Characterization of novel extracellular proteases produced by Acanthamoeba castellanii after contact with human corneal epithelial cells and their relevance to pathogenesis.

BACKGROUND: Proteases produced by Acanthamoeba spp. play an important role in their virulence and may be the key to understanding Acanthamoeba pathogenesis; thus, increasing attention has been directed towards these proteins. The present study aimed to investigate the lytic factors produced by Acanthamoeba castellanii during the first hours of in vitro co-culture with human corneal epithelial cells (HCECs). METHODS: We used one old and one recent Acanthamoeba isolate, both from patients with severe keratitis, and subsets of these strains with enhanced pathogenic potential induced by sequential passaging over HCEC monolayers. The proteolytic profiles of all strains and substrains were examined using 1D in-gel zymography. RESULTS: We observed the activity of additional proteases (ranging from 33 to 50 kDa) during the early interaction phase between amoebae and HCECs, which were only expressed for a short time. Based on their susceptibilities to protease inhibitors, these proteases were characterized as serine proteases. Protease activities showed a sharp decline after 4 h of co-incubation. Interestingly, the expression of Acanthamoeba mannose-binding protein did not differ between amoebae in monoculture and those in co-culture. Moreover, we observed the activation of matrix metalloproteinases in HCECs after contact with Acanthamoeba. CONCLUSIONS: This study revealed the involvement of two novel serine proteases in Acanthamoeba pathogenesis and suggests a pivotal role of serine proteases during Acanthamoeba-host cell interaction, contributing to cell adhesion and lysis.

Biodegradation of Keratin Waste by Bacillus velezensis HFS_F2 through Optimized Keratinase Production Medium.

Enormous aggregates of keratinous wastes are produced annually by the poultry and leather industries which cause environmental degradation globally. To combat this issue, microbially synthesized extracellular proteases known as keratinase are used widely which is effective in degrading keratin found in hair and feathers. In the present work, keratinolytic bacteria were isolated from poultry farm soil and feather waste, and various cultural conditions were optimized to provide the highest enzyme production for efficient keratin waste degradation. Based on the primary and secondary screening methods, the potent keratinolytic strain (HFS_F2T) with the highest enzyme activity 32.65 ± 0.16 U/mL was genotypically characterized by 16S rRNA sequencing and was confirmed as Bacillus velezensis HFS_F2T ON556508. Through one-variable-at-a-time approach (OVAT), the keratinase production medium was optimized with sucrose (carbon source), beef extract (nitrogen source) pH-7, inoculum size (5%), and incubation at 37 °C). The degree of degradation (%DD) of keratin wastes was evaluated after 35 days of degradation in the optimized keratinase production medium devoid of feather meal under submerged fermentation conditions. Further, the deteriorated keratin wastes were visually examined and the hydrolysed bovine hair with 77.32 ± 0.32% degradation was morphologically analysed through Field Emission Scanning Electron Microscopy (FESEM) to confirm the structural disintegration of the cuticle. Therefore, the current study would be a convincing strategy for reducing the detrimental impact of pollutants from the poultry and leather industries by efficient keratin waste degradation through the production of microbial keratinase.

Bacillus velezensis strain NA16 shows high poultry feather-degrading efficiency, protease and amino acid production.

Isolated Bacillus velezensis strain NA16, which produces proteases, amino acids and the transcription levels of different keratinolytic enzymes and disulfide reductase genes in whole gene sequencing, was evaluated during feather degradation. The result shows under optimum fermentation conditions, chicken feather fermentation showed total amino acid concentration of 7599 mg/L, degradation efficiency of 99.3% at 72 h, and protease activity of 1058 U/mL and keratinase activity of 288 U/mL at 48 h. Goose feather fermentation showed total amino acid concentration of 4918 mg/L (96 h), and degradation efficiency was 98.9% at 120 h. Chicken feather fermentation broth at 72 h showed high levels of 17 amino acids, particularly phenylalanine (1050 ± 1.90 mg/L), valine (960 ± 1.04 mg/L), and glutamic (950 ± 3.00 mg/L). Scanning electron microscopy and Fourier transform infrared analysis revealed the essential role of peptide bond cleavage in structural changes and degradation of feathers. Protein purification and zymographic analyses revealed a key role in feather degradation of the 39-kDa protein encoded by gene1031, identified as an S8 family serine peptidase. Whole genome sequencing of NA16 revealed 26 metalloproteinase genes and 22 serine protease genes. Among the proteins, S8 family serine peptidase (gene1031, gene1428) and S9 family peptidase (gene3132) were shown by transcription analysis to play major roles in chicken feather degradation. These findings revealed the transcription levels of different families of keratinolytic enzymes in the degradation of feather keratin by microorganisms, and suggested potential applications of NA16 in feather waste management and amino acid production.

Low frequency magnetic field assisted production of acidic protease by Aspergillus niger.

Acid protease is widely used in industries such as food processing and feed additives. In the study, low frequency magnetic field (LF-MF) as an aid enhances acid protease production by Aspergillus niger (A. niger). The study assessed mycelial biomass, the enzymic activity of the acidic protease and underlying mechanism. At low intensities, alternating magnetic field (AMF) is more effective than static magnetic fields (SMF). Under optimal magnetic field conditions, acid protease activity and biomass increased by 91.44% and 16.31%, as compared with the control, respectively. Maximum 19.87% increase in enzyme activity after magnetic field treatment of crude enzyme solution in control group. Transcriptomics analyses showed that low frequency alternating magnetic field (LF-AMF) treatment significantly upregulated genes related to hydrolases and cell growth. Our results showed that low-frequency magnetic fields can enhance the acid protease production ability of A. niger, and the effect of AMF is better at low intensities. The results revealed that the effect of magnetic field on the metabolic mechanism of A. niger and provided a reference for magnetic field-assisted fermentation of A. niger.

Purification, functional characterization and enhanced production of serratiopeptidase from Serratia marcescens MES-4: An endophyte isolated from Morus rubra.

Serratiopeptidase, a proteolytic enzyme serves as an important anti-inflammatory and analgesic medication. Present study reports the production and purification of extracellular serratiopeptidase from an endophyte, Serratia marcescens MES-4, isolated from Morus rubra. Purification of the enzyme by Ion exchange chromatography led to the specific activity of 13,030 U/mg protein of serratiopeptidase, showcasing about 3.1 fold enhanced activity. The catalytic domain of the purified serratiopeptidase, composed of Zn coordinated with three histidine residues (His 209, His 213, and His 219), along with glutamate (Glu 210) and tyrosine (Tyr 249). The molecular mass, as determined by SDS-PAGE was ∼51 kDa. The purified serratiopeptidase displayed optimal activity at pH 9.0, temperature 50°C. Kinetic studies revealed Vmax and Km values of 33,333 U/mL and 1.66 mg/mL, respectively. Further, optimized conditions for the production of serratiopeptidase by Taguchi design led to the productivity of 87 U/mL/h with 87.9 fold enhanced production as compared to the previous conditions.

Exploring the role of symbiotic modifier peptidases in the legume - rhizobium symbiosis.

Legumes can establish a mutual association with soil-derived nitrogen-fixing bacteria called 'rhizobia' forming lateral root organs called root nodules. Rhizobia inside the root nodules get transformed into 'bacteroids' that can fix atmospheric nitrogen to ammonia for host plants in return for nutrients and shelter. A substantial 200 million tons of nitrogen is fixed annually through biological nitrogen fixation. Consequently, the symbiotic mechanism of nitrogen fixation is utilized worldwide for sustainable agriculture and plays a crucial role in the Earth's ecosystem. The development of effective nitrogen-fixing symbiosis between legumes and rhizobia is very specialized and requires coordinated signaling. A plethora of plant-derived nodule-specific cysteine-rich (NCR or NCR-like) peptides get actively involved in this complex and tightly regulated signaling process of symbiosis between some legumes of the IRLC (Inverted Repeat-Lacking Clade) and Dalbergioid clades and nitrogen-fixing rhizobia. Recent progress has been made in identifying two such peptidases that actively prevent bacterial differentiation, leading to symbiotic incompatibility. In this review, we outlined the functions of NCRs and two nitrogen-fixing blocking peptidases: HrrP (host range restriction peptidase) and SapA (symbiosis-associated peptidase A). SapA was identified through an overexpression screen from the Sinorhizobium meliloti 1021 core genome, whereas HrrP is inherited extra-chromosomally. Interestingly, both peptidases affect the symbiotic outcome by degrading the NCR peptides generated from the host plants. These NCR-degrading peptidases can shed light on symbiotic incompatibility, helping to elucidate the reasons behind the inefficiency of nitrogen fixation observed in certain groups of rhizobia with specific legumes.

Decoupling between the genetic potential and the metabolic regulation and expression in microbial organic matter cleavage across microbiomes.

Metagenomics, metatranscriptomics, and metaproteomics are used to explore the microbial capability of enzyme secretion, but the links between protein-encoding genes and corresponding transcripts/proteins across ecosystems are underexplored. By conducting a multi-omics comparison focusing on key enzymes (carbohydrate-active enzymes [CAZymes] and peptidases) cleaving the main biomolecules across distinct microbiomes living in the ocean, soil, and human gut, we show that the community structure, functional diversity, and secretion mechanisms of microbial secretory CAZymes and peptidases vary drastically between microbiomes at metagenomic, metatranscriptomic, and metaproteomic levels. Such variations lead to decoupled relationships between CAZymes and peptidases from genetic potentials to protein expressions due to the different responses of key players toward organic matter sources and concentrations. Our results highlight the need for systematic analysis of the factors shaping patterns of microbial cleavage on organic matter to better link omics data to ecosystem processes. IMPORTANCE: Omics tools are used to explore adaptive mechanism of microbes in diverse systems, but the advantages and limitations of different omics tools remain skeptical. Here, we reported distinct profiles in microbial secretory enzyme composition revealed by different omics methods. In general, the predicted function from metagenomic analysis decoupled from the expression of corresponding transcripts/proteins. Linking omics results to taxonomic origin, functional capability, substrate specificity, secretion preference, and enzymatic activity measurement suggested the substrate's source, concentration and stoichiometry impose strong filtering on the expression of extracellular enzymes, which may overwrite the genetic potentials. Our results present an integrated perspective on the need for multi-dimensional characterization of microbial adaptation in a changing environment.

Approach towards sustainable leather: Characterization and effective industrial application of proteases from Bacillus sps. for ecofriendly dehairing of leather hide.

Tanneries are one of the most polluted industries known for production of massive amount of solid and liquid wastes without proper management and disposal. In this project we demonstrated the ecofriendly single step dehairing of leather hides with minimum pollution load. In this study, Bacillus species (Bacillus paralicheniformis strain BL.HK, Bacillus cereus strain BS.P) capable of producing proteases was successfully isolated by employing the new optimized selective media named M9-PEA as confirmed by 16sRNA genes sequencing. Sequence of 1493 bp long 16S rRNA genes of Bacillus paralicheniformis strain BL.HK and Bacillus cereus strain BS. P was submitted to GenBank under the accession number OP612692.1, OP612721.1 respectively The Bacillus paralicheniformis strain BL.HK, Bacillus cereus strain BS.P produced extracellur proteases of 28 and 37 KDa as resolved by SDS-PAGE respectively. The enzymes showed temperature optima at 50 °C and 55 °C and pH optima at 8.5, 9.5 respectively. The Proteases of Bacillus paralicheniformis strain BL.HK, Bacillus cereus strain BS.P were employed for dehairing of animal hides. The process resulted in significant removal of interfibriller substances without damage to collagen layer after one hour treatment, which was confirmed by histology, scanning electron microscopy. The quantification of various skin constituents (collagen, uronic acid, hexosamines, and GAGs) and pollution load parameters revealed that enzymatic treatment are more reliable. The results of skin application trials at industrial level with complete elimination of chemicals remark the biotechnological potential of these proteases for ecofriendly dehairing of animal hides without affecting the quality of the leathers produced.

Plasmodium falciparum proteases as new drug targets with special focus on metalloproteases.

Malaria poses a significant global health threat particularly due to the prevalence of Plasmodium falciparum infection. With the emergence of parasite resistance to existing drugs including the recently discovered artemisinin, ongoing research seeks novel therapeutic avenues within the malaria parasite. Proteases are promising drug targets due to their essential roles in parasite biology, including hemoglobin digestion, merozoite invasion, and egress. While exploring the genomic landscape of Plasmodium falciparum, it has been revealed that there are 92 predicted proteases, with only approximately 14 of them having been characterized. These proteases are further distributed among 26 families grouped into five clans: aspartic proteases, cysteine proteases, metalloproteases, serine proteases, and threonine proteases. Focus on metalloprotease class shows further role in organelle processing for mitochondria and apicoplasts suggesting the potential of metalloproteases as viable drug targets. Holistic understanding of the parasite intricate life cycle and identification of potential drug targets are essential for developing effective therapeutic strategies against malaria and mitigating its devastating global impact.

Molecular strategies to enhance the keratinase gene expression and its potential implications in poultry feed industry.

The tons of keratin waste are produced by the poultry and meat industry which is an insoluble and protein-rich material found in hair, feathers, wool, and some epidermal wastes. These waste products could be degraded and recycled to recover protein, which can save our environment. One of the potential strategy to achieve this target is use of microbial biotreatment which is more convenient, cost-effective, and environment-friendly by formulating hydrolysate complexes that could be administered as protein supplements, bioactive peptides, or animal feed ingredients. Keratin degradation shows great promise for long-term protein and amino acid recycling. According to the MEROPS database, known keratinolytic enzymes currently belong to at least 14 different protease families, including S1, S8, S9, S10, S16, M3, M4, M14, M16, M28, M32, M36, M38, and M55. In addition to exogenous attack (proteases from families S9, S10, M14, M28, M38, and M55), the various keratinolytic enzymes also function via endo-attack (proteases from families S1, S8, S16, M4, M16, and M36). Biotechnological methods have shown great promise for enhancing keratinase expression in different strains of microbes and different protein engineering techniques in genetically modified microbes such as bacteria and some fungi to enhance keratinase production and activity. Some microbes produce specific keratinolytic enzymes that can effectively degrade keratin substrates. Keratinases have been successfully used in the leather, textile, and pharmaceutical industries. However, the production and efficiency of existing enzymes need to be optimized before they can be used more widely in other processes, such as the cost-effective pretreatment of chicken waste. These can be improved more effectively by using various biotechnological applications which could serve as the best and novel approach for recycling and degrading biomass. This paper provides practical insights about molecular strategies to enhance keratinase expression to effectively utilize various poultry wastes like feathers and feed ingredients like soybean pulp. Furthermore, it describes the future implications of engineered keratinases for environment friendly utilization of wastes and crop byproducts for their better use in the poultry feed industry.

An essential protease, FtsH, influences daptomycin resistance acquisition in Enterococcus faecalis.

Daptomycin is a last-line antibiotic commonly used to treat vancomycin-resistant Enterococci, but resistance evolves rapidly and further restricts already limited treatment options. While genetic determinants associated with clinical daptomycin resistance (DAPR) have been described, information on factors affecting the speed of DAPR acquisition is limited. The multiple peptide resistance factor (MprF), a phosphatidylglycerol-modifying enzyme involved in cationic antimicrobial resistance, is linked to DAPR in pathogens such as methicillin-resistant Staphylococcus aureus. Since Enterococcus faecalis encodes two paralogs of mprF and clinical DAPR mutations do not map to mprF, we hypothesized that functional redundancy between the paralogs prevents mprF-mediated resistance and masks other evolutionary pathways to DAPR. Here, we performed in vitro evolution to DAPR in mprF mutant background. We discovered that the absence of mprF results in slowed DAPR evolution and is associated with inactivating mutations in ftsH, resulting in the depletion of the chaperone repressor HrcA. We also report that ftsH is essential in the parental, but not in the ΔmprF, strain where FtsH depletion results in growth impairment in the parental strain, a phenotype associated with reduced extracellular acidification and reduced ability for metabolic reduction. This presents FtsH and HrcA as enticing targets for developing anti-resistance strategies.

Unravelling the Intricate Roles of FAM111A and FAM111B: From Protease-Mediated Cellular Processes to Disease Implications.

Proteases are critical enzymes in cellular processes which regulate intricate events like cellular proliferation, differentiation and apoptosis. This review highlights the multifaceted roles of the serine proteases FAM111A and FAM111B, exploring their impact on cellular functions and diseases. FAM111A is implicated in DNA replication and replication fork protection, thereby maintaining genome integrity. Additionally, FAM111A functions as an antiviral factor against DNA and RNA viruses. Apart from being involved in DNA repair, FAM111B, a paralog of FAM111A, participates in cell cycle regulation and apoptosis. It influences the apoptotic pathway by upregulating anti-apoptotic proteins and modulating cell cycle-related proteins. Furthermore, FAM111B's association with nucleoporins suggests its involvement in nucleo-cytoplasmic trafficking and plays a role in maintaining normal telomere length. FAM111A and FAM111B also exhibit some interconnectedness and functional similarity despite their distinct roles in cellular processes and associated diseases resulting from their dysfunction. FAM111A and FAM111B dysregulation are linked to genetic disorders: Kenny-Caffey Syndrome type 2 and Gracile Bone Dysplasia for FAM111A and POIKTMP, respectively, and cancers. Therefore, the dysregulation of these proteases in diseases emphasizes their potential as diagnostic markers and therapeutic targets. Future research is essential to unravel the intricate mechanisms governing FAM111A and FAM111B and explore their therapeutic implications comprehensively.

Revealing extracellular protein profile and excavating spoilage-related proteases of Aeromonas salmonicida based on multi-omics investigation.

Aeromonas is a ubiquitous aquatic bacteria, and it is a significant factor contributing to meat spoilage during processing and consumption. The abilities of Aeromonas salmonicida 29 and 57, which exhibit spoilage heterogeneity, to secrete protease, lipase, hemolysin, gelatinase, amylase, and lecithinase were confirmed by plate method. A total of 3948 proteins were identified by ITRAQ in extracellular secretions of A. salmonicida, and 16 proteases were found to be potentially related to spoilage ability. The complete genome sequence of A. salmonicida 57 consists of one circular chromosome and three plasmids, while A. salmonicida 29 consists of one circular chromosome, without a plasmid. Transcriptomic analysis revealed a significant number of DEGs were up-regulated in A. salmonicida 29, which were mainly enriched in metabolic pathways (e.g., amino acid metabolism, carbohydrate metabolism), indicating that A. salmonicida 29 had better potential to decompose and utilize nutrients in meat. Six protease genes (2 pepB, hap, pepA, ftsI, and pepD) were excavated by combined ITRAQ with transcriptome analysis, which potentially contribute to bacterial spoilage ability and exhibit universality among other dominant spoilage bacteria. This investigation provides new insights and evidence for elucidating metabolic and spoilage phenotypic differences and provides candidate genes and strategies for future prevention and control technology development.

Increased Proteolytic Activity of Serratia marcescens Clinical Isolate HU1848 Is Associated with Higher eepR Expression.

Serratia marcescens is a global opportunistic pathogen. In vitro cytotoxicity of this bacterium is mainly related to metalloprotease serralysin (PrtS) activity. Proteolytic capability varies among the different isolates. Here, we characterized protease production and transcriptional regulators at 37°C of two S. marcescens isolates from bronchial expectorations, HU1848 and SmUNAM836. As a reference strain the insect pathogen S. marcescens Db10 was included. Zymography of supernatant cultures revealed a single (SmUNAM836) or double proteolytic zones (HU1848 and Db10). Mass spectrometry confirmed the identity of PrtS and the serralysin-like protease SlpB from supernatant samples. Elevated proteolytic activity and prtS expression were evidenced in the HU1848 strain through azocasein degradation and qRT-PCR, respectively. Evaluation of transcriptional regulators revealed higher eepR expression in HU1848, whereas cpxR and hexS transcriptional levels were similar between studied strains. Higher eepR expression in HU1848 was further confirmed through an in vivo transcriptional assay. Moreover, two putative CpxR binding motifs were identified within the eepR regulatory region. EMSA validated the interaction of CpxR with both motifs. The evaluation of eepR transcription in a cpxR deletion strain indicated that CpxR negatively regulates eepR. Sequence conservation suggests that regulation of eepR by CpxR is common along S. marcescens species. Overall, our data incorporates CpxR to the complex regulatory mechanisms governing eepR expression and associates the increased proteolytic activity of the HU1848 strain with higher eepR transcription. Based on the global impact of EepR in secondary metabolites production, our work contributes to understanding virulence factors variances across S. marcescens isolates.

Oxidative Stress-Mediated Repression of Virulence Gene Transcription and Biofilm Formation as Antibacterial Action of Cinnamomum burmannii Essential Oil on Staphylococcus aureus.

This work aimed to identify the chemical compounds of Cinnamomum burmannii leaf essential oil (CBLEO) and to unravel the antibacterial mechanism of CBLEO at the molecular level for developing antimicrobials. CBLEO had 37 volatile compounds with abundant borneol (28.40%) and showed good potential to control foodborne pathogens, of which Staphylococcus aureus had the greatest inhibition zone diameter (28.72 mm) with the lowest values of minimum inhibitory concentration (1.0 µg/mL) and bactericidal concentration (2.0 µg/mL). To unravel the antibacterial action of CBLEO on S. aureus, a dynamic exploration of antibacterial growth, material leakage, ROS formation, protein oxidation, cell morphology, and interaction with genome DNA was conducted on S. aureus exposed to CBLEO at different doses (1/2-2×MIC) and times (0-24 h), indicating that CBLEO acts as an inducer for ROS production and the oxidative stress of S. aureus. To highlight the antibacterial action of CBLEO on S. aureus at the molecular level, we performed a comparative association of ROS accumulation with some key virulence-related gene (sigB/agrA/sarA/icaA/cidA/rsbU) transcription, protease production, and biofilm formation in S. aureus subjected to CBLEO at different levels and times, revealing that CBLEO-induced oxidative stress caused transcript suppression of virulence regulators (RsbU and SigB) and its targeted genes, causing a protease level increase destined for the biofilm formation and growth inhibition of S. aureus, which may be a key bactericidal action. Our findings provide valuable information for studying the antibacterial mechanism of essential oil against pathogens.

Engineering Anti-CRISPR Proteins to Create CRISPR-Cas Protein Switches for Activatable Genome Editing and Viral Protease Detection.

Proteins capable of switching between distinct active states in response to biochemical cues are ideal for sensing and controlling biological processes. Activatable CRISPR-Cas systems are significant in precise genetic manipulation and sensitive molecular diagnostics, yet directly controlling Cas protein function remains challenging. Herein, we explore anti-CRISPR (Acr) proteins as modules to create synthetic Cas protein switches (CasPSs) based on computational chemistry-directed rational protein interface engineering. Guided by molecular fingerprint analysis, electrostatic potential mapping, and binding free energy calculations, we rationally engineer the molecular interaction interface between Cas12a and its cognate Acr proteins (AcrVA4 and AcrVA5) to generate a series of orthogonal protease-responsive CasPSs. These CasPSs enable the conversion of specific proteolytic events into activation of Cas12a function with high switching ratios (up to 34.3-fold). These advancements enable specific proteolysis-inducible genome editing in mammalian cells and sensitive detection of viral protease activities during virus infection. This work provides a promising strategy for developing CRISPR-Cas tools for controllable gene manipulation and regulation and clinical diagnostics.

Unravelling HetC as a peptidase-based ABC exporter driving functional cell differentiation in the cyanobacterium Nostoc PCC 7120.

The export of peptides or proteins is essential for a variety of important functions in bacteria. Among the diverse protein-translocation systems, peptidase-containing ABC transporters (PCAT) are involved in the maturation and export of quorum-sensing or antimicrobial peptides in Gram-positive bacteria and of toxins in Gram-negative organisms. In the multicellular and diazotrophic cyanobacterium Nostoc PCC 7120, the protein HetC is essential for the differentiation of functional heterocysts, which are micro-oxic and non-dividing cells specialized in atmospheric nitrogen fixation. HetC shows similarities to PCAT systems, but whether it actually acts as a peptidase-based exporter remains to be established. In this study, we show that the N-terminal part of HetC, encompassing the peptidase domain, displays a cysteine-type protease activity. The conserved catalytic residues conserved in this family of proteases are essential for the proteolytic activity of HetC and the differentiation of heterocysts. Furthermore, we show that the catalytic residue of the ATPase domain of HetC is also essential for cell differentiation. Interestingly, HetC has a cyclic nucleotide-binding domain at its N-terminus which can bind ppGpp in vitro and which is required for its function in vivo. Our results indicate that HetC is a peculiar PCAT that might be regulated by ppGpp to potentially facilitate the export of a signaling peptide essential for cell differentiation, thereby broadening the scope of PCAT role in Gram-negative bacteria.IMPORTANCEBacteria have a great capacity to adapt to various environmental and physiological conditions; it is widely accepted that their ability to produce extracellular molecules contributes greatly to their fitness. Exported molecules are used for a variety of purposes ranging from communication to adjust cellular physiology, to the production of toxins that bacteria secrete to fight for their ecological niche. They use export machineries for this purpose, the most common of which energize transport by hydrolysis of adenosine triphosphate. Here, we demonstrate that such a mechanism is involved in cell differentiation in the filamentous cyanobacterium Nostoc PCC 7120. The HetC protein belongs to the ATP-binding cassette transporter superfamily and presumably ensures the maturation of a yet unknown substrate during export. These results open interesting perspectives on cellular signaling pathways involving the export of regulatory peptides, which will broaden our knowledge of how these bacteria use two cell types to conciliate photosynthesis and nitrogen fixation.

Variably protease-sensitive prionopathy with methionine homozygosity at codon 129 in the prion protein gene.

Variably protease-sensitive prionopathy (VPSPr) is a recently characterised rare subtype of sporadic prion disease, mainly affecting individuals with valine homozygosity at codon 129 in the prion protein gene, with only seven methionine homozygote cases reported to date. This case presents clinical, neuropathological and biochemical features of the eighth VPSPr case worldwide with methionine homozygosity at codon 129 and compares the features with the formerly presented cases.The patient, a woman in her 70s, presented with cognitive decline, impaired balance and frequent falls. Medical history and clinical presentation were suggestive of a rapidly progressive dementia disorder. MRI showed bilateral thalamic hyperintensity. Cerebrospinal fluid real-time quaking-induced conversion was negative, and the electroencephalogram was unremarkable. The diagnosis was established through post-mortem pathological examinations. VPSPr should be suspected in rapidly progressive dementia lacking typical features or paraclinical results of protein misfolding diseases.

The Deinococcus protease PprI senses DNA damage by directly interacting with single-stranded DNA.

Bacteria have evolved various response systems to adapt to environmental stress. A protease-based derepression mechanism in response to DNA damage was characterized in Deinococcus, which is controlled by the specific cleavage of repressor DdrO by metallopeptidase PprI (also called IrrE). Despite the efforts to document the biochemical, physiological, and downstream regulation of PprI-DdrO, the upstream regulatory signal activating this system remains unclear. Here, we show that single-stranded DNA physically interacts with PprI protease, which enhances the PprI-DdrO interactions as well as the DdrO cleavage in a length-dependent manner both in vivo and in vitro. Structures of PprI, in its apo and complexed forms with single-stranded DNA, reveal two DNA-binding interfaces shaping the cleavage site. Moreover, we show that the dynamic monomer-dimer equilibrium of PprI is also important for its cleavage activity. Our data provide evidence that single-stranded DNA could serve as the signal for DNA damage sensing in the metalloprotease/repressor system in bacteria. These results also shed light on the survival and acquired drug resistance of certain bacteria under antimicrobial stress through a SOS-independent pathway.

The association of TMPRSS6 gene polymorphism with iron status in Egyptian children (a pilot study).

Several studies have shown association of single nucleotide polymorphisms (SNPs) of hepcidin regulatory pathways genes with impaired iron status. The most common is in the TMPRSS6 gene. In Africa, very few studies have been reported. We aimed to investigate the correlation between the common SNPs in the transmembrane protease, serine 6 (TMPRSS6) gene and iron indicators in a sample of Egyptian children for identifying the suitable candidate for iron supplementation.Patients and methods One hundred and sixty children aged 5-13 years were included & classified into iron deficient, iron deficient anemia and normal healthy controls. All were subjected to assessment of serum iron, serum ferritin, total iron binding capacity, complete blood count, reticulocyte count, serum soluble transferrin receptor and serum hepcidin. Molecular study of TMPRSS6 genotyping polymorphisms (rs4820268, rs855791 and rs11704654) were also evaluated.Results There was an association of iron deficiency with AG of rs855791 SNP, (P = 0.01). The minor allele frequency for included children were 0.43, 0.45 & 0.17 for rs4820268, rs855791 & rs11704654 respectively. Genotype GG of rs4820268 expressed the highest hepcidin gene expression fold, the lowest serum ferroportin & iron store compared to AA and AG genotypes (p = 0.05, p = 0.05, p = 0.03 respectively). GG of rs855791 had lower serum ferritin than AA (p = 0.04), lowest iron store & highest serum hepcidin compared to AA and AG genotypes (p = 0.04, p = 0.01 respectively). Children having CC of rs11704654 had lower level of hemoglobin, serum ferritin and serum hepcidin compared with CT genotype (p = 0.01, p = 0.01, p = 0.02) respectively.Conclusion Possible contribution of SNPs (rs855791, rs4820268 and rs11704654) to low iron status.