BCM-7: Opioid-like Peptide with Potential Role in Disease Mechanisms.

Bovine milk is an essential supplement due to its rich energy- and nutrient-rich qualities. Caseins constitute the vast majority of the proteins in milk. Among these, ß-casein comprises around 37% of all caseins, and it is an important type of casein with several different variants. The A1 and A2 variants of ß-casein are the most researched genotypes due to the changes in their composition. It is accepted that the A2 variant is ancestral, while a point mutation in the 67th amino acid created the A1 variant. The digestion derived of both A1 and A2 milk is BCM-7. Digestion of A2 milk in the human intestine also forms BCM-9 peptide molecule. The opioid-like characteristics of BCM-7 are highlighted for their potential triggering effect on several diseases. Most research has been focused on gastrointestinal-related diseases; however other metabolic and nervous system-based diseases are also potentially triggered. By manipulating the mechanisms of these diseases, BCM-7 can induce certain situations, such as conformational changes, reduction in protein activity, and the creation of undesired activity in the biological system. Furthermore, the genotype of casein can also play a role in bone health, such as altering fracture rates, and calcium contents can change the characteristics of dietary products. The context between opioid molecules and BCM-7 points to a potential triggering mechanism for the central nervous system and other metabolic diseases discussed.

[Age-dependent expression of HSP90 in the hippocampus of APP/PS1 mice].

The present study aims to observe the change in expression of heat shock protein 90 (HSP90) along with amyloid-ß (Aß) and phosphorylated Tau (p-Tau) protein levels in the hippocampus tissue of Alzheimer's disease (AD) transgenic animal model with age. APP/PS1 transgenic mice at age of 6-, 9- and 12-month and C57BL/6J mice of the same age were used. The cognitive abilities of these animals were evaluated using a Morris water maze. Western blot or immunohistochemistry was used to detect the expressions of HSP90 and Aß1-42, as well as the phosphorylation levels of Tau protein in the hippocampus. The hsp90 mRNA levels and the morphology and number of cells in the hippocampus were detected with real-time quantitative polymerase chain reaction (qRT-PCR) and Nissl staining, respectively. The results showed that compared with C57BL/6J mice of the same age, HSP90 and hsp90 mRNA expression were decreased (P < 0.05 or P < 0.01), while Aß1-42 and p-Tau protein levels were increased (P < 0.05 or P < 0.01) in the hippocampal tissue of APP/PS1 transgenic mice. Meanwhile, the decrease in HSP90 and hsp90 mRNA expression (P < 0.05 or P < 0.01), the increase in Aß1-42 and p-Tau levels (P < 0.01 or P < 0.05) in hippocampal tissue and the reduction in behavioral ability showed a progressive development with the advancing of age in the APP/PS1 transgenic mice. In conclusion, in the hippocampal tissue of APP/PS1 mice, the decrease in HSP90 expression and the increase in Aß1-42 and p-Tau levels together with the decline of their cognitive ability are age-dependent.

A two-step regulatory circuit involving Spo0A-AbrB activates mersacidin biosynthesis in Bacillus subtilis.

Due to intramolecular ring structures, the ribosomally produced and post-translationally modified peptide mersacidin shows antimicrobial properties comparable to those of vancomycin without exhibiting cross-resistance. Although the principles of mersacidin biosynthesis are known, there is no information on the molecular control processes for the initial stimulation of mersacidin bioproduction. By using Bacillus subtilis for heterologous biosynthesis, a considerable amount of mersacidin could be produced without the mersacidin-specific immune system and the mersacidin-activating secretory protease. By using the established laboratory strain Bacillus subtilis 168 and strain 3NA, which is used for high cell density fermentation processes, in combination with the construction of reporter strains to determine the promoter strengths within the mersacidin core gene cluster, the molecular regulatory circuit of Spo0A, a master regulator of cell differentiation including sporulation initiation, and the global transcriptional regulator AbrB, which is involved in cell adaptation processes in the transient growth phase, was identified to control the initial stimulation of the mersacidin core gene cluster. In a second downstream regulatory step, the activator MrsR1, encoded in the core gene cluster, acts as a stimulatory element for mersacidin biosynthesis. These findings are important to understand the mechanisms linking environmental conditions and microbial responses with respect to the bioproduction of bioactive metabolites including antimicrobials such as mersacidin. This information will also support the construction of production strains for bioactive metabolites with antimicrobial properties.

Genome-wide epistasis analysis reveals gene-gene interaction network on an intermediate endophenotype P-tau/Aß42 ratio in ADNI cohort.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder and the most common cause of dementia in the elderly worldwide. The exact etiology of AD, particularly its genetic mechanisms, remains incompletely understood. Traditional genome-wide association studies (GWAS), which primarily focus on single-nucleotide polymorphisms (SNPs) with main effects, provide limited explanations for the "missing heritability" of AD, while there is growing evidence supporting the important role of epistasis. In this study, we performed a genome-wide SNP-SNP interaction detection using a linear regression model and employed multiple GPUs for parallel computing, significantly enhancing the speed of whole-genome analysis. The cerebrospinal fluid (CSF) phosphorylated tau (P-tau)/amyloid-[Formula: see text] (A[Formula: see text]) ratio was used as a quantitative trait (QT) to enhance statistical power. Age, gender, and clinical diagnosis were included as covariates to control for potential non-genetic factors influencing AD. We identified 961 pairs of statistically significant SNP-SNP interactions, explaining a high-level variance of P-tau/A[Formula: see text] level, all of which exhibited marginal main effects. Additionally, we replicated 432 previously reported AD-related genes and found 11 gene-gene interaction pairs overlapping with the protein-protein interaction (PPI) network. Our findings may contribute to partially explain the "missing heritability" of AD. The identified subnetwork may be associated with synaptic dysfunction, Wnt signaling pathway, oligodendrocytes, inflammation, hippocampus, and neuronal cells.

Cerebrospinal fluid neutral lipids predict progression from mild cognitive impairment to Alzheimer's disease.

Genetic, metabolic, and clinical evidence links lipid dysregulation to an increased risk of Alzheimer's disease (AD). However, the role of lipids in the pathophysiological processes of AD and its clinical progression is unclear. We investigated the association between cerebrospinal fluid (CSF) lipidome and the pathological hallmarks of AD, progression from mild cognitive impairment (MCI) to AD, and the rate of cognitive decline in MCI patients. The CSF lipidome was analyzed by liquid chromatography coupled to mass spectrometry in an LC-ESI-QTOF-MS/MS platform for 209 participants: 91 AD, 92 MCI, and 26 control participants. The MCI patients were followed up for a median of 58 (± 12.5) months to evaluate their clinical progression to AD. Forty-eight (52.2%) MCI patients progressed to AD during follow-up. We found that higher CSF levels of hexacosanoic acid and ceramide Cer(d38:4) were associated with an increased risk of amyloid beta 42 (Aß42) positivity in CSF, while levels of phosphatidylethanolamine PE(40:0) were associated with a reduced risk. Higher CSF levels of sphingomyelin SM(30:1) were positively associated with pathological levels of phosphorylated tau in CSF. Cholesteryl ester CE(11D3:1) and an unknown lipid were recognized as the most associated lipid species with MCI to AD progression. Furthermore, TG(O-52:2) was identified as the lipid most strongly associated with the rate of progression. Our results indicate the involvement of membrane and intracellular neutral lipids in the pathophysiological processes of AD and the progression from MCI to AD dementia. Therefore, CSF neutral lipids can be used as potential prognostic markers for AD.

Deep intronic variant causes aberrant splicing of ATP7A in a family with a variable occipital horn syndrome phenotype.

Genetic variants in ATP7A are associated with a spectrum of X-linked disorders. In descending order of severity, these are Menkes disease, occipital horn syndrome, and X-linked distal spinal muscular atrophy. After 30 years of diagnostic investigation, we identified a deep intronic ATP7A variant in four males from a family affected to variable degrees by a predominantly skeletal phenotype, featuring bowing of long bones, elbow joints with restricted mobility which dislocate frequently, coarse curly hair, chronic diarrhoea, and motor coordination difficulties. Analysis of whole genome sequencing data from the Genomics England 100,000 Genomes Project following clinical re-evaluation identified a deep intronic ATP7A variant, which was predicted by SpliceAI to have a modest splicing effect. Using a mini-gene splicing assay, we determined that the intronic variant results in aberrant splicing. Sanger sequencing of patient cDNA revealed ATP7A transcripts with exon 5 skipping, or inclusion of a novel intron 4 pseudoexon. In both instances, frameshift leading to premature termination are predicted. Quantification of ATP7A mRNA transcripts using a qPCR assay indicated that the majority of transcripts (86.1 %) have non-canonical splicing, with 68.0 % featuring exon 5 skipping, and 18.1 % featuring the novel pseudoexon. We suggest that the variability of the phenotypes within the affected males results from the stochastic effects of splicing. This deep intronic variant, resulting in aberrant ATP7A splicing, expands the understanding of intronic variation on the ATP7A-related disease spectrum.

Blood-Brain Barrier Dysfunction and Aß42/40 Ratio Dose-Dependent Modulation with the ApoE Genotype within the ATN Framework.

The definition of Alzheimer's disease (AD) now considers the presence of the markers of amyloid (A), tau deposition (T), and neurodegeneration (N) essential for diagnosis. AD patients have been reported to have increased blood-brain barrier (BBB) dysfunction, but that has not been tested within the ATN framework so far. As the field is moving towards the use of blood-based biomarkers, the relationship between BBB disruption and AD-specific biomarkers requires considerable attention. Moreover, other factors have been previously implicated in modulating BBB permeability, including age, gender, and ApoE status. A total of 172 cognitively impaired individuals underwent cerebrospinal fluid (CSF) analysis for AD biomarkers, and data on BBB dysfunction, demographics, and ApoE status were collected. Our data showed that there was no difference in BBB dysfunction across different ATN subtypes, and that BBB damage was not correlated with cognitive impairment. However, patients with BBB disruption, if measured with a high Qalb, had low Aß40 levels. ApoE status did not affect BBB function but had a dose-dependent effect on the Aß42/40 ratio. These results might highlight the importance of understanding dynamic changes across the BBB in future studies in patients with AD.

The underlying mechanism of transcription factor IRF1, PRDM1, and ZNF263 involved in the regulation of NPPB rs3753581 on pulse pressure hypertension.

To investigate the correlation between NPPB gene variants and pulse pressure hypertension and the underlying regulatory mechanisms and try to confirm that NPPB may be a potential molecular target of gene therapy for pulse pressure hypertension. A total of 898 participants were recruited from the First Affiliated Hospital of Fujian Medical University and the plasmids with differential expression of NPPB were constructed. Genotype distribution of NPPB(rs3753581, rs198388, and rs198389)was analyzed and the expression of N-terminal pro-B-type natriuretic peptide(NT-proBNP) and renin-angiotensin -aldosterone system(RAAS) related indicators were identified in the groups studied. According to a genotype analysis, there was a significant difference in the genotype distribution of NPPB rs3753581 among the groups (P = 0.034). In logistic regression analysis, NPPB rs3753581 TT was associated with a 1.8-fold greater risk of pulse pressure hypertension than NPPB rs3753581 GG (odds ratio = 1.801; 95% confidence interval: 1.070-3.032; P = 0.027). The expression of NT-proBNP and RAAS related indicators in clinical and laboratory samples showed striking differences. The activity of firefly and Renilla luciferase in pGL-3-NPPB-luc (-1299G) was higher than pGL-3-NPPBmut-luc(-1299 T)(P < 0.05). The binding of NPPB gene promoter rs3753581 (-1299G) with transcription factors IRF1, PRDM1, and ZNF263 was predicted and validated by the bioinformatics software TESS and chromatin immunoprecipitation(P < 0.05). NPPB rs3753581 was correlated with genetic susceptibility to pulse pressure hypertension and the transcription factors IRF1, PRDM1, and ZNF263 may be involved in the regulation of NPPB rs3753581 promoter (-1299G) on the expression of NT-proBNP/RAAS.

Genetic architecture of plasma Alzheimer disease biomarkers.

Genome-wide association studies (GWAS) of cerebrospinal fluid (CSF) Alzheimer's Disease (AD) biomarker levels have identified novel genes implicated in disease risk, onset and progression. However, lumbar punctures have limited availability and may be perceived as invasive. Blood collection is readily available and well accepted, but it is not clear whether plasma biomarkers will be informative for genetic studies. Here we perform genetic analyses on concentrations of plasma amyloid-ß peptides Aß40 (n = 1,467) and Aß42 (n = 1,484), Aß42/40 (n = 1467) total tau (n = 504), tau phosphorylated (p-tau181; n = 1079) and neurofilament light (NfL; n = 2,058). GWAS and gene-based analysis was used to identify single variant and genes associated with plasma levels. Finally, polygenic risk score and summary statistics were used to investigate overlapping genetic architecture between plasma biomarkers, CSF biomarkers and AD risk. We found a total of six genome-wide significant signals. APOE was associated with plasma Aß42, Aß42/40, tau, p-tau181 and NfL. We proposed 10 candidate functional genes on the basis of 12 single nucleotide polymorphism-biomarker pairs and brain differential gene expression analysis. We found a significant genetic overlap between CSF and plasma biomarkers. We also demonstrate that it is possible to improve the specificity and sensitivity of these biomarkers, when genetic variants regulating protein levels are included in the model. This current study using plasma biomarker levels as quantitative traits can be critical to identification of novel genes that impact AD and more accurate interpretation of plasma biomarker levels.

Familial Alzheimer's Disease-Related Mutations Differentially Alter Stability of Amyloid-Beta Aggregates.

Amyloid-beta (Aß) deposition as senile plaques is a pathological hallmark of Alzheimer's disease (AD). AD is characterized by a large level of heterogeneity in amyloid pathology, whose molecular origin is poorly understood. Here, we employ NMR spectroscopy and MD simulation at ambient and high pressures and investigate how AD-related mutations in Aß peptide influence the stability of Aß aggregates. The pressure-induced monomer dissociation from Aß aggregates monitored by NMR demonstrated that the Iowa (D23N), Arctic (E22G), and Osaka (ΔE22) mutations altered the pressure stability of Aß40 aggregates in distinct manners. While the NMR data of monomeric Aß40 showed only small localized effects of mutations, the MD simulation of mutated Aß fibrils revealed their distinct susceptibility to elevated pressure. Our data propose a structural basis for the distinct stability of various Aß fibrils and highlights "stability" as a molecular property potentially contributing to the large heterogeneity of amyloid pathology in AD.

ATP7A-related copper transport disorders: A systematic review and definition of the clinical subtypes.

In patients with ATP7A-related disorders, counseling is challenging due to clinical overlap between the entities, the absence of predictive biomarkers and a clear genotype-phenotype correlation. We performed a systematic literature review by querying the MEDLINE and Embase databases identifying 143 relevant papers. We recorded data on the phenotype and genotype in 162 individuals with a molecularly confirmed ATP7A-related disorder in order to identify differentiating clinical criteria, evaluate genotype-phenotype correlations and propose management guidelines. Early seizures are specific for classical Menkes disease (CMD), that is characterized by early-onset neurodegenerative disease with high mortality rates. Ataxia is an independent indicator for atypical Menkes disease, that shows better survival rates than CMD. Bony exostoses, radial head dislocations, herniations and dental abnormalities are specific for occipital horn syndrome (OHS) that may further present with developmental delay and connective tissue manifestations. Intracranial tortuosity and bladder diverticula, both with high risk of complications, are common among all subtypes. Low ceruloplasmin is a more sensitive and discriminating biomarker for ATP7A-related disorders than serum copper. Truncating mutations are frequently associated with CMD, in contrast with splice site and intronic mutations which are more prevalent in OHS.

A Genome-Wide Functional Screen Identifies Enhancer and Protective Genes for Amyloid Beta-Peptide Toxicity.

Alzheimer's disease (AD) is known to be caused by amyloid ß-peptide (Aß) misfolded into ß-sheets, but this knowledge has not yet led to treatments to prevent AD. To identify novel molecular players in Aß toxicity, we carried out a genome-wide screen in Saccharomyces cerevisiae, using a library of 5154 gene knock-out strains expressing Aß1-42. We identified 81 mammalian orthologue genes that enhance Aß1-42 toxicity, while 157 were protective. Next, we performed interactome and text-mining studies to increase the number of genes and to identify the main cellular functions affected by Aß oligomers (oAß). We found that the most affected cellular functions were calcium regulation, protein translation and mitochondrial activity. We focused on SURF4, a protein that regulates the store-operated calcium channel (SOCE). An in vitro analysis using human neuroblastoma cells showed that SURF4 silencing induced higher intracellular calcium levels, while its overexpression decreased calcium entry. Furthermore, SURF4 silencing produced a significant reduction in cell death when cells were challenged with oAß1-42, whereas SURF4 overexpression induced Aß1-42 cytotoxicity. In summary, we identified new enhancer and protective activities for Aß toxicity and showed that SURF4 contributes to oAß1-42 neurotoxicity by decreasing SOCE activity.

A minimal construct of nuclear-import receptor Karyopherin-ß2 defines the regions critical for chaperone and disaggregation activity.

Karyopherin-ß2 (Kapß2) is a nuclear-import receptor that recognizes proline-tyrosine nuclear localization signals of diverse cytoplasmic cargo for transport to the nucleus. Kapß2 cargo includes several disease-linked RNA-binding proteins with prion-like domains, such as FUS, TAF15, EWSR1, hnRNPA1, and hnRNPA2. These RNA-binding proteins with prion-like domains are linked via pathology and genetics to debilitating degenerative disorders, including amyotrophic lateral sclerosis, frontotemporal dementia, and multisystem proteinopathy. Remarkably, Kapß2 prevents and reverses aberrant phase transitions of these cargoes, which is cytoprotective. However, the molecular determinants of Kapß2 that enable these activities remain poorly understood, particularly from the standpoint of nuclear-import receptor architecture. Kapß2 is a super-helical protein comprised of 20 HEAT repeats. Here, we design truncated variants of Kapß2 and assess their ability to antagonize FUS aggregation and toxicity in yeast and FUS condensation at the pure protein level and in human cells. We find that HEAT repeats 8 to 20 of Kapß2 recapitulate all salient features of Kapß2 activity. By contrast, Kapß2 truncations lacking even a single cargo-binding HEAT repeat display reduced activity. Thus, we define a minimal Kapß2 construct for delivery in adeno-associated viruses as a potential therapeutic for amyotrophic lateral sclerosis/frontotemporal dementia, multisystem proteinopathy, and related disorders.

Associations of the APOE ε2 and ε4 alleles and polygenic profiles comprising APOE-TOMM40-APOC1 variants with Alzheimer's disease biomarkers.

Capturing the genetic architecture of Alzheimer's disease (AD) is challenging because of the complex interplay of genetic and non-genetic factors in its etiology. It has been suggested that AD biomarkers may improve the characterization of AD pathology and its genetic architecture. Most studies have focused on connections of individual genetic variants with AD biomarkers, whereas the role of combinations of genetic variants is substantially underexplored. We examined the associations of the APOE ε2 and ε4 alleles and polygenic profiles comprising the ε4-encoding rs429358, TOMM40 rs2075650, and APOC1 rs12721046 polymorphisms with cerebrospinal fluid (CSF) and plasma amyloid ß (Aß40 and Aß42) and tau biomarkers. Our findings support associations of the ε4 alleles with both plasma and CSF Aß42 and CSF tau, and the ε2 alleles with baseline, but not longitudinal, CSF Aß42 measurements. We found that the ε4-bearing polygenic profiles conferring higher and lower AD risks are differentially associated with tau but not Aß42. Modulation of the effect of the ε4 alleles by TOMM40 and APOC1 variants indicates the potential genetic mechanism of differential roles of Aß and tau in AD pathogenesis.

Key Residue for Aggregation of Amyloid-ß Peptides.

It is known that oligomers of amyloid-ß (Aß) peptide are associated with Alzheimer's disease. Aß has two isoforms: Aß40 and Aß42. Although the difference between Aß40 and Aß42 is only two additional C-terminal residues, Aß42 aggregates much faster than Aß40. It is unknown what role the C-terminal two residues play in accelerating aggregation. Since Aß42 is more toxic than Aß40, its oligomerization process needs to be clarified. Moreover, clarifying the differences between the oligomerization processes of Aß40 and Aß42 is essential to elucidate the key factors of oligomerization. Therefore, to investigate the dimerization process, which is the early oligomerization process, Hamiltonian replica-permutation molecular dynamics simulations were performed for Aß40 and Aß42. We identified a key residue, Arg5, for the Aß42 dimerization. The two additional residues in Aß42 allow the C-terminus to form contact with Arg5 because of the electrostatic attraction between them, and this contact stabilizes the ß-hairpin. This ß-hairpin promotes dimer formation through the intermolecular ß-bridges. Thus, we examined the effects of amino acid substitutions of Arg5, thereby confirming that the mutations remarkably suppressed the aggregation of Aß42. Moreover, the mutations of Arg5 suppressed the Aß40 aggregation. It was found by analyzing the simulations that Arg5 is important for Aß40 to form intermolecular contacts. Thus, it was clarified that the role of Arg5 in the oligomerization process varies due to the two additional C-terminal residues.

3D Blockage Mapping for Identifying Familial Point Mutations in Single Amyloid-ß Peptides with a Nanopore.

Accurate discrimination of amyloid-ß (Aß) peptides containing familial point mutations would advance the knowledge of their roles in early-onset Alzheimer's disease. Herein, we simultaneously identified the mutant A21G, E22G, E22Q, and the wild-type (WT) Aß18-26 peptides with aerolysin nanopore using a 3D blockage mapping strategy. The standard deviation of current blockade fluctuations (σb ) was proposed as a new supplement to current blockage (Ib /I0 ) and duration time (tD ) to profile the blockage characteristics of single molecules. Although the WT and A21G Aß18-26 are indistinguishable in a traditional Ib /I0 -tD 2D description, ∼87 % of the blockade events can be accurately classified with half reduction of false identification using a combination of Ib /I0 , tD, and σb . This work offers an easy and reliable strategy to promote nanopore sensitivity of peptide mutants, leading to a more precise analysis of pathogenic mutations for developing effective diagnosis and treatment.

HER2 c-Terminal Fragments Are Expressed via Internal Translation of the HER2 mRNA.

The HER2/neu signaling pathway is one of the most frequently mutated in human cancer. Although therapeutics targeting this pathway have good efficacy, cancer cells frequently develop resistance. The HER2 gene encodes the full-length HER2 protein, as well as smaller c-terminal fragments (CTFs), which have been shown to be a cause of resistance. Here, we show that HER2 CTFs, exclusive from the full-length HER2 protein, are generated via internal translation of the full-length HER2 mRNA and identify regions which are required for this mechanism to occur. These regions of the HER2 mRNA may present novel sites for therapeutic intervention via small molecules or antisense oligonucleotides (ASOs).

Presepsin Predicts Severity and Secondary Bacterial Infection in COVID-19 by Bioinformatics Analysis.

Introduction: Novel coronavirus pneumonia (COVID-19) is an acute respiratory disease caused by the novel coronavirus SARS-CoV-2. Severe and critical illness, especially secondary bacterial infection (SBI) cases, accounts for the vast majority of COVID-19-related deaths. However, the relevant biological indicators of COVID-19 and SBI are still unclear, which significantly limits the timely diagnosis and treatment. Methods: The differentially expressed genes (DEGs) between severe COVID-19 patients with SBI and without SBI were screened through the analysis of GSE168017 and GSE168018 datasets. By performing Gene Ontology (GO) enrichment analysis for significant DEGs, significant biological processes, cellular components, and molecular functions were selected. To understand the high-level functions and utilities of the biological system, the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was performed. By analyzing protein-protein interaction (PPI) and key subnetworks, the core DEGs were found. Results: 85 DEGs were upregulated, and 436 DEGs were downregulated. The CD14 expression was significantly increased in the SBI group of severe COVID-19 patients (P < 0.01). The area under the curve (AUC) of CD14 in the SBI group in severe COVID-19 patients was 0.9429. The presepsin expression was significantly higher in moderate to severe COVID-19 patients (P < 0.05). Presepsin has a diagnostic value for moderate to severe COVID-19 with the AUC of 0.9732. The presepsin expression of COVID-19 patients in the nonsurvivors was significantly higher than that in the survivors (P < 0.05). Conclusion: Presepsin predicts severity and SBI in COVID-19 and may be associated with prognosis in COVID-19.

Investigating the effect of bacteriophages on bacterial FtsZ localisation.

Escherichia coli is one of the most common Gram-negative pathogens and is responsible for infection leading to neonatal meningitis and sepsis. The FtsZ protein is a bacterial tubulin homolog required for cell division in most species, including E. coli. Several agents that block cell division have been shown to mislocalise FtsZ, including the bacteriophage λ-encoded Kil peptide, resulting in defective cell division and a filamentous phenotype, making FtsZ an attractive target for antimicrobials. In this study, we have used an in vitro meningitis model system for studying the effect of bacteriophages on FtsZ using fluorescent E. coli EV36/FtsZ-mCherry and K12/FtsZ-mNeon strains. We show localisation of FtsZ to the bacterial cell midbody as a single ring during normal growth conditions, and mislocalisation of FtsZ producing filamentous multi-ringed bacterial cells upon addition of the known inhibitor Kil peptide. We also show that when bacteriophages K1F-GFP and T7-mCherry were applied to their respective host strains, these phages can inhibit FtsZ and block bacterial cell division leading to a filamentous multi-ringed phenotype, potentially delaying lysis and increasing progeny number. This occurs in the exponential growth phase, as actively dividing hosts are needed. We present that ZapA protein is needed for phage inhibition by showing a phenotype recovery with a ZapA mutant strain, and we show that FtsI protein is also mislocalised upon phage infection. Finally, we show that the T7 peptide gp0.4 is responsible for the inhibition of FtsZ in K12 strains by observing a phenotype recovery with a T7Δ0.4 mutant.

Tryptophan degradation enzymes and Angiotensin (1-7) expression in human placenta.

Indoleamine 2,3-dioxygenase 1 (IDO1) and tryptophan 2,3-dioxygenase (TDO) are key enzymes for tryptophan degradation, regulating immune tolerance during pregnancy. The intrauterine renin-angiotensin system is also involved in the progression of a healthy pregnancy. Angiotensin(1-7) maintains the integrity of fetal membranes via counteracting the pro-inflammatory actions of Angiotensin II. No data are available on placental Angiotensin(1-7) co-expression with TDO. We aimed to characterize TDO mRNA expression and its localization in different areas of the placenta of physiological pregnancies delivered at term; its co-expression with Angiotensin(1-7) and its correlation with the plasma kynurenine/tryptophan (Kyn/Trp) ratio was investigated. This prospective observational study included a nonconsecutive series of 20 singleton uncomplicated pregnancies delivered vaginally. TDO mRNA was expressed in both maternal and fetal sides of the placentas and TDO protein also in the villi and it was co-expressed with IDO1 in almost half of the placental cells at these sites. The percentage of TDO+ and IDO1+ cells appeared to be influenced by maternal pre-gestational smoking and newborn weight. A strong correlation was found between the percentage of TDO+ and IDO1+ cells in the villi. TDO+ cells also expressed Angiotensin(1-7), with a higher percentage on the fetal side and in the villi compared to the maternal one. Kyn/Trp plasma ratio was not correlated with IDO and TDO expression nor with the patient's characteristics. Collectively, our data indicate that TDO is detectable in placental tissue and is co-expressed with IDO and with Angiotensin(1-7)+ on the fetal side and in the villi.