MOUSE PANETH CELL DEFENSINS - PRIMARY STRUCTURES AND ANTIBACTERIAL ACTIVITIES OF NUMEROUS CRYPTDIN ISOFORMS

Cryptdins are antimicrobial peptides of the defensin family that are p roduced by intestinal Paneth cells. mRNAs encoding 17 cryptdin isoform s have been characterized from a cDNA library generated from a single jejunal crypt. Six cryptdin cDNAs correspond to known peptides, and th e remainder predict 11 novel Paneth cell defensins. Most cryptdin cDNA s have greater than or equal to 93% nucleotide sequence identity overa ll, except for cryptdin 4 and 5 cDNAs, whose respective mature peptide encoding regions are only 74 and 78% identical to that of cryptdin 1. Cryptdin cDNAs differ at a small number of nucleotide positions: freq uent substitutions were found in codons 38 and 52 of the propiece and in codons 68, 73, 76, 87, and 89 of the deduced peptides; cDNA clones with changes in codons 74, 83, and 88 were found, but there were fewer of these. The antimicrobial activities of cryptdins 1 to 6 were teste d against Escherichia call ML35 in two assays. In an agar diffusion as say, the potencies of cryptdins 1 to 3, 5, and 6 were approximately eq uivalent to that of rabbit neutrophil defensin NP-1 but cryptdin 4 was 30 times more active than NP-1. In a bactericidal assay system, crypt dins 1 and 3 to 6 were equally active at 10 mu g/ml but cryptdin 2 and rabbit NP-1 were not active at this concentration. Since cryptdins 2 and 3 differ only ah residue 10 (Thr and Lys, respectively), this amin o acid appears to function in bactericidal interaction with E. coli. T he demonstration that Paneth cells express a diverse population of mic robicidal defensins further implicates cryptdins in restricting coloni zation or invasion of small intestinal epithelium by bacteria.