Subcompartmentalization by cross-membranes during early growth of Streptomyces hyphae

Abstract

Bacteria of the genus Streptomyces are a model system for bacterial multicellularity. Their mycelial life style involves the formation of long multinucleated hyphae during vegetative growth, with occasional cross-walls separating long compartments. Reproduction occurs by specialized aerial hyphae, which differentiate into chains of uninucleoid spores. While the tubulin-like FtsZ protein is required for the formation of all peptidoglycan-based septa in Streptomyces, canonical divisome-dependent cell division only occurs during sporulation. Here we report extensive subcompartmentalization in young vegetative hyphae ofStreptomyces coelicolor, whereby 1 μm compartments are formed by nucleic acid stain-impermeable barriers. These barriers possess the permeability properties of membranes and at least some of them are cross-membranes without detectable peptidoglycan. Z-ladders form during the early growth, but cross-membrane formation does not depend on FtsZ. Thus, a new level of hyphal organization is presented involving unprecedented high-frequency compartmentalization, which changes the old dogma that Streptomyces vegetative hyphae have scarce compartmentalization.

Introduction

Streptomycetes are filamentous Gram-positive bacteria that are of great importance for biotechnology given their ability to produce a large array of natural products, including antibiotics, anticancer agents and immunosuppressants, as well as a plethora of industrial enzymes1,2.

The Streptomyces life cycle has largely been studied during the growth of surface-grown cultures3,4,5,6,7,8 (Fig. 1). The life cycle starts with the germination of a spore, which expands out via tip growth and hyphal branching to form a vegetative mycelium consisting of multinucleate compartments3. When dispersal is required, for example, after nutrient depletion, the vegetative mycelium eventually differentiates into a new so-called aerial mycelium, which grows into the air. The aerial hyphae are also initially multinucleated, but these eventually develop sporogenic structures that differentiate into chains of unigenomic spores4. The lysis of the substrate mycelium and, later, the early aerial mycelium, exhibits the hallmarks of programmed cell death (PCD), with the involvement of specific lytic enzymes (nucleases, proteases and muramidases)5,6. New features affecting early development have been described over the last decade7,8. An early compartmentalized mycelium (MI) undergoes an early PCD-like process affecting the substrate and aerial hyphae6. Remarkably, live and dying cells are alternately observed in the MI hyphae6. The lifespan of this young mycelium is very short under laboratory conditions, but it is likely the predominant mycelium in cultures grown under natural conditions, such as in non-amended soils7. The substrate and aerial hyphae in the MII phase6 are physiologically different from those in the MI phase. MI corresponds to the vegetative mycelium, whereas the substrate and aerial mycelia are the reproductive stages driving towards sporulation8. Secondary metabolism is typically restricted to the MII phase8.

Figure 1: Development of Streptomyces on solid agar plates.

The Streptomyces developmental cycle3,4,5,6,7,8 along the transverse axis of the plate is illustrated (the agar border is indicated with a dashed line). Discontinuities in hyphal membranes represent changes in membrane permeability in dying cells. Red and green colours represent PI fluorescence (dying cells) and SYTO9 fluorescence (live cells), respectively. The traditional nomenclature for substrate and aerial mycelia is indicated in black letters (right); the new nomenclature for MI and MII stages is indicated in red letters (left).