Phase Variation of Campylobacter jejuni 81-176 Lipooligosaccharide Affects Ganglioside Mimicry and Invasiveness In Vitro

ABSTRACT The outer cores of the lipooligosaccharides (LOS) of many strains of Campylobacter jejuni mimic human gangliosides in structure. A population of cells of C. jejuni strain 81-176 produced a mixture of LOS cores which consisted primarily of structures mimicking GM2 and GM3 gangliosides, with minor amounts of structures mimicking GD1b and GD2. Genetic analyses of genes involved in the biosynthesis of the outer core of C. jejuni 81-176 revealed the presence of a homopolymeric tract of G residues within a gene encoding CgtA, an N-acetylgalactosaminyltransferase. Variation in the number of G residues within cgtA affected the length of the open reading frame, and these changes in cgtA corresponded to a change in LOS structure from GM2 to GM3 ganglioside mimicry. Site-specific mutation of cgtA in 81-176 resulted in a major LOS core structure that lacked GalNAc and resembled GM3 ganglioside. Compared to wild-type 81-176, the cgtA mutant showed a significant increase in invasion of INT407 cells. In comparison, a site-specific mutation of the neuC1 gene resulted in the loss of sialic acid in the LOS core and reduced resistance to normal human serum but had no affect on invasion of INT407 cells.

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The potential of a site-specific delivery of thiamine hydrochloride as a novel insect repellent exerting long-term protection on human skin: In-vitro, ex-vivo study and clinical assessment

Journal of Pharmaceutical Sciences ◽

10.1016/j.xphs.2021.07.017 ◽

2021 ◽

Author(s):

Mai El Halawany ◽

Randa Latif ◽

Alia Badawi

Keyword(s):

Human Skin ◽

Clinical Assessment ◽

Ex Vivo ◽

Thiamine Hydrochloride ◽

Insect Repellent ◽

Site Specific ◽

A Site ◽

Ex Vivo Study

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Slipped strand mispairing in the gene encoding sialidase NanH3 in Gardnerella spp

10.1101/2020.09.16.300947 ◽

2020 ◽

Author(s):

Shakya P. Kurukulasuriya ◽

Mo H. Patterson ◽

Janet E. Hill

Keyword(s):

Cell Wall ◽

Biofilm Formation ◽

Gene Sequence ◽

Phase Variation ◽

Cell Wall Proteins ◽

Vaginal Microbiome ◽

Gene Encoding ◽

Reading Frame ◽

Sialidase Activity ◽

Mucosal Surfaces

AbstractCell wall proteins with sialidase activity are involved in carbohydrate assimilation, adhesion to mucosal surfaces, and biofilm formation. Gardnerella spp. inhabit the human vaginal microbiome and encode up to three sialidase enzymes, two of which are suspected to be cell wall associated. Here we demonstrate that the gene encoding extracellular sialidase NanH3 is found almost exclusively in G. piotii and closely related Gardnerella genome sp. 3, and its presence correlates with sialidase positive phenotype in a collection of 112 Gardnerella isolates. The nanH3 gene sequence includes a homopolymeric repeat of cytosines that varies in length within cell populations, indicating that this gene is subject to slipped-strand mispairing, a mechanisms of phase variation in bacteria. Variation in the length of the homopolymer sequence results in encoding of either the full length sialidase protein or truncated peptides lacking the sialidase domain due to introduction of reading-frame shifts and premature stop codons. Phase variation in NanH3 may be involved in immune evasion or modulation of adhesion to host epithelial cells, and formation of biofilms characteristic of the vaginal dysbiosis known as bacterial vaginosis.

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The Hrq1 helicase stimulates Pso2 translesion nuclease activity to promote DNA inter-strand crosslink repair

10.1101/773267 ◽

2019 ◽

Cited By ~ 2

Author(s):

Cody M. Rogers ◽

Chun-Ying Lee ◽

Samuel Parkins ◽

Nicholas J. Buehler ◽

Sabine Wenzel ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Dna Damage Response ◽

Nuclease Activity ◽

Site Specific ◽

Damage Response ◽

Physical Barriers ◽

A Site ◽

Translesion Polymerases ◽

Stimulation Of

AbstractDNA inter-strand crosslink (ICL) repair requires a complicated network of DNA damage response pathways. Removal of these lesions is vital as they are physical barriers to essential DNA processes that require the separation of duplex DNA, such as replication and transcription. The Fanconi anemia (FA) pathway is the principle mechanism for ICL repair in metazoans and is coupled to replication. In Saccharomyces cerevisiae, a degenerate FA pathway is present, but ICLs are predominantly repaired by a pathway involving the Pso2 nuclease that is hypothesized to digest through the lesion to provide access for translesion polymerases. However, Pso2 lacks translesion nuclease activity in vitro, and mechanistic details of this pathway are lacking, especially relative to FA. We recently identified the Hrq1 helicase, a homolog of the disease-linked RECQL4, as a novel component of Pso2- mediated ICL repair. Here, we show that Hrq1 stimulates the Pso2 nuclease in a mechanism that requires Hrq1 catalytic activity. Importantly, Hrq1 also stimulates Pso2 translesion nuclease activity through a site- specific ICL in vitro. Stimulation of Pso2 nuclease activity is specific to eukaryotic RecQ4 subfamily helicases, and Hrq1 likely interacts with Pso2 through their N-terminal domains. These results advance our understanding of FA-independent ICL repair and establish a role for the RecQ4 helicases in the repair of these dangerous lesions.

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CapA, an Autotransporter Protein of Campylobacter jejuni, Mediates Association with Human Epithelial Cells and Colonization of the Chicken Gut

Journal of Bacteriology ◽

10.1128/jb.01427-06 ◽

2006 ◽

Vol 189 (5) ◽

pp. 1856-1865 ◽

Cited By ~ 82

Author(s):

Sami S. A. Ashgar ◽

Neil J. Oldfield ◽

Karl G. Wooldridge ◽

Michael A. Jones ◽

Greg J. Irving ◽

...

Keyword(s):

Campylobacter Jejuni ◽

Protein A ◽

Phase Variation ◽

Subcellular Fractionation ◽

Immunogold Electron Microscopy ◽

Insertion Mutant ◽

Genes Encoding ◽

Monospecific Antisera ◽

Recombinant Polypeptides

ABSTRACT Two putative autotransporter proteins, CapA and CapB, were identified in silico from the genome sequence of Campylobacter jejuni NCTC11168. The genes encoding each protein contain homopolymeric tracts, suggestive of phase variation mediated by a slipped-strand mispairing mechanism; in each case the gene sequence contained frameshifts at these positions. The C-terminal two-thirds of the two genes, as well as a portion of the predicted signal peptides, were identical; the remaining N-terminal portions were gene specific. Both genes were cloned and expressed; recombinant polypeptides were purified and used to raise rabbit polyclonal monospecific antisera. Using immunoblotting, expression of the ca.116-kDa CapA protein was demonstrated for in vitro-grown cells of strain NCTC11168, for 4 out of 11 recent human fecal isolates, and for 2 out of 8 sequence-typed strains examined. Expression of CapB was not detected for any of the strains tested. Surface localization of CapA was demonstrated by subcellular fractionation and immunogold electron microscopy. Export of CapA was inhibited by globomycin, reinforcing the bioinformatic prediction that the protein is a lipoprotein. A capA insertion mutant had a significantly reduced capacity for association with and invasion of Caco-2 cells and failed to colonize and persist in chickens, indicating that CapA plays a role in host association and colonization by Campylobacter. In view of this demonstrated role, we propose that CapA stands for Campylobacter adhesion protein A.

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Involvement of a site-specific trans-acting factor and a common RNA-binding protein in the editing of chloroplast mRNAs: development of a chloroplast in vitro RNA editing system

The EMBO Journal ◽

10.1093/emboj/20.5.1144 ◽

2001 ◽

Vol 20 (5) ◽

pp. 1144-1152 ◽

Cited By ~ 109

Author(s):

T. Hirose

Keyword(s):

Rna Editing ◽

Rna Binding ◽

Binding Protein ◽

Rna Binding Protein ◽

Site Specific ◽

A Site

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Thermo-Sensitive Vesicles in Controlled Drug Delivery for Chemotherapy

Pharmaceutics ◽

10.3390/pharmaceutics10030150 ◽

2018 ◽

Vol 10 (3) ◽

pp. 150 ◽

Cited By ~ 14

Author(s):

Elisabetta Mazzotta ◽

Lorena Tavano ◽

Rita Muzzalupo

Keyword(s):

Drug Delivery ◽

Cancer Treatment ◽

Controlled Drug Delivery ◽

Solid Tumours ◽

Site Specific ◽

Mild Hyperthermia ◽

Promising Tool ◽

A Site

Thermo-sensitive vesicles are a promising tool for triggering the release of drugs to solid tumours when used in combination with mild hyperthermia. Responsivity to temperature makes them intelligent nanodevices able to provide a site-specific chemotherapy. Following a brief introduction concerning hyperthermia and its advantageous combination with vesicular systems, recent investigations on thermo-sensitive vesicles useful for controlled drug delivery in cancer treatment are reported in this review. In particular, the influence of bilayer composition on the in vitro and in vivo behaviour of thermo-sensitive formulations currently under investigation have been extensively explored.

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Surface Diversity in Mycoplasma agalactiae Is Driven by Site-Specific DNA Inversions within the vpma Multigene Locus

Journal of Bacteriology ◽

10.1128/jb.184.21.5987-5998.2002 ◽

2002 ◽

Vol 184 (21) ◽

pp. 5987-5998 ◽

Cited By ~ 32

Author(s):

Michelle D. Glew ◽

Marc Marenda ◽

Renate Rosengarten ◽

Christine Citti

Keyword(s):

Promoter Sequence ◽

Untranslated Regions ◽

Dna Rearrangements ◽

Site Specific ◽

Reading Frame ◽

Mycoplasma Agalactiae ◽

Variable Surface ◽

A Site ◽

Active Promoter ◽

Minimal Region

ABSTRACT The ruminant pathogen Mycoplasma agalactiae possesses a family of abundantly expressed variable surface lipoproteins called Vpmas. Phenotypic switches between Vpma members have previously been correlated with DNA rearrangements within a locus of vpma genes and are proposed to play an important role in disease pathogenesis. In this study, six vpma genes were characterized in the M. agalactiae type strain PG2. All vpma genes clustered within an 8-kb region and shared highly conserved 5′ untranslated regions, lipoprotein signal sequences, and short N-terminal sequences. Analyses of the vpma loci from consecutive clonal isolates showed that vpma DNA rearrangements were site specific and that cleavage and strand exchange occurred within a minimal region of 21 bp located within the 5′ untranslated region of all vpma genes. This process controlled expression of vpma genes by effectively linking the open reading frame (ORF) of a silent gene to a unique active promoter sequence within the locus. An ORF (xer1) immediately adjacent to one end of the vpma locus did not undergo rearrangement and had significant homology to a distinct subset of genes belonging to the λ integrase family of site-specific xer recombinases. It is proposed that xer1 codes for a site-specific recombinase that is not involved in chromosome dimer resolution but rather is responsible for the observed vpma-specific recombination in M. agalactiae.

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Purification and Characterization of the R64 Shufflon-Specific Recombinase

Journal of Bacteriology ◽

10.1128/jb.182.10.2787-2792.2000 ◽

2000 ◽

Vol 182 (10) ◽

pp. 2787-2792 ◽

Cited By ~ 14

Author(s):

Atsuko Gyohda ◽

Teruya Komano

Keyword(s):

Electrophoretic Analysis ◽

Recombination Reaction ◽

Site Specific ◽

Dna Inversion ◽

Repeat Sequences ◽

In Vitro Recombination ◽

Recombinase Gene ◽

A Site

ABSTRACT The shufflon, a multiple DNA inversion system in plasmid R64, consists of four invertible DNA segments which are separated and flanked by seven 19-bp repeat sequences. The product of a site-specific recombinase gene, rci, promotes site-specific recombination between any two of the inverted 19-bp repeat sequences of the shufflon. To analyze the molecular mechanism of this recombination reaction, Rci protein was overproduced and purified. The purified Rci protein promoted the in vitro recombination reaction between the inverted 19-bp repeats of supercoiled DNA of a plasmid carrying segment A of the R64 shufflon. The recombination reaction was enhanced by the bacterial host factor HU. Gel electrophoretic analysis indicated that the Rci protein specifically binds to the DNA segments carrying the 19-bp sequences. The binding affinity of the Rci protein to the four shufflon segments as well as four synthetic 19-bp sequences differed greatly: among the four 19-bp repeat sequences, the repeat-a and -d sequences displayed higher affinity to Rci protein. These results suggest that the differences in the affinity of Rci protein for the 19-bp repeat sequences determine the inversion frequencies of the four segments.

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