Membrane vesicles (MVs) are nanoparticles composed of lipid membranes that are produced by both Gram-negative and Gram-positive bacteria. MVs have been assigned diverse biological functions, and they show great potential for applications in various fields. However, the mechanisms underlying their functions and biogenesis are not completely understood. Accumulating evidence shows that MVs are heterogenous, and different types of MVs with different compositions are released from the same species. To understand the origin and function of these MVs, determining the biochemical properties of MVs is important. In this review, we will discuss recent progress in understanding the biochemical composition and properties of MVs.

Bacterial membrane vesicles (MVs) are attracting considerable attention in diverse fields of life science and biotechnology due to their potential for various applications. Although there has been progress in determining the mechanisms of MV formation in Gram-negative and Gram-positive bacteria, the mechanisms in mycolic acid-containing bacteria remain an unsolved question due to its complex cell envelope structure. Here, by adapting super-resolution live-cell imaging and biochemical analysis, we show that Corynebacterium glutamicum form distinct types of MVs via different routes in response to environmental conditions. DNA-damaging stress induced MV formation through prophage-triggered cell lysis, whereas envelope stress induced MV formation through mycomembrane blebbing. The MV formation routes were conserved in other mycolic acid-containing bacteria. Our results show how the complex cell envelope structure intrinsically generates various types of MVs and will advance our knowledge on how different types of MVs can be generated from a single cell organism.

Although cyclic imines are present in various bioactive secondary metabolites, their degradative metabolism remains unknown. Here, we report that copper amine oxidases, which are important in metabolism of primary amines, catalyze a cyclic imine cleavage reaction. We isolate a microorganism (Arthrobacter sp. C-4A) which metabolizes a β-carboline alkaloid, harmaline. The harmaline-metabolizing enzyme (HarA) purified from strain C-4A is found to be copper amine oxidase and catalyze a ring-opening reaction of cyclic imine within harmaline, besides oxidative deamination of amines. Growth experiments on strain C-4A and Western blot analysis indicate that the HarA expression is induced by harmaline. We propose a reaction mechanism of the cyclic imine cleavage by HarA containing a post-translationally-synthesized cofactor, topaquinone. Together with the above results, the finding of the same activity of copper amine oxidase from E. coli suggests that, in many living organisms, these enzymes may play crucial roles in metabolism of ubiquitous cyclic imines.

Extracellular contractile injection systems (eCISs) are structurally similar to headless phages and are versatile nanomachines conserved among diverse classes of bacteria. Herein, Streptomyces species, which comprise filamentous Gram-positive bacteria and are ubiquitous in soil, were shown to produce Streptomyces phage tail-like particles (SLPs) from eCIS-related genes that are widely conserved among Streptomyces species. In some Streptomyces species, these eCIS-related genes are regulated by a key regulatory gene, which is essential for Streptomyces life cycle and is involved in morphological differentiation and antibiotic production. Deletion mutants of S. lividans of the eCIS-related genes appeared phenotypically normal in terms of morphological differentiation and antibiotic production, suggesting that SLPs are involved in other aspects of Streptomyces life cycle. Using co-culture method, we found that colonies of SLP-deficient mutants of S. lividans were more severely invaded by fungi, including Saccharomyces cerevisiae and Schizosaccharomyces pombe. In addition, microscopic and transcriptional analyses demonstrated that SLP expression was elevated upon co-culture with the fungi. In contrast, co-culture with Bacillus subtilis markedly decreased SLP expression and increased antibiotic production. Our findings demonstrate that in Streptomyces, eCIS-related genes affect microbial competition, and the patterns of SLP expression can differ depending on the competitor species.

Recent bioinformatic analyses have revealed that CIS-related gene clusters are highly conserved in Gram-positive actinomycetes, especially members of the genus Streptomyces known for their ability to produce therapeutic antibiotics. While typical CISs are released from the cells and can act as protein translocation systems that inject effector proteins into the target cells, our results indicate the unique intracellular localization of SLPs, CIS-related nanostructures produced by S. lividans . In addition, the direct and indirect interactions of SLPs with cytoplasmic proteins and SLP localization within specific regions of mycelia suggest that the biological significance of SLPs is related to intracellular processes.