The homologous operons for P1 and P7 plasmid partition are autoregulated from dissimilar operator sites

The plasmid-partition regions of the P1 and P7 plasmid prophages in Escherichia coli are homologues which each encode two partition proteins, ParA and ParB. The equivalent PI and P7 proteins are closely related. In each case, the proteins are encoded by an operon that is autoregulated by the ParA and ParB proteins in concert. This regulation is species-specific, as the P1 proteins are unable to repress the P7 par operon and vice versa. The homologous ParA proteins are primarily responsible for repression and bind to regions that overlap the operon promoter in both cases. The DNA-binding domain of the P7 auto-repressor lies in the amino-terminal end of the P7 ParA protein. This region includes a helix-turn-helix motif that has a clear counterpart in the P1 ParA sequence. However, despite the common regulatory mechanism and the similarity of the proteins involved in repression, the promoter-operator sequences of these two operons are very different in sequence and organization. The operator is located downstream of the promoter in P1 and upstream of it in P7, and the two regions show little, if any, homology. How these differences may have arisen from a common ancestral form is discussed.     

mTOR activity is essential for retinal pigment epithelium regeneration in zebrafish

The retinal pigment epithelium (RPE) plays numerous critical roles in maintaining vision and this is underscored by the prevalence of degenerative blinding diseases like age-related macular degeneration (AMD), in which visual impairment is caused by progressive loss of RPE cells. In contrast to mammals, zebrafish possess the ability to intrinsically regenerate a functional RPE layer after severe injury. The molecular underpinnings of this regenerative process remain largely unknown yet hold tremendous potential for developing treatment strategies to stimulate endogenous regeneration in the human eye. In this study, we demonstrate that the mTOR pathway is activated in RPE cells post-genetic ablation. Pharmacological and genetic inhibition of mTOR activity impaired RPE regeneration, while mTOR activation enhanced RPE recovery post-injury, demonstrating that mTOR activity is essential for RPE regeneration in zebrafish. RNA-seq of RPE isolated from mTOR-inhibited larvae identified a number of genes and pathways dependent on mTOR activity at early and late stages of regeneration; amongst these were components of the immune system, which is emerging as a key regulator of regenerative responses across various tissue and model systems. Our results identify crosstalk between macrophages/microglia and the RPE, wherein mTOR activity is required for recruitment of macrophages/microglia to the RPE injury site. Macrophages/microglia then reinforce mTOR activity in regenerating RPE cells. Interestingly, the function of macrophages/microglia in maintaining mTOR activity in the RPE appeared to be inflammation-independent. Taken together, these data identify mTOR activity as a key regulator of RPE regeneration and link the mTOR pathway to immune responses in facilitating RPE regeneration.     

Identification of nucleotide sequences for the specific and rapid detection of Yersinia pestis

Suppression subtractive hybridization, a cost-effective approach for targeting unique DNA, was used to identify a 41.7-kb Yersinia pestis-specific region. One primer pair designed from this region amplified PCR products from natural isolates of Y. pestis and produced no false positives for near neighbors, an important criterion for unambiguous bacterial identification.     

Bordetella pertussis lipopolysaccharide resists the bactericidal effects of pulmonary surfactant protein A

Surfactant protein A (SP-A) plays an important role in the innate immune defense of the respiratory tract. SP-A binds to lipid A of bacterial LPS, induces aggregation, destabilizes bacterial membranes, and promotes phagocytosis by neutrophils and macrophages. In this study, SP-A interaction with wild-type and mutant LPS of Bordetella pertussis, the causative agent of whooping cough, was examined. B. pertussis LPS has a branched core structure with a nonrepeating trisaccharide, rather than a long-chain repeating O-Ag. SP-A did not bind, aggregate, nor permeabilize wild-type B. pertussis. LPS mutants lacking even one of the sugars in the terminal trisaccharide were bound and aggregated by SP-A. SP-A enhanced phagocytosis by human monocytes of LPS mutants that were able to bind SP-A, but not wild-type bacteria. SP-A enhanced phagocytosis by human neutrophils of LPS-mutant strains, but only in the absence of functional adenylate cyclase toxin, a B. pertussis toxin that has been shown to depress neutrophil activity. We conclude that the LPS of wild-type B. pertussis shields the bacteria from SP-A-mediated clearance, possibly by sterically limiting access to the lipid A region.     

Genomic heterogeneity and instability in colorectal cancer: spectral karyotyping, glutathione transferase-Ml and ras

Genomic instability in cancer is frequently described as being either chromosomal instability or microsatellite instability, although when events within chromosomes are monitored, extensive intrachromosomal instability is also found. Spectral karyotyping was used to visualize how extensively genomic instability gives rise to intratumor genomic heterogeneity in sporadic colorectal carcinomas. Two factors were then examined which might relate to intrachromosomal instability in colorectal cancers: the presence of the glutathione transferase-Ml gene to detoxify potential carcinogens, and the presence of activated ras which has been associated with chromosomal instability when first expressed. Intrachromosomal genomic instability was previously determined by inter-(simple sequence repeat) PCR (inter-SSR PCR) and by fractional allelic loss rate for 348 markers. GSTM1 status was determined for each of 49 tumors through use of specific PCR, and 28 of the tumors showed the GSTM1 null genotype. A significant association was found between GSTMl-null status and elevated inter-(simple sequence repeat) PCR instability. In contrast, no association was found with fractional allelic loss rate. The first exons of the K-ras and H-ras oncogenes were sequenced in 72 colorectal cancers; 19 of the tumors had a mutation in codon 12 of the K-ras gene (24.5%), but no H-ras mutations were found. A weak correlation (p=0.10) was observed between mutant K-ras and inter-(simple sequence repeat) PCR genomic instability, and no association existed with fractional allelic loss rate.     

Recursive SVM feature selection and sample classification for mass-spectrometry and microarray data

Background  

Like microarray-based investigations, high-throughput proteomics techniques require machine learning algorithms to identify biomarkers that are informative for biological classification problems. Feature selection and classification algorithms need to be robust to noise and outliers in the data.     

Hematological indicators of stress in longline-captured sharks

For many shark species, little information exists about the stress response to capture and release in commercial longline fisheries. Recent studies have used hematological profiling to assess the secondary stress response, but little is known about how, and to what degree, these indicators vary interspecifically. Moreover, there is little understanding of the extent to which the level of relative swimming activity (e.g., sluggish vs. active) or the general ecological classification (e.g., coastal vs. pelagic) correlates to the magnitude of the exercise-induced (capture-related) stress response. This study compared plasma electrolytes (Na(+), Cl(-), Mg(2+), Ca(2+), and K(+)), metabolites (glucose and lactate), blood hematocrit, and heat shock protein (Hsp70) levels between 11 species of longline-captured sharks (n=164). Statistical comparison of hematological parameters revealed species-specific differences in response to longline capture, as well as differences by ecological classification. Taken together, the blood properties of longline-captured sharks appear to be useful indicators of interspecific variation in the secondary stress response to capture, and may prove useful in the future for predicting survivorship of longline-captured sharks where new technologies (i.e., pop-up satellite tags) can verify post-release mortality.