor 1α (ef-1α) and peptidyl-prolyl isomerase 12 (ppi12) as the most stable genes in L3 and L4 developmental stages of A. simplex s. s. Validation of the selected reference genes was performed by profiling the expression of the nuclear hormone receptor gene (nhr 48) in different developmental stages. Conclusions This first analysis selecting suitable reference genes for RT-qPCR in A. simplex s. s. will facilitate future functional analyses and deep mining of genetic resources in this parasitic nematode.

scholarly journals Comparative Transcriptomics Reveals Clues for Differences in Pathogenicity between Hysterothylacium aduncum, Anisakis simplex sensu stricto and Anisakis pegreffii

Genes ◽  
2020 ◽  
Vol 11 (3) ◽  
pp. 321 ◽  
Author(s):  
Serena Cavallero ◽  
Fabrizio Lombardo ◽  
Marco Salvemini ◽  
Antonella Pizzarelli ◽  
Cinzia Cantacessi ◽  
...  
Keyword(s):  
Ribosomal Proteins ◽  
De Novo ◽  
Proteolytic Enzymes ◽  
Sensu Stricto ◽  
Anisakis Simplex ◽  
Anisakis Pegreffii ◽  
Primary Hemostasis ◽  
Differential Gene ◽  
Immunomodulatory Peptides ◽  
Hysterothylacium Aduncum

Ascaridoid nematodes are widespread in marine fishes. Despite their major socioeconomic importance, mechanisms associated to the fish-borne zoonotic disease anisakiasis are still obscure. RNA-Seq and de-novo assembly were herein applied to RNA extracted from larvae and dissected pharynx of Hysterothylacium aduncum (HA), a non-pathogenic nematode. Assembled transcripts in HA were annotated and compared to the transcriptomes of the zoonotic species Anisakis simplex sensu stricto (AS) and Anisakis pegreffii (AP). Approximately 60,000,000 single-end reads were generated for HA, AS and AP. Transcripts in HA encoded for 30,254 putative peptides while AS and AP encoded for 20,574 and 20,840 putative peptides, respectively. Differential gene expression analyses yielded 471, 612 and 526 transcripts up regulated in the pharynx of HA, AS and AP. The transcriptomes of larvae and pharynx of HA were enriched in transcripts encoding collagen, peptidases, ribosomal proteins and in heat-shock motifs. Transcripts encoding proteolytic enzymes, anesthetics, inhibitors of primary hemostasis and virulence factors, anticoagulants and immunomodulatory peptides were up-regulated in AS and AP pharynx. This study represents the first transcriptomic characterization of a marine parasitic nematode commonly recovered in fish and probably of negligible concern for public health.

scholarly journals Immunoreactive Proteins in the Esophageal Gland Cells of Anisakis Simplex Sensu Stricto Detected by MALDI-TOF/TOF Analysis

Genes ◽  
2020 ◽  
Vol 11 (6) ◽  
pp. 683
Author(s):  
Lee Robertson ◽  
Susana C. Arcos ◽  
Sergio Ciordia ◽  
Noelia Carballeda-Sanguiao ◽  
María del Carmen Mena ◽  
...  
Keyword(s):  
Sensu Stricto ◽  
Anisakis Simplex ◽  
2D Electrophoresis ◽  
Western Blots ◽  
Gland Cells ◽  
Maldi Tof ◽  
Nematode Parasites ◽  
Esophageal Gland ◽  
Allergenic Potential ◽  
Phylogenetic Origin

In plant and animal nematode parasites, proteins derived from esophageal gland cells have been shown to be important in the host-nematodes relationship but little is known about the allergenic potential of these proteins in the genus Anisakis. Taking into account the increase of anisakiasis and allergies related to these nematodes, immunoreactive properties of gland cell proteins were investigated. Two hundred ventricles were manually dissected from L3 stage larvae of Aniskakis simplex s.s. to allow direct protein analysis. Denaturing gel electrophoresis followed by monochromatic silver staining which revealed the presence of differential (enriched) proteins when compared to total nematode extracts. Such comparison was performed by means of 1D and 2D electrophoresis. Pooled antisera from Anisakis spp.-allergic patients were used in western blots revealing the presence of 13 immunoreactive bands in the ventricular extracts in 1D, with 82 spots revealed in 2D. The corresponding protein bands and spots were excised from the silver-stained gel and protein assignation was made by MALDI-TOF/TOF. A total of 13 (including proteoforms) were unambiguously identified. The majority of these proteins are known to be secreted by nematodes into the external environment, of which three are described as being major allergens in other organisms with different phylogenetic origin and one is an Anisakis simplex allergen.

scholarly journals Relapsing Fever Spirochetes Borrelia recurrentis and B. duttonii Acquire Complement Regulators C4b-Binding Protein and Factor H

2006 ◽  
Vol 74 (7) ◽  
pp. 4157-4163 ◽  
Author(s):  
T. Meri ◽  
S. J. Cutler ◽  
A. M. Blom ◽  
S. Meri ◽  
T. S. Jokiranta
Keyword(s):  
Binding Protein ◽  
Protein A ◽  
Sensu Stricto ◽  
Factor H ◽  
Complement Pathway ◽  
Relapsing Fever ◽  
Borrelia Recurrentis ◽  
First Report ◽  
Complement Regulators

ABSTRACT Relapsing fever is a rapidly progressive and severe septic disease caused by certain Borrelia spirochetes. The disease is divided into two forms, i.e., epidemic relapsing fever, caused by Borrelia recurrentis and transmitted by lice, and the endemic form, caused by several Borrelia species, such as B. duttonii, and transmitted by soft-bodied ticks. The spirochetes enter the bloodstream by the vector bite and live persistently in plasma even after the development of specific antibodies. This leads to fever relapses and high mortality and clearly indicates that the Borrelia organisms utilize effective immune evasion strategies. In this study, we show that the epidemic relapsing fever pathogen B. recurrentis and an endemic relapsing fever pathogen, B. duttonii, are serum resistant, i.e., resistant to complement in vitro. They acquire the host alternative complement pathway regulator factor H on their surfaces in a similar way to that of the less serum-resistant Lyme disease pathogen, B. burgdorferi sensu stricto. More importantly, the relapsing fever spirochetes specifically bind host C4b-binding protein, a major regulator of the antibody-mediated classical complement pathway. Both complement regulators retained their functional activities when bound to the surfaces of the spirochetes. In conclusion, this is the first report of complement evasion by Borrelia recurrentis and B. duttonii and the first report showing capture of C4b-binding protein by spirochetes.

Proteomic profiling and characterization of differential allergens in the nematodes Anisakis simplex sensu stricto and A. pegreffii

PROTEOMICS ◽  
2014 ◽  
Vol 14 (12) ◽  
pp. 1547-1568 ◽  
Author(s):  
Susana C. Arcos ◽  
Sergio Ciordia ◽  
Lee Roberston ◽  
Inés Zapico ◽  
Yolanda Jiménez-Ruiz ◽  
...  
Keyword(s):  
Sensu Stricto ◽  
Anisakis Simplex ◽  
Proteomic Profiling
Sign in / Sign up

Export Citation Format

Share Document