Atack, J. M. | Search bioRxiv

bioRxiv2020

Systematic analysis of REBASE identifies numerous Type I restriction-modification systems that contain duplicated, variable hsdS specificity genes that randomly switch methyltransferase specificity by recombination.

N6-adenine DNA methyltransferases associated with some Type I and Type III restriction-modification (R-M) systems are able to randomly switch expression by variation in the length of locus-encoded simple sequence repeats (SSRs). SSR tract-length variation causes ON/OFF switching of methyltransferase expression, resulting in genome-wide methylation differences, and global changes in gene expression. These epigenetic regulatory systems are called phasevarions, phase-variable regulons, and are widespread in bacteria. A distinct switching system has also been described in Type I R-M systems, based on recombination-driven changes in hsdS genes, which dictate the DNA target site. In order to determine the prevalence of recombination-driven phasevarions, we generated a program called RecombinationRepeatSearch to interrogate REBASE and identify the presence and number of inverted repeats of hsdS downstream of Type I R-M loci. We report that 5.9% of Type I R-M systems have duplicated variable hsdS genes containing inverted repeats capable of phase-variation. We report the presence of these systems in the major pathogens Enterococcus faecalis and Listeria monocytogenes, which will have important implications for pathogenesis and vaccine development. These data suggest that in addition to SSR-driven phasevarions, many bacteria have independently evolved phase-variable Type I R-M systems via recombination between multiple, variable hsdS genes. ImportanceMany bacterial species contain DNA methyltransferases that have random on/off switching of expression. These systems called phasevarions (phase-variable regulons) control the expression of multiple genes by global methylation changes. In every previously characterised phasevarion, genes involved in pathobiology, antibiotic resistance, and potential vaccine candidates are randomly varied in their expression, commensurate with methyltransferase switching. A systematic study to determine the extent of phasevarions controlled by invertible Type I R-M systems has never before been performed. Understanding how bacteria regulate genes is key to the study of physiology, virulence, and vaccine development; therefore it is critical to identify and characterize phase-variable methyltransferases controlling phasevarions.

microbiology↗

bioRxiv2020

The non-typeable Haemophilus influenzae major adhesin Hia is a dual function lectin that binds to human-specific respiratory tract sialic acid glycan receptors.

NTHi is a human-adapted pathogen that colonises the human respiratory tract. Strains of NTHi express multiple adhesins, however there is a unique, mutually exclusive relationship between the major adhesins Hia and HMW1/2. Approximately 25% of NTHi strains express Hia, a phase-variable autotransporter protein, and which has a critical role in colonisation of the host nasopharynx. The remaining 75% of strains express HMW1/2. Previous work has shown that the HMW1 and HMW2 proteins mediate binding to 2,3- and 2,6-linked sialic acid glycans found in the human respiratory tract. Here we show that that the high affinity binding domain of Hia, binding domain 1 (BD1) is responsible for binding to 2,6-sialyllactosamine glycans. BD1 is highly specific for glycans that incorporate the form of sialic acid expressed by humans, N-acetylneuraminic acid (Neu5Ac). We further show that Hia has lower affinity binding activity for 2,3-linked sialic acid and that this binding activity is mediated via a distinct domain. Thus, Hia with its dual binding activities functionally mimics the combined activities of the HMW1 and 2 adhesins. In addition, we show that Hia has a role in biofilm formation by strains of NTHi that express the adhesin. Knowledge of the binding affinity of a major NTHi adhesin, and putative vaccine candidate, will direct and inform development of future vaccines and therapeutic strategies for this important pathogen. ImportanceHost-adapted bacterial pathogens like NTHi have evolved specific mechanisms to colonize their restricted host niche. Relatively few of the adhesins expressed by NTHi have been characterized as regards their binding affinity at the molecular level. In this work we show that the major NTHi adhesin, Hia, preferentially binds to Neu5Ac-2,6-sialyllactosamine, the form of sialic acid expressed in humans. The receptors targeted by Hia in the human airway mirror those targeted by influenza A virus and indicates the broad importance of sialic acid glycans as receptors for airway pathogens.

microbiology↗