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Bacterial death induced by expression of the intracellular portion of human Fas
1Laboratory of Immune Cell Biology, Division of Basic Sciences, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, MD 20892, USA.
Abstract:
In attempting to produce the intracellular portion of human Fas (IC175 - 319) as a GST-fusion protein we found that expression of GST-IC175 - 319, but not GST alone or GST-IC231 - 298 (containing the Fas death domain), rapidly caused the death of host E. coli cells. Expression of GST-IC175 - 319 with a single amino acid substitution (V238N) corresponding to the mouse lprcg mutation, or E245A, which abolishes the ability of Fas to self-associate, did not kill bacteria. Deletional analysis identified a 20-amino acids region (Asp210 - Lys230) as essential for the killing activity, and introduction of a single amino acid substitution (T225P) in this 20 amino acid region markedly decreased the ability of Fas- IC175 - 319 to cause bacterial death. These data indicate that Fas can deliver a death signal in prokaryotic organisms by a means that shares some features with eukaryotic cells, and raise the possibility that certain mechanisms leading to programmed cell death may be conserved from bacteria to mammalian cells.
Insights
Human Fas protein triggers bacterial death, suggesting conserved cell death mechanisms. Specific regions and mutations in Fas influence its toxicity to E. coli, hinting at shared pathways from bacteria to mammals.
Area of Science:
- Molecular Biology
- Cell Biology
- Microbiology
Background:
- Human Fas is a key regulator of programmed cell death (apoptosis) in eukaryotic cells.
- Investigating conserved cell death mechanisms across different organisms can provide novel insights into fundamental biological processes.
Purpose of the Study:
- To investigate the potential of the intracellular portion of human Fas (IC175-319) to induce cell death in prokaryotic organisms (E. coli).
- To identify specific regions and mutations within Fas that are responsible for its toxicity to bacteria.
Main Methods:
- Expression of GST-fusion proteins of human Fas intracellular domains in E. coli.
- Site-directed mutagenesis to create specific amino acid substitutions (V238N, E245A, T225P).
- Deletional analysis to pinpoint critical regions for bacterial toxicity.
Main Results:
- Expression of GST-IC175-319, but not GST alone or GST-IC231-298, caused rapid E. coli cell death.
- Mutations V238N (lprcg homolog) and E245A (abolishing self-association) prevented bacterial killing.
- A 20-amino acid region (Asp210-Lys230) was identified as essential for toxicity, with T225P substitution reducing this effect.
Conclusions:
- Human Fas can deliver a death signal in E. coli, indicating conserved aspects of cell death pathways.
- These findings suggest that certain programmed cell death mechanisms may be conserved across prokaryotic and eukaryotic life forms.