Four serine proteinases expressed in Manduca sexta haemocytes

Several putative serine proteinases were detected in Manduca sexta larval plasma by labelling with radioactive diisopropyl fluorophosphate. To begin to identify and characterize such enzymes, a polymerase chain reaction was carried out using haemocyte cDNA as template and primers designed to amplify conserved sequences from serine proteinases. Four serine proteinase cDNA fragments were cloned. These were used as probes to screen an M. sexta larval haemocyte cDNA library to obtain full-length clones encoding haemocyte proteinases 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 snake and easter, HP3 and HP4 are most similar to proteinases from mammalian leucocytes. HP1 and HP2 are both present in plasma. HP1 is expressed in haemocytes (granular cells and oenocytoids) and not in fat body. HP2 is expressed in fat body and in granular haemocytes, plasmatocytes and oenocytoids, After injection of larvae with bacteria, the level of HP2 mRNA decreased in haemocytes and increased in fat body.