Four serine proteinases expressed in Manduca sexta haemocytes

Several putative serine proteinases were detected in Manduca sexta larval p lasma by labelling with radioactive diisopropyl fluorophosphate. To begin t o identify and characterize such enzymes, a polymerase chain reaction was c arried out using haemocyte cDNA as template and primers designed to amplify conserved sequences from serine proteinases. Four serine proteinase cDNA f ragments were cloned. These were used as probes to screen an M. sexta larva l haemocyte cDNA library to obtain full-length clones encoding haemocyte pr oteinases 1-4 (HP1, HP2, HP3 and HP4). HP1 and HP2 contain an aminoterminal 'clip' domain similar to those found in horseshoe crab clotting enzyme and clotting factor B and also in the Drosophila melanogaster proteinases snak e and easter, HP3 and HP4 are most similar to proteinases from mammalian le ucocytes. HP1 and HP2 are both present in plasma. HP1 is expressed in haemo cytes (granular cells and oenocytoids) and not in fat body. HP2 is expresse d in fat body and in granular haemocytes, plasmatocytes and oenocytoids, Af ter injection of larvae with bacteria, the level of HP2 mRNA decreased in h aemocytes and increased in fat body.