Four Chromosomal Type IV Secretion Systems in Helicobacter pylori: Composition, Structure and Function

Abstract

The pathogenic bacterium Helicobacter pylori is genetically highly diverse and a major risk factor for the development of peptic ulcer disease and gastric adenocarcinoma in humans. During evolution, H. pylori has acquired multiple type IV secretion systems (T4SSs), and then adapted for various purposes. These T4SSs represent remarkable molecular transporter machines, often associated with an extracellular pilus structure present in many bacteria, which are commonly composed of multiple structural proteins spanning the inner and outer membranes. By definition, these T4SSs exhibit central functions mediated through the contact-dependent conjugative transfer of mobile DNA elements, the contact-independent release and uptake of DNA into and from the extracellular environment as well as the secretion of effector proteins in mammalian host target cells. In recent years, numerous features on the molecular functionality of these T4SSs were disclosed. H. pylori encodes up to four T4SSs on its chromosome, namely the Cag T4SS present in the cag pathogenicity island (cagPAI), the ComB system, as well as the Tfs3 and Tfs4 T4SSs, some of which exhibit unique T4SS functions. The Cag T4SS facilitates the delivery of the CagA effector protein and pro-inflammatory signal transduction through translocated ADP-heptose and chromosomal DNA, while various structural pilus proteins can target host cell receptors such as integrins or TLR5. The ComB apparatus mediates the import of free DNA from the extracellular milieu, whereas Tfs3 may accomplish the secretion or translocation of effector protein CtkA. Both Tfs3 and Tfs4 are furthermore presumed to act as conjugative DNA transfer machineries due to the presence of tyrosine recombinases with cognate recognition sequences, conjugational relaxases, and potential origins of transfer (oriT) found within the tfs3 and tfs4 genome islands. In addition, some extrachromosomal plasmids, transposons and phages have been discovered in multiple H. pylori isolates. The genetic exchange mediated by DNA mobilization events of chromosomal genes and plasmids combined with recombination events could account for much of the genetic diversity found in H. pylori. In this review, we highlight our current knowledge on the four T4SSs and the involved mechanisms with consequences for H. pylori adaptation to the hostile environment in the human stomach.

Keywords: DNA transfer; cag; competence; conjugation; pathogenicity island; recombination; type IV secretion system; virulence.

FIGURE 1

Genetic arrangements of four chromosomally encoded type IV secretion system gene clusters in H. pylori. The arrows show typical arrangements of the cag, comB, tfs3, and tfs4 genes, with coloring according to proven T4SS functions and/or to sequence similarities with prototypical virB/virD genes, as indicated. The Cag, Tfs3, and Tfs4 systems are located on genome islands delimited by left (LJ) and right junctions (RJ), which consist of 31 bp (Cag) or 7 bp (Tfs3, Tfs4) direct sequence repeats. The competence (com) genes are not organized in a contiguous unit, but comprise two operons (comB2 to comB4 and comB6 to comB10) with genes homologous to the corresponding virB genes, and the comEC and comH genes located elsewhere in the genome. Gene designations for cag pathogenicity island genes represent the commonly used names, whereas genes conserved between ICEHptfs3 and ICEHptfs4 (C1 to C21) are indicated, together with some additional or alternative designations, according to Delahay et al. (2018), to illustrate similarities between these two islands. Note that individual genes (e.g., the methylase or ctkA genes) may be absent from some ICEs, and that most ICEHptfs4 genes and some ICEHptfs3 genes (e.g., C2, C3, C5) may occur in distinct allelic variants (regions where such variants occur are indicated by blue bars). On ICEHptfs4 islands, variants are usually grouped as left (L1/L2), central (C1/C2) or right (R1/R2) regions, which can be combined to form distinct ICE variants, such as L1C1R1, L2C1R2, etc.

FIGURE 2

Models for the assembly and subcellular localization of the Cag and ComB type IV secretion systems in the membranes of H. pylori. Proteins sharing sequence homology to the Agrobacterium tumefaciens VirB/VirD4 T4SS are marked with B1 to B11 or D4, respectively. Accessory Cag and Com proteins are designated with letters as given in Figure 1. The Cag T4SS pilus proteins, core complex components, NTP-hydrolysing energetic factors, translocated effector molecules (CagA, ADP-Heptose and dsDNA) and other proteins are emphasized with different colors as denoted in the legend at the bottom. The proposed position of the T4SS proteins is presented in a simplified fashion. In the ComB T4SS, an outer membrane complex might be composed of ComB7, B9 and B10. ComB3, B4, B6, B8 and B10 form a complex at the inner membrane. Hypothetically, the pilin subunit ComB2 binds to dsDNA at the surface of the bacterium (pseudopilus). By retraction of these pilin subunits, powered by the ATPase ComB4, bound dsDNA is translocated into the periplasm. ComH guides incoming dsDNA to the ComEC channel by binding dsDNA with its C-terminus and docking its N-terminus to a periplasmic domain of ComEC. In the cytoplasm, the helicase PriA might empower uptake of ssDNA. Please note that effector molecule delivery in both T4SSs proceeds in opposite directions as indicated. In contrast, almost nothing is known about the assembly and subcellular localization of Tfs3 and Tfs4 components.

FIGURE 3

Schematic presentation for the functional importance of four chromosomally encoded type IV secretion systems of H. pylori. (A) Effector molecule translocation into human host target cells comprises CagA, CtkA, ADP-heptose, and chromosomal DNA. The indicated T4SSs of the cagPAI and ICEHptfs3 are involved. Delivery of CagA requires the interaction of adhesin HopQ with CEACAM receptors. In addition, several indicated pilus-associated cagPAI proteins can interact directly with other cell surface receptors (integrins and TLR5) to induce signaling. (B) The ComB system functions to deliver naked DNA from the environment into H. pylori. Conjugative DNA transfer has been also observed among H. pylori strains (C), or from H. pylori to other bacteria such as Campylobacter jejuni (D).