CHARACTERIZATION OF BETA-DEFENSIN PREPROPEPTIDE MESSENGER-RNA FROM CHICKEN AND TURKEY BONE-MARROW

Four avian beta-defensin prepropeptide cDNA sequences [gallinacins: Ga l 1 (synonym CHP 1, chicken heterophil peptide 1), and Gal 2; turkey h eterophil peptides: THP 1 and THP 2] were amplified from chicken or tu rkey bone marrow mRNA samples, respectively. Partial chicken beta-defe nsin cDNA sequences were obtained using degenerate primers based on ch icken peptide sequences (Gal 1/CHP 1 and Gal 2). The complete cDNA seq uences of the chicken beta-defensins were then determined by designing specific intrapeptidal primers, from the newly acquired sequence, and pairing one primer with a specific poly A primer tail sequence (3' en d) and the other primer with an adapter primer in a 5' rapid amplifica tion of cDNA ends (RACE) reaction. The two, turkey beta-defensins were amplified from turkey marrow using primers designed from chicken beta -defensin preproregions. The complete amino acid sequences for the pre propeptides were deduced for all four avian beta-defensins. Previously , only partial mature peptide sequences for the turkey beta-defensins and complete mature peptide sequences for the chicken beta-defensins w ere known. All sequences obtained translated accurately to complete an d partial amino acid sequences reported for beta-defensins purified fr om chicken and turkey heterophil granules except for one additional am ino acid for Gal 1/CHP 1. The four deduced beta-defensin proregions la ck the long, negatively charged propiece reported in classical defensi n proregions. These regions are thought to stabilize and inactivate th e positively charged mature peptide and target the propeptide to the s torage granule. Instead, these beta-defensin proregions are shorter an d similar to storage granule-free beta-defensins proregions reported f or bovine tracheal antimicrobial peptide (TAP) and lingual antimicrobi al peptide (LAP). These are the first prepropeptide beta-defensins fro m leukocyte granules to be completely characterized.