Transcriptional and Proteolytic Regulation of the Toxin-Antitoxin Locus vapBC10 (ssr2962/slr1767) on the Chromosome of Synechocystis sp. PCC 6803

Abstract

VapBC toxin-antitoxin (TA) systems are defined by the association of a PIN-domain toxin with a DNA-binding antitoxin, and are thought to play important physiological roles in bacteria and archaea. Recently, the PIN-associated gene pair PIN-COG2442 was proposed to encode VapBC-family TA system and found to be abundant in cyanobacteria. However, the features of these predicted TA loci remain under investigation. We here report characterization of the PIN-COG2442 locus vapBC10 (ssr2962/slr1767) on the chromosome of Synechocystis sp. PCC 6803. RT-PCR analysis revealed that the vapBC10 genes were co-transcribed under normal growth conditions. Ectopic expression of the PIN-domain protein VapC10 caused growth arrest of Escherichia coli that does not possess vapBC TA locus. Coincidentally, this growth-inhibition effect could be neutralized by either simultaneous or subsequent production of the COG2442-domain protein VapB10 through formation of the TA complex VapBC10 in vivo. In contrast to the transcription repression activity of the well-studied antitoxins, VapB10 positively auto-regulated the transcription of its own operon via specific binding to the promoter region. Furthermore, in vivo experiments in E. coli demonstrated that the Synechocystis protease ClpXP2s, rather than Lons, could cleave VapB10 and proteolytically activate the VapC10 toxicity. Our results show that the PIN-COG2442 locus vapBC10 encodes a functional VapBC TA system with an alternative mechanism for the transcriptional auto-regulation of its own operon.

Type II Toxin–Antitoxin Systems in the Unicellular Cyanobacterium Synechocystis sp. PCC 6803

“…Hence, growth inhibition by the respective toxin and corresponding attenuation by the antitoxin has been demonstrated for altogether 11 Synechocystis 6803 TA systems: five located on plasmid pSYSA were previously analyzed [ 23 ], one located on plasmid pSYSG in this work ( Figure 3 ) and five chromosomal TA systems analyzed before [ 18 , 20 ] or in this work ( Figure 3 ).…”

“…According to the excludon concept [ 34 ], enhanced expression of the slr1767–ssr2962 dicistron might lead to operon discoordination and lowered expression of the glutamate decarboxylase gene compared to the first two genes within TU520. The Slr1767–Ssr2962 TA system is functional, and has previously been characterized as a bona fide TA system [ 20 ]. However, while this is the first report of a TA system in such an arrangement, the comparison to the closely related strain Synechocystis 6714 revealed the homologs of the genes slr1639–slr1641 to be conserved and within one operon, while the TA locus was missing.…”

“…Makarova et al (2009) [ 3 ] identified a total of 29 Type II TA pairs in Synechocystis 6803, whereas the TA Database [ 16 , 17 ] listed a total of 36 TA pairs, which are distributed over the chromosome and the four larger plasmids (Chr: 19; pSYSM: 5; pSYSX: 2; pSYSA: 6; pSYSG: 4). From these, only four chromosomally located TA pairs [ 18 , 19 , 20 , 21 ], one pair on plasmid pSYSX [ 22 ] and five located on plasmid pSYSA [ 23 ] were targeted experimentally thus far.…”

Towards Exploring Toxin-Antitoxin Systems in <em>Geobacillus</em>: A Screen for Type II Toxin-Antitoxin System Families in A Thermophilic Genus

“…VapBC (Virulence associated protein) is the most widespread TA family in bacteria and archaea [89], but the least well-described [90,91]. The VapBC family is composed of a protein harboring a PIN domain as the toxin, and a DNA-binding domain as the antitoxin [90].…”

“…The PIN (PilT N-terminus) domain, is a type of pili protein [90] associated with the ribonuclease activity of the VapC toxin, which has a bacteriostatic effect [89,92]. The PIN-like domain-harboring toxin could associate with any protein harboring a DNA-binding domain as an antitoxin [90]. Different TA composites have been reported [63,91], one of which is the COG2886-PIN composite [62].…”