Chromosomal type I toxin-antitoxin conservation, regulation, and function in E. coli

Bacterial chromosomal type I toxin-antitoxin systems consist of a small protein, typically under 60 amino acids, and a small RNA (sRNA) that represses toxin translation. These gene pairs have gained attention for their contribution to antibiotic resistance and resilience. However, biological functions for many remain elusive. For many pairs, it is also still unknown when the toxin and/or antitoxin gene are natively expressed within the bacterium. In this work, I outline how I used a simple BLASTn based protocol utilizing publicly available data to identify DNA sequence, protein sequence, and copy number conservation of three chromosomal type I toxin-antitoxins, tisB/istR-1, shoB/ohsC, and zor/orz and features important for both their regulation and their function. I also outline how I constructed various fluorescent reporter constructs to test the transcriptional and posttranscriptional regulation of zorO/orzO under various environmental conditions, rapidly and robustly. Finally, I examined the ability of the shoB/ohsC to confer antibiotic resistance to E. coli. Together, this work expands our understanding of chromosomal type I toxin-antitoxins in E. coli and their native function.