# Bacterial Type I Toxins: Folding and Membrane Interactions
###### Abstract  
Bacterial type I toxin-antitoxin systems are two-component genetic modules that encode a stable toxic protein whose ectopic overexpression can lead to growth arrest or cell death, and an unstable RNA antitoxin that inhibits toxin translation during growth. These systems are widely spread among bacterial species. Type I antioxins are _cis_- or _trans_-encoded antisense small RNAs that interact with toxin-encoding mRNAs by pairing, thereby inhibiting toxin mRNA translation and/or inducing its degradation. Under environmental stress conditions, the up-regulation of the toxin and/or the antitoxin degradation by specific RNases promote toxin translation. Most type I toxins are small hydrophobic peptides with a predicted \(\alpha\)-helical transmembrane domain that induces membrane depolarization and/or permeabilization followed by a decrease of intracellular ATP, leading to plasmid maintenance, growth adaptation to environmental stresses, or persister cell formation. In this review, we describe the current state of the art on the folding and the membrane interactions of these membrane-associated type I toxins from either Gram-negative or Gram-positive bacteria and establish a chronology of their toxic effects on the bacterial cell. This review also includes novel structural results obtained by NMR concerning the _sprG1_-encoded membrane peptides that belong to the _sprG1_/SprF1 type I TA system expressed in _Staphylococcus aureus_ and discusses the putative membrane interactions allowing the lysis of competing bacteria and host cells. toxin-antitoxin systems; type I toxins; mechanisms of action; membrane depolarization; membrane permeabilization; pore formation; nucleoid condensation; structure; folding 2021 13 490 10.3900 1307090 10.3900 10.  
described in which the DarG antitoxin interacts with the DarT toxin to inhibit its expression, but also acts on the target of DarT [5; 6]. Antitoxins are unstable and more susceptible to degradation by ribonucleases or proteases than toxins, leaving toxins free to interfere with essential cellular functions such as replication, translation, or cell division [7]. In this review, we will focus exclusively on type I TA systems, characterized by a small antisense RNA that base-pairs with its cognate toxin-encoding messenger RNA (mRNA) to prevent the toxin synthesis under normal growth conditions. These systems are first predicted in the bacterial genomes through computational approaches [8]. However, only a few of them are experimentally characterized and mostly focused on _E. coli_. Type I TA systems were initially identified on plasmids, where they ensure plasmid maintenance through a post-segregational killing (PSK) mechanism [9]. In this process, because of the antitoxin degradation, the plasmid loss results in the decrease of the antitoxin cell concentration and the killing of the plasmid-free cell by the stable toxin. Later on, homologues of known type I TA systems were discovered on bacterial chromosomes [8; 10]. Although the biological role of chromosomal type I TA systems is still elusive, they can be involved in mobile genetic elements maintenance, growth adaptation to environmental stresses, or persister cells formation [11]. Persister cells represent a subpopulation of genetically identical and metabolically slow-growing bacteria that are tolerant to extremely high antibiotic doses after selection through repeated antibiotic therapy. They can result in treatment failure, relapse, and persistent bacterial infections [12]. Type I toxin expression is tightly controlled by RNA antioxins and _cis_-encoded mRNA functional elements [13]. Most type I antioxins are _cis_-acting antiotoxins (e.g., _holz_/Sok, _bsrG_/SR4, _sprG_/SPrF), meaning that the antitoxin and toxin loci overlap, resulting in a perfect pairing between the two RNAs. The _trans_-acting antiotoxins (e.g., _tisB_/IstR1, _din_Q/AgrB) are located away from the toxin locus and share often limited sequence complementarities.