Ramachandran plot for the predicted tertiary structure of the Msa protein pre (A) and post (B) refinement

<p><b>Copyright information:</b></p><p>Taken from "Structure and function predictions of the Msa protein in "</p><p>http://www.biomedcentral.com/1471-2105/8/S7/S5</p><p>BMC Bioinformatics 2007;8(Suppl 7):S5-S5.</p><p>Published online 1 Nov 2007</p><p>PMCID:PMC2099497.</p><p></p

GeneVenn - A web application for comparing gene lists using Venn diagrams-1

<p><b>Copyright information:</b></p><p>Taken from "GeneVenn - A web application for comparing gene lists using Venn diagrams"</p><p>Bioinformation 2007;1(10):420-422.</p><p>Published online 10 Apr 2007</p><p>PMCID:PMC1899164.</p><p></p

GeneVenn - A web application for comparing gene lists using Venn diagrams-0

<p><b>Copyright information:</b></p><p>Taken from "GeneVenn - A web application for comparing gene lists using Venn diagrams"</p><p>Bioinformation 2007;1(10):420-422.</p><p>Published online 10 Apr 2007</p><p>PMCID:PMC1899164.</p><p></p

A Novel Specificity Protein 1 (SP1)-like Gene Regulating Protein Kinase C-1 (Pkc1)-dependent Cell Wall Integrity and Virulence Factors in Cryptococcus neoformans

Eukaryotic cells utilize complex signaling systems to detect their environments, responding and adapting as new conditions arise during evolution. The basidiomycete fungus Cryptococcus neoformans is a leading cause of AIDS-related death worldwide and utilizes the calcineurin and protein kinase C-1 (Pkc1) signaling pathways for host adaptation and expression of virulence. In the present studies, a C-terminal zinc finger transcription factor, homologous to both the calcineurin responsive zinc fingers (Crz1) of ascomycetes and to the Pkc1 dependent specificity protein-1 (Sp1) transcription factors of metazoans, was identified and named SP1 because of its greater similarity to the metazoan factors. Structurally, the Cn Sp1 protein was found to have acquired an additional Zn finger motif from that of Crz1 and showed Pkc1 dependent phosphorylation, nuclear localization and whole genome epistatic associations under starvation conditions. Transcriptional targets of Cn Sp1 shared functional similarities with Crz1 factors such as cell wall synthesis, but gained the regulation of processes involved in carbohydrate metabolism including trehalose metabolism and lost others such as the induction of autophagy. In addition, overexpression of Cn Sp1 in a pkc1Δ mutant showed restoration of altered phenotypes involved in virulence including cell wall stability, nitrosative stress and extracellular capsule production. Cn Sp1 was also found to be important for virulence of the fungus using a mouse model. In summary, these data suggest an evolutionary shift in C-terminal Zn finger proteins during fungal evolution, transforming them from calcineurin-dependent to PKC1- dependent transcription factors, helping to shape the role of fungal pathogenesis of Cryptococcus neoformans

Development of Genetic System to Inactivate a <i>Borrelia turicatae</i> Surface Protein Selectively Produced within the Salivary Glands of the Arthropod Vector

<div><p>Background</p><p><i>Borrelia turicatae</i>, an agent of tick-borne relapsing fever, is an example of a pathogen that can adapt to disparate conditions found when colonizing the mammalian host and arthropod vector. However, little is known about the genetic factors necessary during the tick-mammalian infectious cycle, therefore we developed a genetic system to transform this species of spirochete. We also identified a plasmid gene that was up-regulated <i>in vitro</i> when <i>B. turicatae</i> was grown in conditions mimicking the tick environment. This 40 kilodalton protein was predicted to be surface localized and designated the <i>Borrelia repeat protein A</i> (<i>brpA</i>) due to the redundancy of the amino acid motif Gln-Gly-Asn-Val-Glu.</p><p>Methodology/Principal Findings</p><p>Quantitative reverse-transcriptase polymerase chain reaction using RNA from <i>B. turicatae</i> infected ticks and mice indicated differential regulation of <i>brpA</i> during the tick-mammalian infectious cycle. The surface localization was determined, and production of the protein within the salivary glands of the tick was demonstrated. We then applied a novel genetic system for <i>B. turicatae</i> to inactivate <i>brpA</i> and examined the role of the gene product for vector colonization and the ability to establish murine infection.</p><p>Conclusions/Significance</p><p>These results demonstrate the complexity of protein production in a population of spirochetes within the tick. Additionally, the development of a genetic system is important for future studies to evaluate the requirement of specific <i>B. turicatae</i> genes for vector colonization and transmission.</p></div

Inactivation of <i>brpA</i> by allelic exchange.

<p>The gentamicin acetyl transferase gene was flanked by 1,000- and down-stream of <i>brpA</i> forming P<i>_flgB-gent</i>, and <i>brpA</i> was displaced by allelic exchange (A). Deletion of <i>brpA</i> was confirmed by PCR analysis using primers located within <i>gent</i> and 1,000 bp downstream of the gene (A and B), primers within <i>brpA</i> (C), and <i>flaB</i> (D). Arrow heads (A) indicate the primer location and the expected amplicon size. Molecular masses are shown at the left of the gel.</p

Loss of BrpA production in <i>ΔbrpA</i> mutants.

<p>Wild type and <i>ΔbrpA</i> mutants were grown at 22°C and immunoblotting performed using rabbit serum generated against the recombinant protein (A) and pre-immunization rabbit serum (B). The asterisk and arrowhead indicate the expected size of BrpA and FlaB, respectively.</p

IF-LSCM of cryosectioned ticks.

<p>20 µm longitudinal cryosections (A) were double labeled for FlaB (B and C) and BrpA (D and E) using chicken and rabbit serum samples generated against each recombinant protein. Pre-immunization serum samples were used as negative controls on infected ticks (F and G). The secondary antibodies used were Alexa Fluor anti-chicken IgY 568 and anti-rabbit IgG 488. 5 µm bars are shown in each panel.</p

Differential regulation of <i>brpA</i>.

<p>Differential regulation of <i>brpA</i>.</p

IF-LSCM of spirochetes cultivated at 22°C.

<p>Chicken and rabbit serum generated against rFlaB (A) and rBrpA (B), respectively, were used to determine protein production within a population of spirochetes. Images were overlaid and spirochetes producing BrpA were counted (C). Chicken (D) and rabbit (E) preimmunization serum samples were used as negative controls. The secondary antibodies used were Alexa Fluor anti-chicken IgY 568 and anti-rabbit IgG 488. 5 µm bars are shown in each panel.</p