The type IV secretion system of Patescibacteria is homologous to the bacterial monoderm conjugation machinery

Abstract

The Candidate Phyla Radiation, also known as Patescibacteria, represents a vast and diverse division of bacteria that has come to light via culture-independent 'omics' technologies. Their limited biosynthetic capacity, along with evidence of their growth as obligate epibionts on other bacteria, suggests a broad reliance on host organisms for their survival. Nevertheless, our understanding of the molecular mechanisms governing their metabolism and lifestyle remains limited. The type IV secretion system (T4SS) represents a superfamily of translocation systems with a wide range of functional roles. T4SS genes have been identified in the Patescibacteria class Saccharimonadia as essential for their epibiotic growth. In this study, we used a comprehensive bioinformatics approach to investigate the diversity and distribution of T4SS within Patescibacteria. The phylogenetic analysis of the T4SS signature protein VirB4 suggests that most of these proteins cluster into a distinct monophyletic group with a shared ancestry to the MPF_FATA class of T4SS. This class is found in the conjugative elements of Firmicutes, Actinobacteria, Tenericutes and Archaea, indicating a possible horizontal gene transfer from these monoderm micro-organisms to Patescibacteria. We identified additional T4SS components near virB4, particularly those associated with the MPF_FATA class, as well as homologues of other T4SS classes, such as VirB2-like pilins, and observed their varied arrangements across different Patescibacteria classes. The absence of a relaxase in most of these T4SS clusters suggests that the system has been co-opted for other functions in Patescibacteria. The proximity of T4SS components to the origin of replication (gene dnaA) in some Patescibacteria suggests a potential mechanism for increased expression. The broad ubiquity of a phylogenetically distinct T4SS, combined with its chromosomal location, underscores the significance of T4SS in the biology of Patescibacteria.

Keywords: Candidate Phyla Radiation; Patescibacteria; horizontal gene transfer; type IV secretion system.

Fig. 1.. Detection of VirB4 homologues in Patescibacteria. (a) VirB4 abundance in the CPR dataset. Stacked bar plot showing the proportion of genomes with (red) and without (grey) VirB4 homologues for each class in the dataset. The total number of genomes per class (n) is displayed at the top of each bar. (b) Distribution of VirB4 homologues according to genome size. The 3,026 assemblies were ranked by size (307,478–2,280,175 bp), shown on the x-axis. The top panel displays a cumulative distribution function (CDF) plot of genome size, while the lower panels use vertical lines to indicate the presence (red) or absence (grey) of a VirB4 homologue in each corresponding genome.

Fig. 2.. Phylogenetic analysis of VirB4 proteins. Maximum likelihood tree of the VirB4 homologues of the Patescibacteria dataset and members from different T4SS classes rooted at the midpoint. Nodes with UFBoot support values ≥80% are indicated by a grey circle. Rings from inside to outside indicate: (1) T4SS type and (2) Patescibacteria class. The Patescibacteria clade is shadowed in grey.

Fig. 3.. Cumulative distribution function of the minimal patristic distances in the VirB4 tree. The curves show the sum of the lengths of the branches linking two nodes in the VirB4 phylogenetic tree, belonging to the closest homologues within the same class (red) or different classes (blue).

Fig. 4.. Abundance of T4SS components near virB4. Each column represents the abundance of T4SS components at a specific position relative to the virB4 gene (within a range of −20 to +20 coding sequences). Only virB4 genes associated with another T4SS component were considered. The total count of T4SS components at each position is shown above each column and represents 100%. The T4SS components are colour-coded according to the legend. VirB1 comprises homologues retrieved with HMM profiles MPF_FATA TrsG and CD419 and MPF_T VirB1; VirB2 those retrieved with HMM profiles MPF_B TraE, MPF_G Tfc9 and Tfc10, MPF_I TraQ and TraR, and MPF_T VirB2, and Pfam PF04956.16 and PF18895.3; VirB3 with MPF_B TraF, MPF_C Alr705, MPF_F TraL, MPF_FATA PrgI and TrsC, MPF_G Tfc11, MPF_I TraP and MPF_T VirB3; VirB6 with MPF_FATA PrgH and MPF_T VirB6; and VirB8 with MPF_F TraE, MPF_FATA PrgL and MPF_T VirB8.

Fig. 5.. Proximity of the virB4 gene to other chromosomal functions. For the 7 Patescibacteria classes with 15 or more genomes in the dataset, the 6 most prevalent (a) Pfam, (b) NCBIFAM and (c) TIGRFAM families identified near virB4 (within a range of −20 to +20 coding sequences) are shown. Data from the remaining classes are grouped under ‘Other’. The count of detected members for each protein family is displayed above the respective bar, while the total number of genomes analysed per class is also indicated.

Fig. 6.. Genomic context of T4SS in representative genomes. For the four most represented Patescibacteria classes in which a T4SS was identified, the synteny of the virB4 gene neighbourhood is illustrated using a representative from complete Patescibacteria genomes. The genetic organization encompasses 20 CDSs upstream and downstream of the virB4 gene (coloured in red and located at the centre). T4SS genes other than virB4 are coloured in light red, and non-T4SS genes are depicted in different colours according to the legend. Genes for which no homology to NCBIFAM, Pfam-A or TIGRFAM profiles were found are coloured in grey. T4SS prototypes of the MPF_T and MPF_FATA types, as well as an example of the T4SS protein translocators in streptococci, are also included.