-
The expression of human alpha and beta defensin in the endometrium and their effect on implantation.Journal of Assisted Reproduction and... Nov 2007Alpha and beta defensins have been isolated from various human tissues and form an important part of the innate immune system. Their role in implantation of the embryo...
BACKGROUND
Alpha and beta defensins have been isolated from various human tissues and form an important part of the innate immune system. Their role in implantation of the embryo has not yet been studied. This study was designed to detect both alpha and beta defensins in the mid luteal phase endometrium and investigate the correlation between the defensin expression and implantation of the embryo.
METHOD AND RESULTS
An experimental study was designed to detect alpha defensin (HNP1-3) and beta defensin (HBD1) in midluteal phase endometrial samples obtained from women attending the IVF unit at the Liverpool Women's Hospital, UK. Samples were obtained at least two menstrual cycles before IVF treatment was commenced. Immunohistochemical staining was conducted to estimate defensin expression. Some endometrial stromal cells stained positive for HNP1-3 during the midluteal phase. HNP1-3 expression is significantly higher in cases presenting with female factor infertility as compared with purely male factor infertility. A significant increase was not observed in tubal factor or endometriosis when considered separately. Endometrial stromal neutrophils were shown to be the main source of endometrial HNP1-3. HBD1 was the only beta defensin detected by immunochemical staining in the midluteal phase endometrium. The intensity of staining was significantly different in the endometrial stroma, luminal and glandular epithelia. HBD1 expression is not significantly higher in female factor infertility.
CONCLUSION
The study confirmed secretion of HNP1-3 by endometrial stromal neutrophils. Glandular epithelium is the main source of HBD1 expression in the human endometrium. HNP1-3 shows increased expression in female factor infertility. HBD1 expression is not higher in female factor infertility. These defensins do not appear to influence implantation.
Topics: Adult; Embryo Implantation; Endometrium; Female; Gene Expression; Humans; Immunohistochemistry; Infertility, Female; Infertility, Male; Male; alpha-Defensins; beta-Defensins
PubMed: 18026831
DOI: 10.1007/s10815-007-9173-2 -
BMC Biochemistry Apr 2014Defensins are basic, cysteine-rich antimicrobial peptides that are important components of plant defense against pathogens. Previously, we isolated a defensin, PvD1,...
BACKGROUND
Defensins are basic, cysteine-rich antimicrobial peptides that are important components of plant defense against pathogens. Previously, we isolated a defensin, PvD1, from Phaseolus vulgaris L. (common bean) seeds.
RESULTS
The aim of this study was to overexpress PvD1 in a prokaryotic system, verify the biologic function of recombinant PvD1 (PvD1r) by comparing the antimicrobial activity of PvD1r to that of the natural defensin, PvD1, and use a mutant Candida albicans strain that lacks the gene for sphingolipid biosynthesis to unravel the target site of the PvD1r in C. albicans cells. The cDNA encoding PvD1, which was previously obtained, was cloned into the pET-32 EK/LIC vector, and the resulting construct was used to transform bacterial cells (Rosetta Gami 2 (DE3) pLysS) leading to recombinant protein expression. After expression had been induced, PvD1r was purified, cleaved with enterokinase and repurified by chromatographic steps. N-terminal amino acid sequencing showed that the overall process of the recombinant production of PvD1r, including cleavage with the enterokinase, was successful. Additionally, modeling revealed that PvD1r had a structure that was similar to the defensin isolated from plants. Purified PvD1 and PvD1r possessed inhibitory activity against the growth of the wild-type pathogenic yeast strain C. albicans. Both defensins, however, did not present inhibitory activity against the mutant strain of C. albicans. Antifungal assays with the wild-type C. albicans strains showed morphological changes upon observation by light microscopy following growth assays. PvD1r was coupled to FITC, and the subsequent treatment of wild type C. albicans with DAPI revealed that the labeled peptide was intracellularly localized. In the mutant strain, no intracellular labeling was detected.
CONCLUSION
Our results indicate that PvD1r retains full biological activity after recombinant production, enterokinase cleavage and purification. Additionally, our results from the antimicrobial assay, the microscopic analysis and the PvD1r-FITC labeling assays corroborate each other and lead us to suggest that the target of PvD1 in C. albicans cells is the sphingolipid glucosylceramide.
Topics: Antifungal Agents; Base Sequence; Candida albicans; Cloning, Molecular; Defensins; Gene Expression; Molecular Sequence Data; Phaseolus; Protein Structure, Tertiary; Recombinant Proteins; Seeds
PubMed: 24690228
DOI: 10.1186/1471-2091-15-7 -
Cellular and Molecular Life Sciences :... Jul 2011Primate theta-defensins are physically distinguished as the only known fully-cyclic peptides of animal origin. Humans do not produce theta-defensin peptides due to a... (Review)
Review
Primate theta-defensins are physically distinguished as the only known fully-cyclic peptides of animal origin. Humans do not produce theta-defensin peptides due to a premature stop codon present in the signal sequence of all six theta-defensin pseudogenes. Instead, since the putative coding regions of human theta-defensin pseudogenes have remained remarkably intact, their corresponding peptides, called "retrocyclins", have been recreated using solid-phase synthetic approaches. Retrocyclins exhibit an exceptional therapeutic index both as inhibitors of HIV-1 entry and as bactericidal agents, which makes retrocyclins promising candidates for further development as topical microbicides to prevent sexually transmitted diseases. This review presents the evolution, antiretroviral mechanism of action, and potential clinical applications of retrocyclins to prevent sexual transmission of HIV-1.
Topics: Defensins; HIV Infections; HIV-1; Humans; Virus Internalization
PubMed: 21553001
DOI: 10.1007/s00018-011-0715-5 -
Plant, Cell & Environment Sep 2019Although excess cadmium (Cd) accumulation is harmful to plants, the molecular mechanisms underlying Cd detoxification and accumulation in Arabidopsis thaliana remain...
Although excess cadmium (Cd) accumulation is harmful to plants, the molecular mechanisms underlying Cd detoxification and accumulation in Arabidopsis thaliana remain largely undetermined. In this study, we demonstrated that the A. thaliana PLANT DEFENSIN 2 gene AtPDF2.5 is involved in Cd tolerance and accumulation. In vitro Cd-binding assays revealed that AtPDF2.5 has Cd-chelating activity. Site-directed mutagenesis of AtPDF2.5 identified eight cysteine residues that were essential for mediating Cd tolerance and chelation. Histochemical analysis demonstrated that AtPDF2.5 was mainly expressed in root xylem vascular bundles, and that AtPDF2.5 was significantly induced by Cd. Subcellular localization analysis revealed that AtPDF2.5 was localized to the cell wall. The overexpression of AtPDF2.5 significantly enhanced Cd tolerance and accumulation in A. thaliana and its heterologous overexpression in rice increased Cd accumulation; however, the functional disruption of AtPDF2.5 decreased Cd tolerance and accumulation. Physiological analysis suggested that AtPDF2.5 promoted Cd efflux from the protoplast and its subsequent accumulation in the cell wall. These data suggest that AtPDF2.5 promotes cytoplasmic Cd efflux via chelation, thereby enhancing Cd detoxification and apoplastic accumulation.
Topics: Arabidopsis; Cadmium; Cell Wall; Defensins; Plant Roots; Plant Vascular Bundle; Protein Sorting Signals
PubMed: 31115921
DOI: 10.1111/pce.13592 -
International Journal of Molecular... Oct 2022Aphids (Hemiptera: Aphidoidea) are among the most detrimental insects for agricultural plants, and their management is a great challenge in agronomical research. A new...
Aphids (Hemiptera: Aphidoidea) are among the most detrimental insects for agricultural plants, and their management is a great challenge in agronomical research. A new class of proteins, called Bacteriocyte-specific Cysteine-Rich (BCR) peptides, provides an alternative to chemical insecticides for pest control. BCRs were initially identified in the pea aphid . They are small disulfide bond-rich proteins expressed exclusively in aphid bacteriocytes, the insect cells that host intracellular symbiotic bacteria. Here, we show that one of the BCRs, BCR4, displays prominent insecticidal activity against the pea aphid, impairing insect survival and nymphal growth, providing evidence for its potential use as a new biopesticide. Our comparative genomics and phylogenetic analyses indicate that BCRs are restricted to the aphid lineage. The 3D structure of BCR4 reveals that this peptide belongs to an as-yet-unknown structural class of peptides and defines a new superfamily of defensins.
Topics: Animals; Aphids; Phylogeny; Insecticides; Cysteine; Biological Control Agents; Symbiosis; Peptides; Disulfides; Defensins
PubMed: 36293341
DOI: 10.3390/ijms232012480 -
Scientific Reports Nov 2022β-defensins are adsorbable on the sperm surface in the male reproductive tract (MRT) and enhance sperm functional characteristics. The beta-defensin 129 (DEFB129)...
β-defensins are adsorbable on the sperm surface in the male reproductive tract (MRT) and enhance sperm functional characteristics. The beta-defensin 129 (DEFB129) antimicrobial peptide is involved in sperm maturation, motility, and fertilization. However, its role in bovine fertility has not been well investigated. This study examines the relationship between the bovine BBD129 gene and Bos indicus x Bos taurus bull fertility. The complete coding sequence of BBD129 mRNA was identified by RNA Ligase Mediated-Rapid Amplification of cDNA End (RLM-RACE) and Sanger sequencing methodologies. It consisted of 582 nucleotides (nts) including 5' untranslated region (UTR) (46nts) and 3'UTR (23nts). It conserves all beta-defensin-like features. The expression level of BBD129 was checked by RT-qPCR and maximal expression was detected in the corpus-epididymis region compared to other parts of MRT. Polymorphism in BBD129 was also confirmed by Sanger sequencing of 254 clones from 5 high fertile (HF) and 6 low fertile (LF) bulls at two positions, 169 T > G and 329A > G, which change the S57A and N110S in the protein sequence respectively. These two mutations give rise to four types of BBD129 haplotypes. The non-mutated TA-BBD129 (169 T/329A) haplotype was substantially more prevalent among high-fertile bulls (P < 0.005), while the double-site mutated GG-BBD129 (169 T > G/329A > G) haplotype was significantly more prevalent among low-fertile bulls (P < 0.005). The in silico analysis confirmed that the polymorphism in BBD129 results in changes in mRNA secondary structure, protein conformations, protein stability, extracellular-surface availability, post-translational modifications (O-glycosylation and phosphorylation), and affects antibacterial and immunomodulatory capabilities. In conclusion, the mRNA expression of BBD129 in the MRT indicates its region-specific dynamics in sperm maturation. BBD129 polymorphisms were identified as the deciding elements accountable for the changed proteins with impaired functionality, contributing to cross-bred bulls' poor fertility.
Topics: Cattle; Male; Animals; beta-Defensins; Semen; Fertility; Spermatozoa; 3' Untranslated Regions
PubMed: 36352091
DOI: 10.1038/s41598-022-23654-3 -
The Journal of Biological Chemistry Mar 2009Proteolytic processing of defensins is a critical mode of posttranslational regulation of peptide activity. Because mouse alpha-defensin precursors are cleaved and...
Proteolytic processing of defensins is a critical mode of posttranslational regulation of peptide activity. Because mouse alpha-defensin precursors are cleaved and activated by matrix metalloproteinase-7 (MMP-7), we determined if additional defensin molecules, namely human neutrophil defensin pro-HNP-1 and beta-defensins, are targets for MMP-7. We found that MMP-7 cleaves within the pro-domain of the HNP-1 precursor, a reaction that does not generate the mature peptide but produces a 59-amino acid intermediate. This intermediate, which retains the carboxyl-terminal end of the pro-domain, had antimicrobial activity, indicating that the residues important for masking defensin activity reside in the amino terminus of this domain. Mature HNP-1 was resistant to processing by MMP-7 unless the peptide was reduced and alkylated, demonstrating that only the pro-domain of alpha-defensins is normally accessible for cleavage by this enzyme. From the 47-residue HBD-1 precursor, MMP-7 catalyzed removal of 6 amino acids from the amino terminus. Neither a 39-residue intermediate form of HBD-1 nor the mature 36-residue form of HBD-1 was cleaved by MMP-7. In addition, both pro-HBD-2, with its shorter amino-terminal extension, and pro-HBD-3 were resistant to MMP-7. However, human and mouse beta-defensin precursors that lack disulfide bonding contain a cryptic MMP-7-sensitive site within the mature peptide moiety. These findings support and extend accumulating evidence that the native three-dimensional structure of both alpha- and beta-defensins protects the mature peptides against proteolytic processing by MMP-7. We also conclude that sites for MMP-7 cleavage are more common at the amino termini of alpha-defensin rather than beta-defensin precursors, and that catalysis at these sites in alpha-defensin pro-domains results in acquisition of defensin activity.
Topics: Animals; Catalysis; Cell Line; Humans; Matrix Metalloproteinase 7; Mice; Protein Precursors; Protein Structure, Tertiary; alpha-Defensins; beta-Defensins
PubMed: 19181662
DOI: 10.1074/jbc.M809744200 -
Veterinary Research Jun 2021Mannheimia haemolytica-induced bovine respiratory disease causes loss of millions of dollars to Canadian cattle industry. Current antimicrobials are proving to be...
Mannheimia haemolytica-induced bovine respiratory disease causes loss of millions of dollars to Canadian cattle industry. Current antimicrobials are proving to be ineffective and leave residues in meat. Antimicrobial peptides (AMPs) may be effective against M. haemolytica while minimizing the risk of drug residues. Cationic AMPs can kill bacteria through interactions with the anionic bacterial membrane. Human β-Defensin 3 (HBD3) and microcin J25 (MccJ25) are AMPs with potent activity against many Gram-negative bacteria. We tested the microbicidal activity of wild-type HBD3, three HBD3 peptide analogues (28 amino acid, 20AA, and 10AA) derived from the sequence of natural HBD3, and MccJ25 in vitro against M. haemolytica. Three C-terminal analogues of HBD3 with all cysteines replaced with valines were manually synthesized using solid phase peptide synthesis. Since AMPs can act as chemoattractant we tested the chemotactic effect of HBD3, 28AA, 20AA, and 10AA peptides on bovine neutrophils in Boyden chamber. Minimum bactericidal concentration (MBC) assay showed that M. haemolytica was intermediately sensitive to HBD3, 28AA and 20AA analogues with an MBC of 50 µg/mL. The 10AA analogue had MBC 6.3 µg/mL which is likely a result of lower final inoculum size. MccJ25 didn't have significant bactericidal effect below an MBC < 100 µg/mL. Bovine neutrophils showed chemotaxis towards HBD3 and 20AA peptides (P < 0.05) but not towards 28AA analogue. Co-incubation of neutrophils with any of the peptides did not affect their chemotaxis towards N-formyl-L-methionyl-L-leucyl-phenylalanine (fMLP). The data show that these peptides are effective against M. haemolytica and are chemotactic for neutrophils in vitro.
Topics: Animals; Bacteriocins; Cattle; Mannheimia haemolytica; Neutrophils; Protein Engineering; beta-Defensins
PubMed: 34112244
DOI: 10.1186/s13567-021-00956-4 -
Toxins Dec 2021β-defensins are antimicrobial peptides presenting in vertebrate animals. They participate in innate immunity, but little is known about them in reptiles, including...
β-defensins are antimicrobial peptides presenting in vertebrate animals. They participate in innate immunity, but little is known about them in reptiles, including snakes. Although several β-defensin genes were described in Brazilian snakes, their function is still unknown. The peptide sequence from these genes was deduced, and synthetic peptides (with approximately 40 amino acids and derived peptides) were tested against pathogenic bacteria and fungi using microbroth dilution assays. The linear peptides, derived from β-defensins, were designed applying the bioisosterism strategy. The linear β-defensins were more active against , , , and . The derived peptides (7-14 mer) showed antibacterial activity against those bacteria and on . Nonetheless, they did not present activity against , , , and showing that the cysteine substitution to serine is deleterious to antifungal properties. Tryptophan residue showed to be necessary to improve antibacterial activity. Even though the studied snake β-defensins do not have high antimicrobial activity, they proved to be attractive as template molecules for the development of antibiotics.
Topics: Animals; Anti-Infective Agents; Bacteria; Fungi; Reptilian Proteins; Snakes; Species Specificity; beta-Defensins
PubMed: 35050978
DOI: 10.3390/toxins14010001 -
Scientific Reports Aug 2018β-Defensins are small antimicrobial proteins expressed in various organisms and have great potential for improving animal health and selective breeding programs. Giant...
β-Defensins are small antimicrobial proteins expressed in various organisms and have great potential for improving animal health and selective breeding programs. Giant pandas have a distinctive lineage in Carnivora, and it is unclear whether β-defensin genes have experienced different selective pressures during giant panda evolution. We therefore characterized the giant panda (Ailuropoda melanoleuca) β-defensin gene family through gap filling, TBLASTN, and HMM searches. Among 36 β-defensins identified, gastrointestinal disease may induce the expression of the DEFB1 and DEFB139 genes in the digestive system. Moreover, for DEFB139, a significant positive selection different from that of its homologs was revealed through branch model comparisons. A Pro-to-Arg mutation in the giant panda DEFB139 mature peptide may have enhanced the peptide's antimicrobial potency by increasing its stability, isoelectric point, surface charge and surface hydrophobicity, and by stabilizing its second β-sheet. Broth microdilution tests showed that the increase in net charge caused by the Pro-to-Arg mutation has enhanced the peptide's potency against Staphylococcus aureus, although the increase was minor. We expect that additional gene function and expression studies of the giant panda DEFB139 gene could improve the existing conservation strategies for the giant panda.
Topics: Amino Acid Sequence; Animals; Anti-Infective Agents; Isoelectric Point; Mutation; Phylogeny; Ursidae; beta-Defensins
PubMed: 30120296
DOI: 10.1038/s41598-018-29898-2