Ovodefensins, an Oviduct-Specific Antimicrobial Gene Family, Have Evolved in Birds and Reptiles to Protect the Egg by Both Sequence and Intra-Six-Cysteine Sequence Motif Spacing1

Abstract

Ovodefensins are a novel beta defensin related family of antimicrobial peptides containing conserved glycine and six cysteine residues. Originally thought to be restricted to the albumen producing region of the avian oviduct, expression was found in chicken, turkey, duck and zebra finch in large quantities in many parts of the oviduct, but this varied between species and between gene forms in the same species. Using new search strategies the ovodefensin family now has 35 members including reptiles, but no representatives outside birds and reptiles have been found. Analysis of their evolution shows that ovodefensins divide into 6 groups based on the intra cysteine amino acid spacing, representing a unique mechanism alongside traditional evolution of sequence. The groups have been used to base a nomenclature for the family. Antimicrobial activity for three ovodefensins from chicken and duck was confirmed against E. coli and a pathogenic E. coli strain as well as a gram +ve organism, S. aureus, for the first time. However, activity varied greatly between peptides, with Gallus gallus OvoDA1 being the most potent, suggesting a link with the different structures. Expression of Gallus gallus OvoDA1 (gallin) in the oviduct was increased by oestrogen and progesterone and in the reproductive state. Overall the results support the hypothesis that ovodefensins evolved to protect the egg but they are not necessarily restricted to the egg white. There divergent motif structure and sequence present an interesting area of research for antimicrobial peptide design and understanding protection of the cleidoic egg.Summary sentence: Bird and reptile oviduct specific antimicrobial gene family, the ovodefensins, have evolved both their sequence and the amino acid spacing between a conserved cysteine sequence motif.

The Komodo dragon (Varanus komodoensis) genome and identification of innate immunity genes and clusters

“…Reptile neutrophils appear to have granules that contain both cathelicidin-like peptides as well as β-defensin peptides. β-defensin-like peptides are also found in reptile eggs [26]. It is well-known that some species of lizard can lose their tails as a method of predator escape, and that these tails then regenerate from the wound site without inflammation or infection.…”

“…The family of vertebrate defensin peptides includes alpha-, beta-, theta- and ovo-defensin subclasses, with alpha- and theta-defensins being unique to mammals and ovodefensins to birds and reptiles [26, 27]. Peptides in each subclass exhibit compact three-dimensional conformations stabilized by characteristic conserved patterns of cysteine residues and associated disulfide bond networks.…”

Gastrointestinal distribution of chicken gastrin-cholecystokinin family transcript expression and response to short-term nutritive state

“…Brilliant III Ultra-fast SYBR Green qPCR Mastermix and the Mx3005p qPCR System with MxPro software (Agilent Technologies) were employed according to the manufacturers’ guidelines and as described previously ( Whenham et al, 2015 ). Briefly, 10 μl SYBR mix, 8 μl cDNA product, 0.4 μl 20 μM forward primer, 0.4 μl 20 μM reverse primer, 0.3 μl 1/500 rox reference dye solution and 0.9 μl H 2 O were mixed for each 20 μl reaction.…”

Pancreatic PYY but not PPY expression is responsive to short-term nutritional state and the pancreas constitutes the major site of PYY mRNA expression in chickens

“…All qPCR assays employed Brilliant III Ultra-fast SYBR Green qPCR Mastermix and the Mx3005p qPCR System with MxPro software (Agilent Technologies) according to the manufacturers’ guidelines and as described previously ( Whenham et al, 2015 ). Briefly, 10 μl SYBR mix, 8 μl cDNA product, 0.4 μl 20 μM forward primer, 0.4 μl 20 μM reverse primer, 0.3 μl 1/500 rox reference dye solution and 0.9 μl H 2 O were mixed for each 20 μl reaction.…”