Genome Sequence of the Filamentous Actinomycete
            <i>Kitasatospora viridifaciens</i>

Ramijan, Karina; Wezel, Gilles P. van; Claessen, Dennis

doi:10.1128/genomea.01560-16

Cited by 12 publications

(15 citation statements)

References 12 publications

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“…This phenotype is reminiscent of the defective sporulation seen in colonies of Streptomyces clavuligerus that have lost the large linear plasmid pSCL4 following protoplast formation and regeneration 28 . Given that K. viridifaciens contains a large megaplasmid (KVP1 29 ), we reasoned that S-cell formation could increase the frequency of the loss of this plasmid. To test this assumption, we performed quantitative real-time PCR using four genes contained on the megaplasmid ( orf1, parA, tetR , and allC ).…”

Section: Resultsmentioning

confidence: 99%

“…Whole-genome sequencing followed by de novo assembly (Illumina and PacBio) and variant calling analyses were performed by BaseClear (Leiden, The Netherlands). The unique mutations were identified by direct comparison to the parental strain Kitasatospora viridifaciens DSM40239 (GenBank accession number PRJNA353578 29 ). The single and multiple nucleotide variations were identified using a minimum sequencing coverage of 50 and a variant frequency of 70%.…”

Section: Methodsmentioning

confidence: 99%

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Stress-induced formation of cell wall-deficient cells in filamentous actinomycetes

Ramijan

Ultee

Willemse

et al. 2016

Preprint

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The cell wall is a shape-defining structure that envelopes almost all bacteria. One of its main functions is to serve as a protection barrier to environmental stresses. Bacteria can be forced in a cell wall-deficient state under highly specialized conditions, which are invariably aimed at interrupting cell wall synthesis. Therefore, the relevance of such cells has remained obscure. Here we show that many filamentous actinomycetes have a natural ability to generate a new, cell wall-deficient cell type in response to hyperosmotic stress, which we call S-cells. This wall-deficient state is transient, as S-cells are able to switch to the canonical mycelial mode-of-growth. Remarkably, prolonged exposure of S-cells to hyperosmotic stress yielded variants that are able to proliferate indefinitely without their cell wall. This is the first report that demonstrates the formation of wall-deficient cells as a natural adaptation strategy and their potential transition into stable wall-less forms solely caused by prolonged exposure to osmotic stress. Given that actinomycetes are potent antibiotic producers, our work also provides important insights into how biosynthetic gene clusters and resistance determinants may disseminate into the environment.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Stress-induced formation of cell wall-deficient cells in filamentous actinomycetes

Ramijan

Ultee

Willemse

et al. 2016

Preprint

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“…Since the cytoskeletal protein FilP localizes to actively growing hyphal tips and is linked to the sturdiness of the tip in Streptomyces 10 , we reasoned that it could play a significant role in the release of S-cells from osmotically stressed hyphae. We therefore created a knock-out mutant strain of the filP homologue in K. viridifaciens (BOQ63_RS19300 18 , Supplementary Fig. 1).…”

Section: Resultsmentioning

confidence: 99%

“…The wild-type strain Kitasatospora viridifaciens DSM40239 18 was obtained from DSMZ. Spores were harvested after 7 days of growth on MYM agar medium 35 .…”

Section: Methodsmentioning

confidence: 99%

Formation of wall-less cells inKitasatospora viridifaciensrequires cytoskeletal protein FilP in oxygen-limiting conditions

Ultee

Briegel

Claessen

2020

Preprint

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13The cell wall is considered an essential component for bacterial survival, providing structural support 14 and protection from environmental insults. Under normal growth conditions, filamentous 15 actinobacteria insert new cell wall material at the hyphal tips regulated by the coordinated activity of 16 cytoskeletal proteins and cell wall biosynthetic enzymes. Despite the importance of the cell wall, some 17 filamentous actinobacteria can produce wall-deficient S-cells upon prolonged exposure to 18 hyperosmotic stress. Here we performed cryo-electron tomography and live cell imaging to further 19 characterize S-cell extrusion in Kitasatospora viridifaciens. We show that exposure to hyperosmotic 20 stress leads to DNA compaction, membrane and S-cell extrusion and thinning of the cell wall at 21 hyphal tips. Additionally, we find that the extrusion of S-cells is abolished in a cytoskeletal mutant 22 strain that lacks the intermediate filament-like protein FilP. Furthermore, micro-aerobic culturing 23 promotes the formation of S-cells in the wild-type, but the limited oxygen still impedes S-cell 24 formation in the ΔfilP mutant. These results demonstrate that S-cell formation is stimulated by 25 oxygen-limiting conditions and dependent on the presence of an intact cytoskeleton. 26 27

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“…We previously sequenced K. viridifaciens and identified KVP1 as a novel megaplasmid [26]. Analysis of the biosynthetic gene clusters (BGCs) using antiSMASH 5.0 [27] located 11 BGCs on KVP1 and 33 clusters on the chromosome (Fig.…”

Section: Resultsmentioning

confidence: 99%

Genome rearrangements and megaplasmid loss in the filamentous bacteriumKitasatospora viridifaciensare associated with protoplast formation and regeneration

Ramijan

Zhang

Wezel

et al. 2019

Preprint

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20Filamentous Actinobacteria are multicellular bacteria with linear replicons. Kitasatospora 21 viridifaciens DSM 40239 contains a linear 7.8 Mb chromosome and an autonomously 22 replicating plasmid KVP1 of 1.7 Mb. Here we show that lysozyme-induced protoplast 23 formation of the multinucleated mycelium of K. viridifaciens drives morphological diversity. 24Characterization and sequencing of an individual revertant colony that had lost the ability to 25 differentiate revealed that the strain had not only lost most of KVP1 but also carried lesions 26 in the right arm of the chromosome. Strikingly, the lesion sites were preceded by insertion 27 sequence elements, suggesting that the rearrangements may have been caused by 28 replicative transposition and homologous recombination between both replicons. These 29 data indicate that protoplast formation is a stressful process that can lead to profound 30 genetic changes. 31 34 metabolites are mostly used as weapons that provide protection against other 35 microorganisms and phages in the environment [1][2][3]. This is particularly useful for 36 filamentous organisms, given that they generally lack the ability to make flagella for escaping 37 dangerous situations. In addition, these bacteria are able to generate resistant spores that 38 can invade new environments after their dispersal. Germination of spores leads to the 39 formation of 1-2 germ tubes, which grow by tip extension, thereby establishing filamentous 40 cells called hyphae. Branching of hyphae leads to the formation of a multinucleated 41 vegetative mycelium, which forages and acquires nutrients by decomposing polymeric 42 substances. Stressful conditions (such as nutrient depletion) induce program cell death 43 (PCD) of the mycelium, which in turn triggers morphological and chemical differentiation [4].44 This developmental transition leads to the formation of specialized hyphae that grow into the 45 air, and the onset of production of a suite of bioactive compounds [5]. Eventually, the aerial 46 hyphae metamorphose into chains of grey-pigmented spores. Mutants that are unable to 47 establish an aerial mycelium are called bald (bld), while those that are not capable to form 48 spores are called white (whi) after their whitish color [6, 7]. 49Genome mining has been instrumental for the revival of drug discovery [8, 9]. Many 50 of the biosynthetic gene clusters that specify bioactive natural products are contained on 51 giant linear plasmids [10-13]. Although linear replicons are rare in many bacterial taxa, they 52 are common in Actinobacteria [14, 15]. In fact, Streptomyces chromosomes (between 8 and 53 10 Mb in size) are also linear and typically comprise a "core region" containing the essential 54 genes, and two variable "arms" with lengths ranging from 1.5 Mb to 2.3 Mb [16]. Like linear 55 plasmids, the linear chromosomes are capped by terminal proteins bound to the 5' end of 56 the DNA [17]. The chromosomal ends are genetically unstable, and readily undergo large 57 (up to 2 Mb) DNA rearrangements. Such...

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Genome Sequence of the Filamentous Actinomycete Kitasatospora viridifaciens

Cited by 12 publications

References 12 publications

Stress-induced formation of cell wall-deficient cells in filamentous actinomycetes

Stress-induced formation of cell wall-deficient cells in filamentous actinomycetes

Formation of wall-less cells inKitasatospora viridifaciensrequires cytoskeletal protein FilP in oxygen-limiting conditions

Genome rearrangements and megaplasmid loss in the filamentous bacteriumKitasatospora viridifaciensare associated with protoplast formation and regeneration

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