Bacteria-sensitive lethal mutants

Figure 6. Bacteria-sensitive lethal mutants are inducible for antibacterial proteins.

Mutants were screened by mutagenizing males with 4000 R of gamma-rays and mating them to y v attached-X females. The G1 males were mated again to the attached-X females to avoid mosaics and the G2 males were individually mated to attached-X females. Five G3 sons of each individual G2 male were inoculated with 0.1 microliters of an overnight culture of Escherichia coli A-585. Three days later lines were scored for survival. Lines in which 4 or 5 of the G3 males had died were retested to confirm that they were bacteria-sensitive lethal mutations. 

From our bacteria sensitive lethal mutations we can make the surprising conclusion that the presence of ABs detected after overlaying an isoelectric focusing gel with bacteria is insufficient for a fly to survive a bacterial attack. Therefore, at least some of our bsl mutations identify genes required for surviving a bacterial infection that are independent of the ABs. We hypothesize that the necessary function identified by the bsl mutants is a cellular immune function, perhaps phagocytosis of bacteria. 

The immune system of Drosophila seems to have two main subsystems, one involving the antibacterial proteins and the other identified by bacteria-sensitive lethal mutations. The humoral subsystem results in the production of diffusible antibacterial proteins of at least three species. Stock Cu, which lacks the ABs found in wild type Drosophila and which is correspondingly more sensitive to infection, blocks a part of this subsystem. That the ABs do help flies to survive bacterial infections was shown by inducing ABs to appear in bsl mutants by injection of dead bacteria and then finding that the inoculated mutants would survive an otherwise lethal injection of live bacteria. Evidently inoculation induces processes (perhaps secretion of ABs) that can overcome the deficiency in the bsl mutations. Similar experiments showed that inoculation protects locusts from a lethal dose of bacteria (19). 

Finally, the current work provides bacteria-sensitive lethal mutations that point out pest control possibilities. Since bsl mutants die when injected with bacteria, they focus our attention directly on the immune steps at which the insect is particularly vulnerable. In future work we need to identify the cellular and molecular processes that are blocked by these mutations. If we could learn to artificially interfere with an immune step that is blocked by a bsl, then an effective insecticide might be developed. Because of the demonstrated similarities in the antibacterial immune systems of Drosophila melanoqaster and fruit fly pests, it is hoped that such methods would lead to crop protection. 

Temperature-sensitive ts) mutants have proven to be the most useful type of mutants for a number of viruses and bacteria because of their conditional-lethal phenotype. The (ts) mutants are produced by alteration in the nucleotide sequence of a gene so that the resulting protein product of the gene is unable to assume or maintain its functional configuration at the non-permissive (37-39°C) temperature. The protein, however, is able to assume a functional configuration at the permissive temperature (32-34° C), e.g., herpesviruses, adenoviruses, and influenza viruses. Thus, these mutants can replicate in mucosal sites with a lower temperature, e.g., the nasal cavity, but are unable to cause systemic infections and disease. 

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