Replisome Trafficking in Growing Vegetative Hyphae ofStreptomyces coelicolorA3(2)

Abstract

We observed movies of replisome trafficking during Streptomyces coelicolor growth. A replisome(s) in the spore served as a replication center(s) until hyphae reached a certain length, when a tip-proximal replisome formed and moved at a fixed distance behind the tip at a speed equivalent to the extension rate of the tip.Members of the bacterial genus Streptomyces exhibit mycelial growth and sporulation that are reminiscent of those of filamentous fungi. Spores germinate to form germ tubes (7,18), which extend to elongated hyphae through peptidoglycan incorporation at the hyphal tip (2, 4). Eventually, a mycelium is formed by branch emergence from the lateral hyphal walls. Incorporation of new cell wall material at the hyphal tip and branch points is, either directly or indirectly, dependent on the essential protein DivIVA (3). Typical cell division does not occur during vegetative growth, and elongated, multigenomic compartments are delimited by occasionally spaced septa. In contrast to most other bacteria, in this genus many genes required for cell division are inessential for vegetative growth and are required only during sporulation (14). The chromosomes in the vegetative hyphae seem to remain uncondensed and do not undergo typical segregation. Early studies using pulse-labeling revealed that hyphae did not show any region of preferential incorporation of the label and that replicating nucleoids were evenly distributed along the hyphae (12, 13). This indicates that DNA replication does not depend on nucleoid location; the corollary of this is that mechanisms must exist to allow chromosomes to populate the extending hypha. However, Yang and Losick (19) were unable to find any evidence that DNA replication activity was concentrated at the apex. Further studies using a functional DnaN-enhanced green fluorescent protein (EGFP) fusion demonstrated that DNA replication takes place in both apical and subapical compartments of Streptomyces coelicolor vegetative hyphae (17). Moreover, replication is asynchronous, and only selected chromosomes undergo replication at any given time (17). Recently, a new method for monitoring Streptomyces hyphal growth in real time using time-lapse microscopy was established by Jyothikumar et al. (11). It showed that, following germination, hyphal extension occurred at about 20 m h Ϫ1 when grown on mannitol-supplemented minimal medium at 30°C. Here, we present the results obtained by application of this method (11) to study replisome dynamics in vegetative mycelium. Captured images were processed using IPlabs 3.7 image processing software (BD Biosciences Bioimaging, Rockville, MD). Eleven

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The conjugative DNA-transfer apparatus of Streptomyces

“…Unlike most other bacteria that divide by binary fission, Streptomyces grow by apical tip extension at a rate of ∼16 m h −1 (Wolanski et al, 2011). The incorporation of new peptidoglycan (PG) precursors at the tips depends on the coiled coil proteins DivIVA, Scy and FilP, which form the polarisome (Hempel et al, 2008;Fuchino et al, 2013).…”

“…Streptomyces genomes with a size of 5.3-11.8 Mb (Chandra and Chater, 2013) belong to the largest bacterial genomes. During replication the replisomes follow tips at a speed equivalent to the rate of hyphal extension (Wolanski et al, 2011).…”

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