Academic literature on the topic 'Pro-rich, defensins, antimicrobial peptides'

Common bed bugs, Cimex lectularius, can carry, but do not transmit, pathogens to the vertebrate hosts on which they feed. Some components of the innate immune system of bed bugs, such as antimicrobial peptides (AMPs), eliminate the pathogens. Here, we determined the molecular characteristics, structural properties, and phylogenetic relatedness of two new defensins (CL-defensin1 (XP_024085718.1), CL-defensin2 (XP_014240919.1)), and two new defensin isoforms (CL-defensin3a (XP_014240918.1), CL-defensin3b (XP_024083729.1)). The complete amino acid sequences of CL-defensin1, CL-defensin2, CL-defensin3a, and CL-defensin3b are strongly conserved, with only minor differences in their signal and pro-peptide regions. We used a combination of comparative transcriptomics and real-time quantitative PCR to evaluate the expression of these defensins in the midguts and the rest of the body of insects that had been injected with bacteria or had ingested blood containing the Gram-positive (Gr+) bacterium Bacillus subtilis and the Gram-negative (Gr–) bacterium Escherichia coli. We demonstrate, for the first time, sex-specific and immunization mode-specific upregulation of bed bug defensins in response to injection or ingestion of Gr+ or Gr– bacteria. Understanding the components, such as these defensins, of the bed bugs’ innate immune systems in response to pathogens may help unravel why bed bugs do not transmit pathogens to vertebrates.

Defensins have long been considered as merely antimicrobial peptides. Throughout the years, more immune-related functions have been discovered for both the α-defensin and β-defensin subfamily. This review provides insights into the role of defensins in tumor immunity. Since defensins are present and differentially expressed in certain cancer types, researchers started to unravel their role in the tumor microenvironment. The human neutrophil peptides have been demonstrated to be directly oncolytic by permealizing the cell membrane. Further, defensins can inflict DNA damage and induce apoptosis of tumor cells. In the tumor microenvironment, defensins can act as chemoattractants for subsets of immune cells, such as T cells, immature dendritic cells, monocytes and mast cells. Additionally, by activating the targeted leukocytes, defensins generate pro-inflammatory signals. Moreover, immuno-adjuvant effects have been reported in a variety of models. Therefore, the action of defensins reaches beyond their direct antimicrobial effect, i.e., the lysis of microbes invading the mucosal surfaces. By causing an increase in pro-inflammatory signaling events, cell lysis (generating antigens) and attraction and activation of antigen presenting cells, defensins could have a relevant role in activating the adaptive immune system and generating anti-tumor immunity, and could thus contribute to the success of immune therapy.

Plant defensins are small, cysteine-rich antimicrobial peptides. These peptides have previously been shown to primarily inhibit the growth of fungal plant pathogens. Plant defensins have a γ-core motif, defined as GXCX3-9C, which is required for their antifungal activity. To evaluate plant defensins as a potential control for a problematic agricultural disease (alfalfa crown rot), short, chemically synthesized peptides containing γ-core motif sequences were screened for activity against numerous crown rot pathogens. These peptides showed both antifungal and, surprisingly, antibacterial activity. Core motif peptides from Medicago truncatula defensins (MtDef4 and MtDef5) displayed high activity against both plant and human bacterial pathogens in vitro. Full-length defensins had higher antimicrobial activity compared with the peptides containing their predictive γ-core motifs. These results show the future promise for controlling a wide array of economically important fungal and bacterial plant pathogens through the transgenic expression of a plant defensin. They also suggest that plant defensins may be an untapped reservoir for development of therapeutic compounds for combating human and animal pathogens.

Beta-defensins consist in a group of cysteine-rich antimicrobial peptides (AMPs), widely found throughout vertebrate species, including teleost fish, with antimicrobial and immunomodulatory activities. However, although the European sea bass (Dicentrarchus labrax) is one of the most commercially important farmed fish species in the Mediterranean area, the characterization of its beta-defensins and its potential applications are still missing. In this study, we characterized two members of the beta-defensin family in this species. Phylogenetic and synteny analysis places sea bass peptides in the beta-defensin subfamilies 1 and 2, sharing similar features with the other members, including the six cysteines and the tertiary structure, that consists in three antiparallel beta-sheets, with beta-defensin 1 presenting an extra alpha-helix at the N-terminal. Further studies are necessary to uncover the functions of sea bass beta-defensins, particularly their antimicrobial and immunomodulatory properties, in order to develop novel prophylactic or therapeutic compounds to be used in aquaculture production.

Abstract Bin1b is a rat epididymis specific β-defensin which may have fertility related functions in addition to its antimicrobial activity. β-defensins are cysteine-rich cationic antimicrobial peptides that have their important implications in innate and adaptive immunity. Though considerable numbers of new β-defensins have been discovered, few corresponding antibodies have been reported. The small peptide with special structure and antimicrobial nature of β-defensins make them very difficult to express in prokaryotic system. Here we adopted a double-copy protein expression scheme based on which not only the mBin1b protein was successfully expressed but also the immunity of the antigen was enhanced. The validity of the antisera was verified by using Western blotting and immunohistochemical analyses. It will be a useful tool for deeply investigating the roles of Bin1b and also provide a simple but effective method in raising antisera against other members of the β-defensin gene family.

The purpose of the study was to analyze literature sources containing information about defensins, cysteine-rich cationic amphipathic peptides produced by circulating white blood cells and tissue cells. This review describes the antimicrobial, antiviral, anti-inflammatory and immunomodulatory properties of defensins, as well as their molecular and cellular interactions. These substances, which are present on the epithelium and body fluids, are active against bacteria, fungi and viruses, as well as produced by immune and epithelial cells. These natural antimicrobial cationic peptides play an important role in innate and adaptive immunity. Defensins are divided into alpha and beta families. Alpha-defensins (α-defensins) are found in neutrophils, macrophages and Paneth cells in the intestine. Beta-defensins (β-defensins) are secreted by most leukocytes and epithelial cells. Extensive antimicrobial activity and multifaceted immunomodulatory functions of defensins confirm their role in innate immunity as the main protective component of the human body against bacterial, viral and fungal infections. Thus, they are key effector molecules in protecting the organism from infection due to their broad-spectrum antimicrobial activity. Their common antimicrobial function is the formation of destructive pores in the membranes of pathogens, including enveloped viruses. Antiviral activity includes the direct effect of defensin on viral envelopes, glycoproteins and capsids. Binding and modulation of host cell surface receptors and disruption of intracellular signaling by defensins may also inhibit virus replication. These peptides block infection with enveloped and non-enveloped viruses by aggregating particles, blocking receptor binding, inhibiting virus penetration or depletion of particles, inhibiting stem cell signaling, or viral gene expression. In addition, defensins may function as chemokines to enhance and alter adaptive immune responses by exhibiting an indirect antiviral mechanism. Conclusion. However, sources of scientific information have shown that defensins attract immune cells and modulate adaptive immune responses. It has also been shown that defensins can both induce inflammation and suppress inflammatory responses by acting on certain cells through various mechanisms. Due to this, they can be used as one of the markers in the development of inflammatory diseases of the mouth and oropharynx. The main drugs that activate the production of defensins are probiotics, vitamin D and leukotriene B4. This expands the possibility of their use as a new class of non-toxic antimicrobials and immunomodulators

Plant defensins are promising future strategy as antimicrobial agents; however the number of characterized plant defensins is very low. We isolated and studied the expression of plant defensins in six plants from Taif region. Using RT-PCR and two pairs of common as well as three pairs of specific primers, the defensin gene expression was analyzed. Plants showed differences in defensin gene expression in floral buds and leaves, though floral buds represented higher gene expression. The amino acid sequence of the six isolated cDNA sequences showed high similarity with other defensin accessions from the nucleotide database, especially in the cysteine rich motif. Phylogenetic analysis revealed that the isolated sequences share the common features of plant defensins, especially the 8 conserved cysteines. The results of this study help to detect some valuable Saudi plant sources for the development of natural peptides as a replacement for chemical antibiotics.

Abstract Defensins constitute a family of 3- to 4-kDa antimicrobial peptides that are stored in the cytoplasmic granules of neutrophils, some macrophages, and intestinal Paneth cells. We have assessed defensin gene expression during myeloid differentiation by first characterizing cDNAs for each of the four known rat neutrophil defensins (RatNP 1-4). The cDNA sequences revealed that the peptides are synthesized as 87-94 amino acid precursors, each containing signal, pro-, and mature peptide segments. RatNP-3 and -4 mRNAs, but not those for RatNP-1 and -2 or other myeloid defensins, contained unique polypurine tracts located close to the termination codon in the 3' untranslated region. By using cDNA probes and/or riboprobes, we evaluated defensin transcript levels in a variety of tissues and in the full spectrum of neutrophil precursors. By in situ hybridization, defensin mRNAs were localized to neutrophil precursors in the bone marrow, with the highest mRNA levels occurring in promyelocytes and somewhat lower signals occurring in myeloblasts and myelocytes. Defensin mRNAs were not detectable in bands or mature neutrophils, nor at significant levels in tissues other than bone marrow. The accumulation of defensin protein in bone marrow cells was assessed by immunohistochemical staining with anti-RatNP-1 Ab. RatNP 1-4 mRNAs and protein levels were then correlated for each stage of neutrophilic differentiation to reveal the maturational profile of myeloid defensin gene expression in the rat.

Defensins are cysteine-rich cationic antimicrobial peptides that play an important role in innate immunity and are known to contribute to the regulation of host adaptive immunity. In addition to direct antimicrobial activities, it has been recently reported that α-defensins, mainly present in neutrophils in the lung, have a cytotoxic effect and induce IL-8 production from airway epithelial cells. Although β-defensins are expressed in epithelial cells in various tissues, including lung, there are no reports of their effects on cytokine synthesis in airway epithelial cells. The aim of the present study was to determine the effects of both α- and β-defensins on the cytokine production, transcription factor binding activity, and cytotoxicity in primary cultured human bronchial epithelial cells (HBECs). We used human neutrophil peptide-1 (HNP-1; α-defensin) and human β-defensin-2 (HBD-2) to stimulate HBECs. The results showed that treatment of HBECs with HNP-1, but not HBD-2, increased IL-8 and IL-1β mRNA expression in a dose-dependent manner and also enhanced IL-8 protein secretion and NF-κB DNA binding activity. The 24-h treatments with >20 μg/ml of HNP-1 or >50 μg/ml of HBD-2 were cytotoxic to HBECs. These results suggest that α- and β-defensins have different effects on cytokine synthesis by airway epithelial cells, and we speculate that they play different roles in inflammatory lung diseases.

Traditional medicinal plants contain a variety of bioactive natural products including cysteine-rich (Cys-rich) antimicrobial peptides (AMPs). Cys-rich AMPs are often crosslinked by multiple disulfide bonds which increase their resistance to chemical and enzymatic degradation. However, this class of molecules is relatively underexplored. Herein, in silico analysis predicted 80–100 Cys-rich AMPs per species from three edible traditional medicinal plants: Linum usitatissimum (flax), Trifolium pratense (red clover), and Sesamum indicum (sesame). Bottom-up proteomic analysis of seed peptide extracts revealed direct evidence for the translation of 3–10 Cys-rich AMPs per species, including lipid transfer proteins, defensins, α-hairpinins, and snakins. Negative activity revealed by antibacterial screening highlights the importance of employing a multi-pronged approach for AMP discovery. Further, this study demonstrates that flax, red clover, and sesame are promising sources for further AMP discovery and characterization.

2010/2011
Antimicrobial peptides (AMPs) are an important group of innate immunity effectors, needed to prevent or arrest microbial infections. In this thesis I describe an investigation on the modes of action and structure activity relationships of different types of AMPs, in particular the proline-rich bactenecins and β-defensins. The sequence of the bovine cathelicidin Bac7, a reference Pro-rich AMP, has suggested several previous studies identifying the N-terminal region as responsible for the antimicrobial activity. To search for the minimal entry sequence into bacteria, and to investigate whether this overlaps with the minimal antimicrobial fragment a set of progressively shortened labelled N-terminal fragments of Bac7 were synthesized and tested for their antibacterial activity and internalization capacity into E. coli cells by flow cytometry and confocal microscopy. I confirmed the precise 16-residue fragment which is still fully active and is efficiently internalized into cells. Further shortening leads to a dramatic decrease of both functions. Furthermore it was found that a continuous chain is required for transmembrane transport and/or antimicrobial activity. In addition I have worked to determine the role of the two key N-terminal Arg residues of Bac7 on penetration of the outer membrane and translocation through a putative inner membrane transporter i) synthesizing Bac7 analogues with systematically altered physico-chemical properties of first two Arg residues, ii) using mutants of E.coli strains with deleted transport system or altered outer membrane characteristics iii) testing their differential potencies and internalization efficiency by flow cytometric assays and biological assay. The results indicated that stereochemistry, charge and H-bonding all seem to be important requirements for the activity and internalization of this Pro-rich AMPs. These are relevant to both OM transit and the translocating role of inner membrane transporter, which was confirmed. These studies have helped evaluate Bac7 as a potential anti-infective agent selective for Gram-negative bacteria, as well as a possible vehicle for internalization of antibiotic cargo into these. β-Defensins of various origins all show six highly conserved cysteines that form three S-S bridges to stabilize a β-sheet structure, suggesting a strong structure/activity relationship in their mode of action. Starting from the active and multimeric human β-defensin hBD3, I have synthesized a monomeric analogue that contains only the N-terminal α-helical stretch and one C-terminal strand from this AMP, removing most of the β-sheet core. Its activity was compared with that of hBD3 and a monomeric full sequence primate variant hcBD3. The antimicrobial activity of brevi-hBD3 was found to be comparable in potency to that of hBD3, but with significantly increased bacterial permeabilization capacity and decreased haemolysis with respect to hBD3. It appears to have switched to a different killing mechanism. It helped define the role of the long, amphipathic helical segment at the N-terminus of hBD3, a member of the -defensins which has a robust antimicrobial activity and is therefore considered a potential lead for novel anti-infective agents. To study the interaction between antimicrobial peptides of interest to me and model membranes I have used Surface Plasmon Resonance. I observed a disparate capacity of AMPs to bind with membranes that can depend on very limited alterations in the peptides and have confirmed SPR as a valuable tool for understanding the mode-of-action of AMPs. Throughout by PhD project, I have also actively collaborated in several studies aimed at understanding the modes of antimicrobial action and potential of different types of AMPs. In collaboration with groups at the Universities of Chieti, Udine, and Split, I have had the opportunity to study the roles of these AMPs against pathogens of cystic fibrosis or against the pathogenic alga Prototheca, causing bovine mastitis, and contributed to the identification and characterization of novel AMPs from teleosts and anurans. As these studies have resulted in publications of which I am co-author, I have included these with a brief description of methods used and the principal results.
XXIV Ciclo
1980