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The biology and the importance of Photobacterium species

Moi IM, Roslan NN, Leow ATC, Ali MSM, Rahman RNZRA, Rahimpour A, et al.

Appl Microbiol Biotechnol, 2017 Jun;101(11):4371-4385.
PMID: 28497204 DOI: 10.1007/s00253-017-8300-y

Photobacterium species are Gram-negative coccobacilli which are distributed in marine habitats worldwide. Some species are unique because of their capability to produce luminescence. Taxonomically, about 23 species and 2 subspecies are validated to date. Genomes from a few Photobacterium spp. have been sequenced and studied. They are considered a special group of bacteria because some species are capable of producing essential polyunsaturated fatty acids, antibacterial compounds, lipases, esterases and asparaginases. They are also used as biosensors in food and environmental monitoring and detectors of drown victim, as well as an important symbiont.

Matched MeSH terms: Bacterial Proteins/genetics*
Fulltext Arctic-adapted dogs emerged at the Pleistocene-Holocene transition

Sinding MS, Gopalakrishnan S, Ramos-Madrigal J, de Manuel M, Pitulko VV, Kuderna L, et al.

Science, 2020 06 26;368(6498):1495-1499.
PMID: 32587022 DOI: 10.1126/science.aaz8599

Although sled dogs are one of the most specialized groups of dogs, their origin and evolution has received much less attention than many other dog groups. We applied a genomic approach to investigate their spatiotemporal emergence by sequencing the genomes of 10 modern Greenland sled dogs, an ~9500-year-old Siberian dog associated with archaeological evidence for sled technology, and an ~33,000-year-old Siberian wolf. We found noteworthy genetic similarity between the ancient dog and modern sled dogs. We detected gene flow from Pleistocene Siberian wolves, but not modern American wolves, to present-day sled dogs. The results indicate that the major ancestry of modern sled dogs traces back to Siberia, where sled dog-specific haplotypes of genes that potentially relate to Arctic adaptation were established by 9500 years ago.

Matched MeSH terms: Mitochondrial Membrane Transport Proteins/genetics
Fulltext Whole genome sequencing of amplified Plasmodium knowlesi DNA from unprocessed blood reveals genetic exchange events between Malaysian Peninsular and Borneo subpopulations

Benavente ED, Gomes AR, De Silva JR, Grigg M, Walker H, Barber BE, et al.

Sci Rep, 2019 07 08;9(1):9873.
PMID: 31285495 DOI: 10.1038/s41598-019-46398-z

The zoonotic Plasmodium knowlesi parasite is the most common cause of human malaria in Malaysia. Genetic analysis has shown that the parasites are divided into three subpopulations according to their geographic origin (Peninsular or Borneo) and, in Borneo, their macaque host (Macaca fascicularis or M. nemestrina). Whilst evidence suggests that genetic exchange events have occurred between the two Borneo subpopulations, the picture is unclear in less studied Peninsular strains. One difficulty is that P. knowlesi infected individuals tend to present with low parasitaemia leading to samples with insufficient DNA for whole genome sequencing. Here, using a parasite selective whole genome amplification approach on unprocessed blood samples, we were able to analyse recent genomes sourced from both Peninsular Malaysia and Borneo. The analysis provides evidence that recombination events are present in the Peninsular Malaysia parasite subpopulation, which have acquired fragments of the M. nemestrina associated subpopulation genotype, including the DBPβ and NBPXa erythrocyte invasion genes. The NBPXb invasion gene has also been exchanged within the macaque host-associated subpopulations of Malaysian Borneo. Our work provides strong evidence that exchange events are far more ubiquitous than expected and should be taken into consideration when studying the highly complex P. knowlesi population structure.

Matched MeSH terms: Protozoan Proteins/genetics
Fulltext Clinical and genetic differences between pustular psoriasis subtypes

Twelves S, Mostafa A, Dand N, Burri E, Farkas K, Wilson R, et al.

J Allergy Clin Immunol, 2019 03;143(3):1021-1026.
PMID: 30036598 DOI: 10.1016/j.jaci.2018.06.038

BACKGROUND: The term pustular psoriasis indicates a group of severe skin disorders characterized by eruptions of neutrophil-filled pustules. The disease, which often manifests with concurrent psoriasis vulgaris, can have an acute systemic (generalized pustular psoriasis [GPP]) or chronic localized (palmoplantar pustulosis [PPP] and acrodermatitis continua of Hallopeau [ACH]) presentation. Although mutations have been uncovered in IL36RN and AP1S3, the rarity of the disease has hindered the study of genotype-phenotype correlations.
OBJECTIVE: We sought to characterize the clinical and genetic features of pustular psoriasis through the analysis of an extended patient cohort.
METHODS: We ascertained a data set of unprecedented size, including 863 unrelated patients (251 with GPP, 560 with PPP, 28 with ACH, and 24 with multiple diagnoses). We undertook mutation screening in 473 cases.
RESULTS: Psoriasis vulgaris concurrence was lowest in PPP (15.8% vs 54.4% in GPP and 46.2% in ACH, P

Matched MeSH terms: Vesicular Transport Proteins/genetics
Fulltext Phylogeny and putative virulence gene analysis of Bartonella bovis

Tay ST, Kho KL, Lye SF, Ngeow YF

J Vet Med Sci, 2018 Apr 18;80(4):653-661.
PMID: 29311425 DOI: 10.1292/jvms.17-0448

Bartonella bovis is a small Gram-negative bacterium recognized as an etiological agent for bacteremia and endocarditis in cattle. As few reports are available on the taxonomic position of B. bovis and its mechanism of virulence, this study aims to resolve the phylogeny of B. bovis and investigate putative virulence genes based on whole genome sequence analysis. Genome-wide comparisons based on single nucleotide polymorphisms (SNP) and orthologous genes were performed in this study for phylogenetic inference of 27 Bartonella species. Rapid Annotation using Subsystem Technology (RAST) analysis was used for annotation of putative virulence genes. The phylogenetic tree generated from the genome-wide comparison of orthologous genes exhibited a topology almost similar to that of the tree generated from SNP-based comparison, indicating a high concordance in the nucleotide and amino acid sequences of Bartonella spp. The analyses show consistent grouping of B. bovis in a cluster related to ruminant-associated species, including Bartonella australis, Bartonella melophagi and Bartonella schoenbuchensis. RAST analysis revealed genes encoding flagellar components, in corroboration with the observation of flagella-like structure of BbUM strain under negative straining. Genes associated with virulence, disease and defence, prophages, membrane transport, iron acquisition, motility and chemotaxis are annotated in B. bovis genome. The flagellin (flaA) gene of B. bovis is closely related to Bartonella bacilliformis and Bartonella clarridgeiae but distinct from other Gram-negative bacteria. The absence of type IV secretion systems, the bona fide pathogenicity factors of bartonellae, in B. bovis suggests that it may have a different mechanism of pathogenicity.

Matched MeSH terms: Bacterial Proteins/genetics
A Rhythmic Gene Entrained to Midnight May Regulate Photoperiod-Controlled Flowering in Arabidopsis

Yeang HY

Yale J Biol Med, 2019 06;92(2):213-223.
PMID: 31249482

The widely held explanation for photoperiod-controlled flowering in long-day plants is largely embodied in the External Coincidence Hypothesis which posits that flowering is induced when activity of a rhythmic gene that regulates it (a putative "flowering gene") occurs in the presence of light. Nevertheless, re-examination of the Arabidopsis flowering data from non 24-hour cycles of Roden et al. suggests that External Coincidence is not tenable if the circadian rhythm of the "flowering gene" were entrained to sunrise as commonly accepted. On the other hand, the hypothesis is supported if circadian cycling of the gene conforms to a solar rhythm, and its entrainment is to midnight on the solar clock. Data available point to flowering being induced by the gene which peaks in its expression between 16 to 19 h after midnight. In the normal 24 h cycle, that would be between 4 p.m. and 7 p.m., regardless of the photoperiod. Such timing of the "flowering gene" expression allows for variable coincidence between gene activity and light, depending on the photoperiod and cycle period. A correlation is found between earliness of flowering and the degree of coincidence of "flowering gene" expression with light (r = 0.88, p<0.01).

Matched MeSH terms: Arabidopsis Proteins/genetics
Comparative genomics reveals differences in algal galactan biosynthesis and related pathways in early and late diverging red algae

Ho CL

Genomics, 2020 03;112(2):1536-1544.
PMID: 31494197 DOI: 10.1016/j.ygeno.2019.09.002

Red algae are a major source of marine sulfated galactans. In this study, orthologs and inparalogs from seven red algae were analyzed and compared with the aim to discover differences in algal galactan biosynthesis and related pathways of these algae. Red algal orthologs for putative carbohydrate sulfotransferases were found to be prevalent in Porphyridium purpureum, Florideophytes and Bangiophytes, while red algal orthologs for putative chondroitin sulfate synthases, sulfurylases, and porphyranases /carrageenases were found exclusively in Florideophytes and Bangiophytes. The acquirement of these genes could have happened after the divergence from Cyanidiales red algae. Cyanidiales red algae were found to have more number and types of putative sulfate permeases, suggesting that these genes could have been acquired in adaptation to the environmental stresses and biogeochemistry of respective habitats. The findings of this study shed lights on the evolution of different homeostasis mechanisms by the early and late diverging red algal orders.

Matched MeSH terms: Plant Proteins/genetics
Fulltext Low Levels of Polymorphisms and Negative Selection in Plasmodum knowlesi Merozoite Surface Protein 8 in Malaysian Isolates

Atique Ahmed M, Kang HJ, Quan FS

Korean J Parasitol, 2019 Aug;57(4):445-450.
PMID: 31533414 DOI: 10.3347/kjp.2019.57.4.445

Human infections due to the monkey malaria parasite Plasmodium knowlesi is increasingly being reported from most Southeast Asian countries specifically Malaysia. The parasite causes severe and fatal malaria thus there is a need for urgent measures for its control. In this study, the level of polymorphisms, haplotypes and natural selection of full-length pkmsp8 in 37 clinical samples from Malaysian Borneo along with 6 lab-adapted strains were investigated. Low levels of polymorphism were observed across the full-length gene, the double epidermal growth factor (EGF) domains were mostly conserved, and non-synonymous substitutions were absent. Evidence of strong negative selection pressure in the non-EGF regions were found indicating functional constrains acting at different domains. Phylogenetic haplotype network analysis identified shared haplotypes and indicated geographical clustering of samples originating from Peninsular Malaysia and Malaysian Borneo. This is the first study to genetically characterize the full-length msp8 gene from clinical isolates of P. knowlesi from Malaysia; however, further functional characterization would be useful for future rational vaccine design.

Matched MeSH terms: Protozoan Proteins/genetics*
Iron-associated protein interaction networks reveal the key functional modules related to survival and virulence of Pasteurella multocida

Jatuponwiphat T, Chumnanpuen P, Othman S, E-Kobon T, Vongsangnak W

Microb Pathog, 2019 Feb;127:257-266.
PMID: 30550841 DOI: 10.1016/j.micpath.2018.12.013

Pasteurella multocida causes respiratory infectious diseases in a multitude of birds and mammals. A number of virulence-associated genes were reported across different strains of P. multocida, including those involved in the iron transport and metabolism. Comparative iron-associated genes of P. multocida among different animal hosts towards their interaction networks have not been fully revealed. Therefore, this study aimed to identify the iron-associated genes from core- and pan-genomes of fourteen P. multocida strains and to construct iron-associated protein interaction networks using genome-scale network analysis which might be associated with the virulence. Results showed that these fourteen strains had 1587 genes in the core-genome and 3400 genes constituting their pan-genome. Out of these, 2651 genes associated with iron transport and metabolism were selected to construct the protein interaction networks and 361 genes were incorporated into the iron-associated protein interaction network (iPIN) consisting of nine different iron-associated functional modules. After comparing with the virulence factor database (VFDB), 21 virulence-associated proteins were determined and 11 of these belonged to the heme biosynthesis module. From this study, the core heme biosynthesis module and the core outer membrane hemoglobin receptor HgbA were proposed as candidate targets to design novel antibiotics and vaccines for preventing pasteurellosis across the serotypes or animal hosts for enhanced precision agriculture to ensure sustainability in food security.

Matched MeSH terms: Bacterial Proteins/genetics*
Optimisation of human VH domain antibodies specific to Mycobacterium tuberculosis heat shock protein (HSP16.3)

Soong JX, Chan SK, Lim TS, Choong YS

J Comput Aided Mol Des, 2019 03;33(3):375-385.
PMID: 30689080 DOI: 10.1007/s10822-019-00186-z

Mycobacterium tuberculosis (Mtb) 16.3 kDa heat shock protein 16.3 (HSP16.3) is a latency-associated antigen that can be targeted for latent tuberculosis (TB) diagnostic and therapeutic development. We have previously developed human VH domain antibodies (dAbs; clone E3 and F1) specific against HSP16.3. In this work, we applied computational methods to optimise and design the antibodies in order to improve the binding affinity with HSP16.3. The VH domain antibodies were first docked to the dimer form of HSP16.3 and further sampled using molecular dynamics simulation. The calculated binding free energy of the HSP16.3-dAb complexes showed non-polar interactions were responsible for the antigen-antibody association. Per-residue free energy decomposition and computational alanine scanning have identified one hotspot residue for E3 (Y391) and 4 hotspot residues for F1 (M394, Y396, R397 and M398). These hotspot residues were then mutated and evaluated by binding free energy calculations. Phage ELISA assay was carried out on the potential mutants (E3Y391W, F1M394E, F1R397N and F1M398Y). The experimental assay showed improved binding affinities of E3Y391W and F1M394E against HSP16.3 compared with the wild type E3 and F1. This case study has thus showed in silico methods are able to assist in optimisation or improvement of antibody-antigen binding.

Matched MeSH terms: Bacterial Proteins/genetics
Biomarkers for ischemic stroke subtypes: A protein-protein interaction analysis

Wei LK, Quan LS

Comput Biol Chem, 2019 Dec;83:107116.
PMID: 31561071 DOI: 10.1016/j.compbiolchem.2019.107116

According to the Trial of Org 10172 in Acute Stroke Treatment, ischemic stroke is classified into five subtypes. However, the predictive biomarkers of ischemic stroke subtypes are still largely unknown. The utmost objective of this study is to map, construct and analyze protein-protein interaction (PPI) networks for all subtypes of ischemic stroke, and to suggest the predominant biological pathways for each subtypes. Through 6285 protein data retrieved from PolySearch2 and STRING database, the first PPI networks for all subtypes of ischemic stroke were constructed. Notably, F2 and PLG were identified as the critical proteins for large artery atherosclerosis (LAA), lacunar, cardioembolic, stroke of other determined etiology (SOE) and stroke of undetermined etiology (SUE). Gene ontology and DAVID analysis revealed that GO:0030193 regulation of blood coagulation and GO:0051917 regulation of fibrinolysis were the important functional clusters for all the subtypes. In addition, inflammatory pathway was the key etiology for LAA and lacunar, while FOS and JAK2/STAT3 signaling pathways might contribute to cardioembolic stroke. Due to many risk factors associated with SOE and SUE, the precise etiology for these two subtypes remained to be concluded.

Matched MeSH terms: Proteins/genetics
Fulltext Application of Reverse Genetics in Functional Genomics of Potyvirus

Kannan M, Zainal Z, Ismail I, Baharum SN, Bunawan H

Viruses, 2020 07 26;12(8).
PMID: 32722532 DOI: 10.3390/v12080803

Numerous potyvirus studies, including virus biology, transmission, viral protein function, as well as virus-host interaction, have greatly benefited from the utilization of reverse genetic techniques. Reverse genetics of RNA viruses refers to the manipulation of viral genomes, transfection of the modified cDNAs into cells, and the production of live infectious progenies, either wild-type or mutated. Reverse genetic technology provides an opportunity of developing potyviruses into vectors for improving agronomic traits in plants, as a reporter system for tracking virus infection in hosts or a production system for target proteins. Therefore, this review provides an overview on the breakthroughs achieved in potyvirus research through the implementation of reverse genetic systems.

Matched MeSH terms: Viral Proteins/genetics
Transactivator protein: An alternative for delivery of recombinant proteins for safer reprogramming of induced Pluripotent Stem Cell

Nordin F, Ahmad RNR, Farzaneh F

Virus Res, 2017 05 02;235:106-114.
PMID: 28408207 DOI: 10.1016/j.virusres.2017.04.007

Induced pluripotent stem cells (iPSC) are somatic cells reprogrammed to pluripotency by forced expression of pluripotency factors. These cells are shown to have the same pluripotent potential as embryonic stem cells (ESC) and considered as an alternative to the much controversial usage of ESC which involved human embryos. However, the traditional method in reprogramming cells into iPSC using genome-integrating retro- or lenti- viruses remains an obstacle for its application in clinical setting. Although numerous studies have been conducted for a safer DNA-based reprogramming, reprogramming of iPSC by genetic modifications may raise the possibility of malignant transformation and has been a major limitation for its usage in clinical applications. Therefore, there is a need for an alternative method to reprogram the cells without the use of gene editing and a much safer way to deliver transcription factors to induce pluripotency on target cells. Using protein transduction approach, a number of studies have demonstrated the generation of human iPSCs from human fibroblasts and mouse embryonic fibroblasts by direct delivery of reprogramming proteins. In this review, the definition and mechanism of HIV-TAT protein (a type of protein transduction domain) in delivering recombinant proteins, including the potential of protein-based delivery to induce iPSC were further discussed.

Matched MeSH terms: Recombinant Proteins/genetics
Complete nucleotide sequence of RNA segment 3 of bluetongue virus serotype 2 (Ona-A). Phylogenetic analyses reveal the probable origin and relationship with other orbiviruses

Pritchard LI, Gould AR, Wilson WC, Thompson L, Mertens PP, Wade-Evans AM

Virus Res, 1995 Mar;35(3):247-61.
PMID: 7785314

The nucleotide sequence of the RNA segment 3 of bluetongue virus (BTV) serotype 2 (Ona-A) from North America was determined to be 2772 nucleotides containing a single large open reading frame of 2703 nucleotides (901 amino acid). The predicted VP3 protein exhibited general physiochemical properties (including hydropathy profiles) which were very similar to those previously deduced for other BTV VP3 proteins. Partial genome segment 3 sequences, obtained by polymerase chain reaction (PCR) sequencing, of BTV isolates from the Caribbean were compared to those from North America, South Africa, India, Indonesia, Malaysia and Australia, as well as other orbiviruses, to determine the phylogenetic relationships amongst them. Three major BTV topotypes (Gould, A.R. (1987) Virus Res. 7, 169-183) were observed which had nucleotide sequences that differed by approximately 20%. At the molecular level, geographic separation had resulted in significant divergence in the BTV genome segment 3 sequences, consistent with the evolution of distinct viral populations. The close phylogenetic relationship between the BTV serotype 2 (Ona-A strain) from Florida and the BTV serotypes 1, 6 and 12 from Jamaica and Honduras, indicated that the presence of BTV serotype 2 in North America was probably due to an exotic incursion from the Caribbean region as previously proposed by Sellers and Maaroof ((1989) Can. J. Vet. Res. 53, 100-102) based on trajectory analysis. Conversely, nucleotide sequence analysis of Caribbean BTV serotype 17 isolates suggested they arose from incursions which originated in the USA, possibly from a BTV population distinct from those circulating in Wyoming.

Matched MeSH terms: Viral Core Proteins/genetics*
Molecular cloning and expression of five glutathione S-transferase (GST) genes from Banana (Musa acuminata L. AAA group, cv. Cavendish)

Wang Z, Huang S, Jia C, Liu J, Zhang J, Xu B, et al.

Plant Cell Rep, 2013 Sep;32(9):1373-80.
PMID: 23652818 DOI: 10.1007/s00299-013-1449-7

KEY MESSAGE: Three tau class MaGSTs responded to abiotic stress, MaGSTF1 and MaGSTL1 responded to signaling molecules, they may play an important role in the growth of banana plantlet. Glutathione S-transferases (GST) are multifunctional detoxification enzymes that participate in a variety of cellular processes, including stress responses. In this study, we report the molecular characteristics of five GST genes (MaGSTU1, MaGSTU2, MaGSTU3, MaGSTF1 and MaGSTL1) cloned from banana (Musa acuminate L. AAA group, cv. Cavendish) using a RACE-PCR-based strategy. The predicted molecular masses of these GSTs range from 23.4 to 27.7 kDa and their pIs are acidic. At the amino acid level, they share high sequence similarity with GSTs in the banana DH-Pahang (AA group) genome. Phylogenetic analysis showed that the deduced amino acid sequences of MaGSTs also have high similarity to GSTs of other plant species. Expression analysis by semi-quantitative RT-PCR revealed that these genes are differentially expressed in various tissues. In addition, their expression is regulated by various stress conditions, including exposure to signaling molecules, cold, salinity, drought and Fusarium oxysporum f specialis(f. Sp) cubense Tropical Race 4 (Foc TR4) infection. The expression of the tau class MaGSTs (MaGSTU1, MaGSTU2 and MaGSTU3) mainly responded to cold, salinity and drought while MaGSTF1 and MaGSTL1 expressions were upregulated by signaling molecules. Our findings suggest that MaGSTs play a key role in both development and abiotic stress responses.

Matched MeSH terms: Plant Proteins/genetics
DNA methylation changes in clonally propagated oil palm

Sarpan N, Taranenko E, Ooi SE, Low EL, Espinoza A, Tatarinova TV, et al.

Plant Cell Rep, 2020 Sep;39(9):1219-1233.
PMID: 32591850 DOI: 10.1007/s00299-020-02561-9

KEY MESSAGE: Several hypomethylated sites within the Karma region of EgDEF1 and hotspot regions in chromosomes 1, 2, 3, and 5 may be associated with mantling. One of the main challenges faced by the oil palm industry is fruit abnormalities, such as the "mantled" phenotype that can lead to reduced yields. This clonal abnormality is an epigenetic phenomenon and has been linked to the hypomethylation of a transposable element within the EgDEF1 gene. To understand the epigenome changes in clones, methylomes of clonal oil palms were compared to methylomes of seedling-derived oil palms. Whole-genome bisulfite sequencing data from seedlings, normal, and mantled clones were analyzed to determine and compare the context-specific DNA methylomes. In seedlings, coding and regulatory regions are generally hypomethylated while introns and repeats are extensively methylated. Genes with a low number of guanines and cytosines in the third position of codons (GC3-poor genes) were increasingly methylated towards their 3' region, while GC3-rich genes remain demethylated, similar to patterns in other eukaryotic species. Predicted promoter regions were generally hypomethylated in seedlings. In clones, CG, CHG, and CHH methylation levels generally decreased in functionally important regions, such as promoters, 5' UTRs, and coding regions. Although random regions were found to be hypomethylated in clonal genomes, hypomethylation of certain hotspot regions may be associated with the clonal mantling phenotype. Our findings, therefore, suggest other hypomethylated CHG sites within the Karma of EgDEF1 and hypomethylated hotspot regions in chromosomes 1, 2, 3 and 5, are associated with mantling.

Matched MeSH terms: Plant Proteins/genetics
Fulltext An integrated linkage map of interspecific backcross 2 (BC2) populations reveals QTLs associated with fatty acid composition and vegetative parameters influencing compactness in oil palm

Yaakub Z, Kamaruddin K, Singh R, Mustafa S, Marjuni M, Ting NC, et al.

BMC Plant Biol, 2020 Jul 29;20(1):356.
PMID: 32727448 DOI: 10.1186/s12870-020-02563-5

BACKGROUND: Molecular breeding has opened new avenues for crop improvement with the potential for faster progress. As oil palm is the major producer of vegetable oil in the world, its improvement, such as developing compact planting materials and altering its oils' fatty acid composition for wider application, is important.
RESULTS: This study sought to identify the QTLs associated with fatty acid composition and vegetative traits for compactness in the crop. It integrated two interspecific backcross two (BC2) mapping populations to improve the genetic resolution and evaluate the consistency of the QTLs identified. A total 1963 markers (1814 SNPs and 149 SSRs) spanning a total map length of 1793 cM were integrated into a consensus map. For the first time, some QTLs associated with vegetative parameters and carotene content were identified in interspecific hybrids, apart from those associated with fatty acid composition. The analysis identified 8, 3 and 8 genomic loci significantly associated with fatty acids, carotene content and compactness, respectively.
CONCLUSIONS: Major genomic region influencing the traits for compactness and fatty acid composition was identified in the same chromosomal region in the two populations using two methods for QTL detection. Several significant loci influencing compactness, carotene content and FAC were common to both populations, while others were specific to particular genetic backgrounds. It is hoped that the QTLs identified will be useful tools for marker-assisted selection and accelerate the identification of desirable genotypes for breeding.

Matched MeSH terms: Plant Proteins/genetics
Phylogenetics of the Old World screwworm fly and its significance for planning control and monitoring invasions in Asia

Wardhana AH, Hall MJ, Mahamdallie SS, Muharsini S, Cameron MM, Ready PD

Int J Parasitol, 2012 Jul;42(8):729-38.
PMID: 22664061 DOI: 10.1016/j.ijpara.2012.04.017

Phylogenetic, genealogical and population relationships of Chrysomya bezziana, the Old World screwworm fly (OWSF), were inferred from DNA sequences of mitochondrial cytochrome b (cyt b), nuclear elongation factor-1α (EF-1α) and nuclear white eye colour (white), using sequences of Chrysomya megacephala and Chrysomya rufifacies as outgroups. Cyt b (717bp, 754 specimens), EF-1α (361bp, 256 specimens) and white (577bp, 242 specimens) were analysed from up to two African and nine Asian countries, including 10 Indonesian islands. We show that OWSF occurs as distinctive African and Asian lineages based on cyt b and white, and that there is a marked differentiation between Sumatran and Javan populations in Indonesia, supported by the genealogy and analysis of molecular variance of cyt b alone. Four cyt b sub-lineages are recognised in Asia: only 2.1 occurs on the Asian mainland, from Yemen to Peninsular Malaysia; only 2.2, 2.3 and 2.4 occur in central Indonesia; 2.4 predominates on New Guinea; and 2.1 co-occurs with others only on Sumatra in western Indonesia. This phylogeography and the genetic distances between cyt b haplotypes indicate pre-historic, natural dispersal of OWSF eastwards into Indonesia and other Malesian islands, followed by vicariant evolution in New Guinea and central Indonesia. OWSF is absent from Australia, where there is surveillance for importation or natural invasion. Judged by cyt b haplotype markers, there is currently little spread of OWSF across sea barriers, despite frequent shipments of Australian livestock through Indonesian seas to the Middle East Gulf region. These findings will inform plans for integrated pest management, which could be applied progressively, for example starting in East Nusa Tenggara (central Indonesia) where OWSF has regional cyt b markers, and progressing westwards to Java where any invasion from Sumatra is unlikely. Cyt b markers would help identify the source of any re-emergence in treated areas.

Matched MeSH terms: Insect Proteins/genetics
Fulltext Limited expression of non-integrating CpG-free plasmid is associated with increased nucleosome enrichment

Habib O, Mohd Sakri R, Ghazalli N, Chau DM, Ling KH, Abdullah S

PLoS One, 2020;15(12):e0244386.
PMID: 33347482 DOI: 10.1371/journal.pone.0244386

CpG-free pDNA was reported to facilitate sustained transgene expression with minimal inflammation in vivo as compared to CpG-containing pDNA. However, the expression potential and impact of CpG-free pDNA in in vitro model have never been described. Hence, in this study, we analyzed the transgene expression profiles of CpG-free pDNA in vitro to determine the influence of CpG depletion from the transgene. We found that in contrast to the published in vivo studies, CpG-free pDNA expressed a significantly lower level of luciferase than CpG-rich pDNA in several human cell lines. By comparing novel CpG-free pDNA carrying CpG-free GFP (pZGFP: 0 CpG) to CpG-rich GFP (pRGFP: 60 CpGs), we further showed that the discrepancy was not influenced by external factors such as gene transfer agent, cell species, cell type, and cytotoxicity. Moreover, pZGFP exhibited reduced expression despite having equal gene dosage as pRGFP. Analysis of mRNA distribution revealed that the mRNA export of pZGFP and pRGFP was similar; however, the steady state mRNA level of pZGFP was significantly lower. Upon further investigation, we found that the CpG-free transgene in non-integrating CpG-free pDNA backbone acquired increased nucleosome enrichment as compared with CpG-rich transgene, which may explain the observed reduced level of steady state mRNA. Our findings suggest that nucleosome enrichment could regulate non-integrating CpG-free pDNA expression and has implications on pDNA design.

Matched MeSH terms: Green Fluorescent Proteins/genetics
Fulltext The Role of Surface Exposed Lysine in Conformational Stability and Functional Properties of Lipase from Staphylococcus Family

Ahmad NN, Ahmad Kamarudin NH, Leow ATC, Rahman RNZRA

Molecules, 2020 Aug 25;25(17).
PMID: 32854267 DOI: 10.3390/molecules25173858

Surface charge residues have been recognized as one of the stability determinants in protein. In this study, we sought to compare and analyse the stability and conformational dynamics of staphylococcal lipase mutants with surface lysine mutation using computational and experimental methods. Three highly mutable and exposed lysine residues (Lys91, Lys177, Lys325) were targeted to generate six mutant lipases in silico. The model structures were simulated in water environment at 25 °C. Our simulations showed that the stability was compromised when Lys177 was substituted while mutation at position 91 and 325 improved the stability. To illustrate the putative alterations of enzyme stability in the stabilising mutants, we characterized single mutant K325G and double mutant K91A/K325G. Both mutants showed a 5 °C change in optimal temperature compared to their wild type. Single mutant K325G rendered a longer half-life at 25 °C (T1/2 = 21 h) while double mutant K91A/K325G retained only 40% of relative activity after 12 h incubation. The optimal pH for mutant K325G was shifted from 8 to 9 and similar substrate preference was observed for the wild type and two mutants. Our findings indicate that surface lysine mutation alters the enzymatic behaviour and, thus, rationalizes the functional effects of surface exposed lysine in conformational stability and activity of this lipase.

Matched MeSH terms: Bacterial Proteins/genetics

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