RAT NEUTROPHIL DEFENSINS - PRECURSOR STRUCTURES AND EXPRESSION DURINGNEUTROPHILIC MYELOPOIESIS

Defensins constitute a family of 3- to 4-kDa antimicrobial peptides th at are stored in the cytoplasmic granules of neutrophils, some macroph ages, and intestinal Paneth cells, We have assessed defensin gene expr ession during myeloid differentiation by first characterizing cDNAs fo r each of the four known rat neutrophil defensins (RatNP 1-4). The cDN A sequences revealed that the peptides are synthesized as 87-94 amino acid precursors, each containing signal, pro-, and mature peptide segm ents, 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 pr obes and/or riboprobes, we evaluated defensin transcript levels in a v ariety of tissues and in the full spectrum of neutrophil precursors, B y in situ hybridization, defensin mRNAs were localized to neutrophil p recursors in the bone marrow, with the highest mRNA levels occurring i n promyelocytes and somewhat lower signals occurring in myeloblasts an d myelocytes. Defensin mRNAs were not detectable in bands or mature ne utrophils, nor at significant levels in tissues other than bone marrow , The accumulation of defensin protein in bone marrow cells was assess ed by immunohistochemical staining with anti-RatNP-1 Ab. RatNP 1-4 mRN As and protein levels were then correlated for each stage of neutrophi lic differentiation to reveal the maturational profile of myeloid defe nsin gene expression in the rat.