THE INDUCTION OF LYSOZYME AND OTHER ANTIBACTERIAL HEMOLYMPH PROTEINS IN THE TOBACCO HORNWORM, MANDUCA SEXTA
Abstract
Injection of formalin-killed bacteria into larvae of Manduca sexta elicited an increase in the concentration of several small (MW < 25000), basic serum proteins which could be detected by polyacrylamide gel electrophoresis beginning at 12 hr after inoculation. The enzyme lysozyme was one of these proteins. After a 6 hr lag, serum lysozyme activity increased for 24-72 hr after injection. By 48 hr after injection of 10('9) killed bacteria, lysozyme activity was approximately ten-fold greater than in untreated insects. This maximal response was observed after injection of every bacterial species tested and after injection of purified cell walls of Micrococcus luteus. Several types of polysaccharides, animal cells, and acellular particles induced much smaller increases in serum lysozyme activity. Injections of saline or protein solutions and some animal cells did not induce increased lysozyme activity. Thus, the lysozyme response has a specificity for bacteria. Injection of peptidoglycan, a component of all bacterial cell walls, induced large increases in concentration of lysozyme and a group of proteins with bactericidal activity against Escherichia coli, bactericidins. Low molecular weight fragments of peptidoglycan, released from M. luteus cell walls by lysozyme digestion, were as active as whole cell walls in inducing lysozyme and bactericidins. Peptidoglycan fragments may provide a signal which initiates a group of antibacterial responses in the insect. Lysozyme and bactericidins were synthesized in and secreted from fat body of M. sexta incubated in vitro. Isolated fat body incorporated ('3)H-leucine into lysozyme, which was released into the culture medium. Lysozyme released by isolated fat body of M. sexta was identified by electrophoretic rate in two systems, immunodiffusion, and electrophoretic analysis of immunoprecipitates as the same enzyme which is present in hemolymph. Fat body from insects previously injected with M. luteus cell walls (treated) released lysozyme at a greater rate than untreated insects. Incubation in the presence of peptidoglycan fragments resulted in the release of lysozyme at an increased rate by fat body from both treated and untreated insects. Bactericidins were not usually released by fat body from untreated or treated insects unless it was cultured in the presence of peptidoglycan fragments.
Degree
Ph.D.
Subject Area
Entomology
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