Trends in Microbiology
Volume 13, Issue 8, August 2005, Pages 360-365
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The PIN-domain toxin–antitoxin array in mycobacteria

https://doi.org/10.1016/j.tim.2005.06.008Get rights and content

PIN-domains (homologues of the pilT N-terminal domain) are small protein domains of ∼140 amino acids. They are found in a diverse range of organisms and recent evidence from bioinformatics, biochemistry, structural biology and microbiology suggest that the majority of the prokaryotic PIN-domain proteins are the toxic components of toxin-antitoxin (TA) operons. Several microorganisms have a large cohort of these operons. For example, the genome of Mycobacterium tuberculosis encodes 48 PIN-domain proteins, of which 38 are thought to be involved in TA interactions. This large array of PIN-domain TA operons raises questions as to their evolutionary origin and contemporary functional significance. We suggest that the evolutionary origin of genes encoding mycobacterial PIN-domain TA operons is linked to the mobile gene pool, but that TA operons can become resident within the chromosome of host cells from where they might be recruited to fulfil a variety of roles associated with retardation of cell growth and persistence in stressful environments.

Introduction

Proteins containing PIN-domains (homologues of the pilT N-terminal domain) are found in the genomes of a wide range of prokaryotes and eukaryotes. The biochemical and biological functions of these proteins is uncertain, however, recent work has resulted in a convergence of opinion. In eukaryotes PIN-domain proteins function as ribonucleases 1, 2 with activity linked to RNAi and nonsense-mediated RNA degradation 1, 3. In prokaryotes, the majority of PIN-domain proteins are the toxic components (by virtue of their ribonuclease activity) of chromosomally encoded toxin-antitoxin (TA) operons 4, 5, 6. A striking feature of the PIN-domain TA operons is their abundance in the genomes of several unrelated prokaryotes. For example, the genome of Mycobacterium tuberculosis harbours 38 PIN-domain TA operons 3, 7 (Arcus, V.L., unpublished results) and the hypertheromophilic chrenarchaeote, Sulfolobus tokodaii, has 25 PIN-domain TAs [6]. These arrays of PIN-domain TA operons in various unrelated prokaryotic genomes pose questions as to their evolutionary origins and contemporary functional significance. Recent results, which suggest that TA operons confer advantages on competing mobile genetic elements 8, 9, combined with the association of PIN-domain TA operons with potential mobile elements in M. tuberculosis, point to the horizontal gene pool as the primary source of TA operons. Indeed, their abundance within the genomes of bacteria such as M. tuberculosis could reflect recent and rapid evolution fuelled by the acquisition of new DNA. Contemporary functions for TA operons are various but point collectively toward a general role in the facilitation of persistence in organisms that inhabit variable and frequently stressful environments [6].

Section snippets

Toxin–antitoxin protein pairs

Toxin–antitoxin (TA) protein pairs were discovered more than 20 years ago as factors that protect low copy number plasmids in bacteria from segregational loss 10, 11. One protein of the pair is toxic to the cell and stable, whereas its cognate antitoxin is unstable and requires continuous transcription to inhibit the toxin. Thus, if the plasmid is not correctly segregated during cell division, one daughter cell lacks the plasmid and is left with just the TA proteins – one stable and toxic, the

PIN-domain proteins

PIN-domain-containing proteins are encoded within the genomes of organisms that span the three domains of life. The PIN-domain was first annotated on the basis of sequence similarity to the N-terminal domain of the type IV pili protein, pilT, from Myxococcus xanthus (PIN, PilT N-terminus) [20], and is represented in the Pfam database (http://www.sanger.ac.uk/Software/Pfam) as PF01850, currently with 468 members. This number is likely to be an underestimate because the Superfamily database,

The biochemical function of PIN-domain proteins

Recent work on the structure and function of PIN-domain-containing proteins in the hyperthermophilic crenarchaeote Pyrobaculum aerophilum and the human pathogen Mycobacterium tuberculosis 22, 23 has focused on a small, well-conserved family of PIN-domains with four homologues in each organism (these eight proteins belong to a ‘Cluster of Orthologous’ proteins, COG4113, NCBI). By solving the 3D structure of one member of the family, a structural and functional relationship was shown between the

Several microorganisms have expanded cohorts of PIN-domain proteins

Bioinformatic surveys of PIN-domain proteins across many fully sequenced organisms showed that several organisms (notably Mycobacterium tuberculosis and Archeoglobus fulgidus) have a dramatically expanded cohort of PIN-domain proteins in their genomes 3, 4. Several other prokaryotic genomes, whose cohort of PIN domains exceeds ten, include Gloeobacter violaceus (a Cyanobacterium and obligate photoautotroph), Nitrosomonas europaea (a Gram-negative obligate chemolithoautotroph) and several

PIN-domain proteins in mycobacteria

M. tuberculosis has 48 PIN-domain-containing proteins [3], which are highly diverse at the DNA sequence level. These PIN-domains can be discovered by protein sequence-based Hidden Markov Model (HMM) methods, such as those used by the TIGRFAM database (47 PIN domains in M. tuberculosis) [7], or structure-based HMM methods such as those used by the Superfamily database [21] (46 PIN-domain sequences in M. tuberculosis, E-values <0.01). Alternatively, using an approach similar to that used by

Possible biological roles for PIN-domain toxin-antitoxin proteins

In assessing the possible reasons for the expanded repertoires of PIN-domain TAs in the mycobacteria and several other evolutionarily distant organisms, it is important to differentiate between the factors leading to their incorporation and/or expansion in the genome, and the consequences of their incorporation (i.e. their present biological functions), which might be only distantly related to their function at the time of incorporation.

The evolutionary origins of TA elements are unclear;

Conclusions

Bioinformaticians were the first to hypothesize a biochemical role for PIN-domain proteins as ribonucleases [1] and suggested that they represented the toxic components of prokaryotic TA operons [3]. Subsequent structural [23], biochemical and microbiological experiments [5] have confirmed these hypotheses. However, there is much yet to learn about these TA systems. For example, are the PIN-domains sequence specific ribonucleases similar to the toxin MazF? What are the targets for the toxic

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