MINIREVIEW
Conjugative DNA transfer in Streptomyces by TraB: is one
protein enough?
€ nther Muth
Lina Thoma & Gu
€r Mikrobiologie und Infektionsmedizin Tu
€bingen IMIT, Eberhard Karls Universit€
€bingen,
Mikrobiologie/Biotechnologie, Interfakult€ares Institut fu
at Tu
€bingen, Germany
Tu
€nther Muth,
Correspondence: Gu
Mikrobiologie/Biotechnologie, Interfakult€ares
€r Mikrobiologie und
Institut fu
€bingen IMIT, Eberhard
Infektionsmedizin Tu
€bingen, Auf der
Karls Universit€
at Tu
€bingen, Germany.
Morgenstelle 28, 72076 Tu
Tel.: +4970712974637; fax: +497071295979;
e-mail: [email protected]
Received 17 September 2012; revised 12
October 2012; accepted 15 October 2012.
Final version published online November
2012.
DOI: 10.1111/1574-6968.12031
Abstract
Antibiotic-producing soil bacteria of the genus Streptomyces form a huge natural reservoir of antibiotic resistance genes for the dissemination within the soil
community. Streptomyces plasmids encode a unique conjugative DNA transfer
system clearly distinguished from classical conjugation involving a singlestranded DNA molecule and a type IV protein secretion system. Only a single
plasmid–encoded protein, TraB, is sufficient to translocate a double-stranded
DNA molecule into the recipient in Streptomyces matings. TraB is a hexameric
pore-forming ATPase that resembles the chromosome segregator protein FtsK
and translocates DNA by recognizing specific 8-bp repeats present in the plasmid clt locus. Mobilization of chromosomal genes does not require integration
of the plasmid, because TraB also recognizes clt-like sequences distributed all
over the chromosome.
MICROBIOLOGY LETTERS
Editor: Klaus Hantke
Keywords
Streptomyces; TraB; FtsK; plasmid transfer;
DNA translocation; chromosome
mobilization.
Introduction
Mycelium-forming actinomycetes do not divide by binary
fission but grow by apical tip extension and undergo a complex life cycle ending in sporulation (Flardh & Buttner,
2009). They are well known for the production of antibiotics, a feature probably developed to inhibit competitors in
the soil community (Allen et al., 2010). During evolution of
the antibiotic biosynthetic gene clusters, they also evolved
specific resistance genes as a part of the cluster to protect
themselves from their own compounds. Because a typical
Streptomyces strain contains 10–20 different gene clusters for
the production of antibiotics and other bioactive secondary
metabolites (Bentley et al., 2002; Medema et al., 2011),
streptomycetes form a huge reservoir of antibiotic resistance
genes in the soil, which can be passed to other bacteria by
horizontal gene transfer (D’Costa et al., 2006; Allen et al.,
2010). Therefore, the antibiotic producers not only compete
with other organisms by the production of antimicrobial
FEMS Microbiol Lett 337 (2012) 81–88
compounds but they also provide resistance genes that can
help others to survive.
In Streptomyces and related actinomycetes, even small
multi-copy plasmids of < 10 kb in size are normally selftransmissible and able to mobilize chromosomal
resistance genes and auxotrophic markers (Kieser et al.,
1982; Kataoka et al., 1991; Servin-Gonzalez et al., 1995).
These plasmids are normally cryptic and do not confer
phenotypic traits (Hopwood & Kieser, 1993; Vogelmann
et al., 2011b). Efficiency of transfer reaches nearly 100%
and between 0.1% and 1% of the transconjugants obtain
chromosomal fragments during mating (Kieser et al.,
1982). DNA transfer takes place only on solid surfaces in
the early growth phase of the life cycle, when Streptomyces
grows as substrate mycelium (Pettis & Cohen, 1996; Possoz et al., 2001). The transfer determinants of many Streptomyces plasmids were initially identified as killing
functions (kilA, traB), which could only be subcloned in
the presence of the corresponding killing override (korA,
ª 2012 Federation of European Microbiological Societies
Published by Blackwell Publishing Ltd. All rights reserved
82
L. Thoma & G. Muth
(a)
(b)
(c)
Fig. 1. Pocks caused by derivatives of the Streptomyces lividans plasmid pIJ101. A total of 105 spores of plasmid-free S. lividans TK23::pSET152
were plated on R5 plates. Subsequently, dilutions of plasmid carrying S. lividans TK64 spores were streaked. After 2 days of incubation at 30 °C
(a), pock structures develop, which are associated with the induction of the blue-red pigmented antibiotic actinorhodin. After full sporulation (b),
the plates were replica plated to LBapramycin/thiostrepton (c) to select for transconjugants.
traR) region (Kendall & Cohen, 1987; Hagege et al., 1993;
Reuther et al., 2006a). Probably due to the toxic effects of
the transfer determinants, plasmid transfer is associated
with the formation of so-called pock structures having
a diameter of 1–3 mm. Pocks are formed when donor
spores germinate on a lawn of a plasmid-free recipient.
Pocks represent temporally retarded growth inhibition
zones and indicate the area, where the recipient mycelium
has obtained a plasmid by conjugation (Fig. 1). Formation of pock structures has been interpreted as the result
of intramycelial plasmid spreading via the septal crosswalls of the recipient mycelium (Hopwood & Kieser,
1993; Grohmann et al., 2003).
Conjugative transfer of a doublestranded plasmid molecule requires
TraB and the noncoding clt region
The small size of the plasmid region determining conjugative transfer already indicated that the Streptomyces
DNA transfer mechanism must differ considerably from
the known conjugation systems of other bacteria, involving a conjugative relaxase and a complex type IV protein
secretion system (Chen et al., 2005; de la Cruz et al.,
2010). Characterization of several Streptomyces plasmids
by subcloning and linker insertions revealed a plasmid
region of about 3 kb being essential for transfer, while
the adjacent region affected only the size of the pock
structures (Kieser et al., 1982; Kataoka et al., 1991; Servin-Gonzalez et al., 1995; Reuther et al., 2006a). When
the nucleotide sequence of the Streptomyces lividans plasmid pIJ101 was available (Kendall & Cohen, 1988) as the
first complete sequence of a conjugative plasmid from a
Gram-positive bacterium, it was realized that korA (traR)
encoded a transcriptional regulator of the GntR family,
while a small region of the KilA (TraB) protein showed
some similarity to the FtsK protein involved in cell division and chromosome segregation (Begg et al., 1995; Wu
et al., 1995; Sherratt et al., 2010).
ª 2012 Federation of European Microbiological Societies
Published by Blackwell Publishing Ltd. All rights reserved
Pettis & Cohen (1994) demonstrated that beside the
TraB protein, a small non-coding plasmid region of about
50 bp was required for the transfer of plasmid pIJ101, the
cis-acting-locus of transfer (clt). When clt was inserted
into a nontransferable plasmid, this plasmid could be
mobilized, if TraB was provided in trans. Interestingly, clt
was only required for plasmid transfer but was dispensable for the mobilization of chromosomal markers (Pettis
& Cohen, 1994), indicating that clt does not represent a
classical origin of transfer (oriT). The clt regions of
different Streptomyces plasmids do not show any sequence
similarity, but often contain repetitive sequences that have
the ability to form secondary structures (Franco et al.,
2003; Vogelmann et al., 2011a).
The first experimental evidence on the novel mechanism
of the Streptomyces conjugative DNA transfer system came
from the work of Possoz et al. (2001) by demonstrating
that conjugative transfer of the Streptomyces ambofaciens
plasmid pSAM2 was sensitive to the presence of the SalI
restriction/modification system in the recipient. In this
study, a pSAM2 derivative could not be transferred into
S. lividans TK23 expressing SalI, whereas pSAM2 was
efficiently transferred to TK23 lacking the SalI restriction
system. Because the transferred DNA was obviously
degraded by SalI and because SalI recognizes only doublestranded DNA as substrate but not single-stranded DNA,
the incoming DNA must be double-stranded.
TraB is a pore-forming hexameric ring
ATPase homologous to FtsK
Multiple alignment of TraB homologues from various
Streptomyces plasmids revealed a highly diverse family of
proteins showing only very limited sequence similarity,
in part < 20%. However, secondary structure predictions
revealed an identical domain architecture for all TraB
homologues, which resembles that of FtsK: a N-terminal
membrane association domain that is followed by a
DNA-translocase/ATPase domain with Walker A and B
FEMS Microbiol Lett 337 (2012) 81–88
83
Conjugative DNA transfer in Streptomyces
(a)
(b)
Fig. 2. Mobilization of chromosomal markers. (a) In most bacteria, the conjugative plasmid integrates into the chromosome at specific positions
by homologous recombination between IS elements (black boxes) generating a HFR strain. The plasmid-encoded relaxase (red arrow) initiates
rolling-circle replication at the oriT (red circle) and guides the single-stranded DNA molecule as a pilot protein to the plasmid-encoded type IV
protein secretion system (grey arrows). (b) In Streptomyces, the translocase TraB (blue arrow) binds noncovalently to the clt locus (blue circle) and
transfers a double-stranded plasmid molecule. Because TraB also recognizes (dotted arrows) clt-like chromosomal sequences (light blue) that are
distributed all over the chromosome, TraB can direct transfer of chromosomal markers not relying on previous plasmid integration.
boxes and a C-terminal winged helix-turn-helix fold
(wHTH) (Vogelmann et al., 2011a). ATPase activity and
membrane association have been experimentally confirmed for TraB proteins of various plasmids (Kosono
et al., 1996; Pettis & Cohen, 1996; Reuther et al.,
2006b). Inactivation of the ATP binding site of TraB
from the Streptomyces nigrifaciens plasmid pSN22
demonstrated that the ATPase activity is essential for
conjugative transfer (Kosono et al., 1996).
The similarity of TraB to the septal DNA translocator
proteins FtsK and SpoIIIE that direct chromosome segregation during cell division and sporulation (Bath et al.,
2000; Massey et al., 2006; Bigot et al., 2007) suggests a
similar function for TraB during conjugation. However,
whereas FtsK translocates the DNA through a closing septum to the daughter cell/spore, TraB has to translocate
the DNA through intact cell envelopes of the donor and
the recipient. Because a TraB–eGFP fusion protein localized to the hyphal tips of substrate mycelium, it was
suggested that Streptomyces conjugation involves the tips
(Reuther et al., 2006b). Up to now it is still unclear,
whether TraB contains a membrane-targeting sequence
and is directed to the tip by the membrane composition
or curvature or whether TraB is recruited to the tips by
its interaction with other proteins, for example, DivIVA
(Hempel et al., 2008; Lenarcic et al., 2009; Jyothikumar
et al., 2012).
Despite the toxic effects of TraB, this protein of the
Streptomyces venezuelae plasmid pSVH1 could be
expressed in S. lividans with an N-terminal Strep tagII
sequence (Voss & Skerra, 1997) and purified (Reuther
et al., 2006b). Chemical crosslinking showed higher oligoFEMS Microbiol Lett 337 (2012) 81–88
meric structures that were also observed when the membrane association domain of TraB was eliminated. After
separation of TraB oligomers from the monomer fraction
by gel filtration chromatography, ring-shaped TraB particles could be detected by electron microscopy. 2D averaging of the images revealed symmetric hexamers of about
12 nm in diameter, which contained a central pore. This
structure was in full agreement with a predicted TraBDNA-translocase structure obtained by homology modelling with the Pseudomonas aeruginosa FtsK translocase
domain crystal structure as a template (Vogelmann et al.,
2011a). Both structures had a central pore of 3.0
and 3.1 nm, respectively, which is of sufficient size to
accommodate a double-stranded DNA molecule.
Conjugative transfer of DNA by direct cell to cell contact implies that the DNA has to pass the cell envelopes
of donor and recipient. For Streptomyces, this means: two
cytoplasmic membranes and two peptidoglycan (PG) layers. PG-binding assays of purified TraB showed that TraB
interacts with PG (Vogelmann et al., 2011a). For the
membrane passage, one has to postulate a pore structure
for TraB. This is in contrast to Escherichia coli FtsK that
probably translocates the chromosome before closure of
the septum and therefore does not rely on a pore-forming
ability (Dubarry & Barre, 2010). The ability of TraB to
form pore structures was analysed by single channel
recordings using planar lipid bilayers. These studies demonstrated that TraB spontaneously inserted into the membrane at various voltages and formed pores with an
opening time of about 47–81 ms (positive voltage
applied) and 105–200 ms, respectively, when a negative
voltage was applied (Vogelmann et al., 2011a).
ª 2012 Federation of European Microbiological Societies
Published by Blackwell Publishing Ltd. All rights reserved
84
L. Thoma & G. Muth
Fig. 3. Model of conjugative plasmid transfer
in Streptomyces. Mycelial tips of donor and
recipient find each other on solid medium
without involvement of a plasmid-encoded
aggregation system. An unknown signal
released by the recipient probably induces
expression of traB in the donor. TraB recruits
proteins of the PG synthesis/remodelling
machinery to direct fusion of the PG layers at
the hyphal tips. TraB hexamers assemble at
the plasmid clt locus and form under partial
fusion of the membranes a pore structure to
the recipient. Under ATP hydrolysis, doublestranded plasmid DNA is translocated. This
step might require further chromosomally
encoded proteins, for example, a
topoisomerase. Following the primary transfer,
the newly transferred DNA spreads via the
septal cross-walls to the neighbouring
compartments resulting in a rapid colonization
of the recipient mycelium with the incoming
plasmid. Plasmid spreading involves various
plasmid-encoded Spd proteins in addition to
TraB.
TraB specifically recognizes 8-bp repeats
in the plasmid clt region
Because only TraB and the non-coding clt region are
required for plasmid transfer, it was studied whether clt
represents the binding site of TraB. This hypothesis turned
out to be correct, because gel retardation assays showed a
specific interaction of TraB with a plasmid region at the
3′ end of traB, which represents the clt region of pSVH1
(Reuther et al., 2006b). The pSVH1 clt region contained
nine imperfectly conserved copies of the GACCCGGA
motif. Subcloning experiments revealed a minimal fragment containing only four copies, which still supported
TraB binding. A more careful analysis detected even binding of TraB to a synthetic 20-bp fragment containing only
ª 2012 Federation of European Microbiological Societies
Published by Blackwell Publishing Ltd. All rights reserved
two copies (Vogelmann et al., 2011a). This study confirmed the GACCCGGA motif as the TraB Recognition
Sequence (TRS). Although two copies of TRS were sufficient for TraB binding in vitro, binding of TraB to a larger
clt fragment containing additional TRS copies was more
efficient and required lower protein concentrations for
retardation (Reuther et al., 2006b) indicating that in vivo
only the complete clt might be effective.
Analysing other Streptomyces plasmids for the presence
of 8-bp repeats also detected specific 8-bp repeats in the
(predicted) clt regions (Franco et al., 2003; Vogelmann
et al., 2011a). With the notable exceptions of pIJ101
(Kieser et al., 1982) and the highly related plasmid p1424
(G. Muth, unpublished), the clt localizes in all Streptomyces
plasmids to the 3′ end of traB, forming a transfer
FEMS Microbiol Lett 337 (2012) 81–88
85
Conjugative DNA transfer in Streptomyces
module of only 2.5 kb in size consisting of the DNAtranslocase-encoding traB gene and its binding site clt
next to it.
To characterize the TraB–clt interaction in more detail,
TraB was incubated with covalently closed circular (ccc)
DNA of the pSVH1 derivative pEB211 in the presence of
ATP and divalent cations. An aliquot was directly loaded
to the gel, while others were heat treated or phenol
extracted to denature TraB previous to gel loading. These
analyses revealed ccc-DNA that had not changed its
conformation demonstrating that TraB binds noncovalently to plasmid DNA and that the plasmid molecule was not
processed by TraB binding (Reuther et al., 2006b).
Because the clt loci of different plasmids contain different 8-bp TRS, it was possible to map the TraB
region determining specific TRS recognition by constructing a series of chimeric proteins and analysing
their DNA binding activities in gel retardation experiments with different clt loci. These studies identified the
very C-terminal end of TraB forming a wHTH fold as
being responsible for clt recognition. Further studies
even narrowed down the TRS recognition region to
helix a3 of the wHTH fold. Exchange of only 13 aa of
TraBpSVH1 against the 13 aa corresponding to helix a3
of TraBpIJ101 switched clt recognition. The chimeric protein was no longer able to bind to the clt of pSVH1
but shifted the clt fragment of pIJ101 (Vogelmann et al.,
2011a).
Colonization of the recipient by
intramycelial plasmid spreading
requires additional plasmid-encoded
proteins
Generation of pock structures during Streptomyces conjugation has been interpreted as the result of intramycelial
plasmid spreading following the primary DNA transfer
from a donor into the recipient (Hopwood & Kieser,
1993; Grohmann et al., 2003). Whereas plasmid transfer
from a donor into the recipient requires only TraB, plasmid spreading involves five to seven plasmid-encoded
proteins (Spd) in addition to TraB. This probably reflects
the challenge to cross the septal cross-walls. The Spd proteins have no significant similarity to any functionally
characterized protein complicating prediction of their
putative function. Inactivation of a single spd gene
reduces the size of the pock structures (Kieser et al.,
1982; Kataoka et al., 1994; Servin-Gonzalez et al., 1995;
Reuther et al., 2006a). Only few reports address the biochemical characterization of the Spd proteins and their
molecular function is more or less unknown. Genetic
organization of the spd genes with overlapping stop and
start codons, analysis of protein–protein interaction by
FEMS Microbiol Lett 337 (2012) 81–88
chemical crosslinking, bacterial two-hybrid analysis or
copurification experiments indicated that the Spd proteins form a multiprotein complex with TraB (Tiffert
et al., 2007) (Thoma, Guezguez and Muth, unpublished).
Intramycelial plasmid spreading might also contribute
to the stable maintenance of Streptomyces plasmids,
because hyphal compartments that have lost a plasmid
can recover a plasmid from the neighbouring compartment. In agreement with this hypothesis, a clear effect of
spd1 inactivation on stable maintenance of the linear plasmid
SLP2 was reported (Hsu & Chen, 2010).
Conjugative Streptomyces plasmids
contribute in different ways to the
evolution of the genomes
Streptomyces plasmids contribute to the evolution and
shaping of the chromosome in different ways (Medema
et al., 2010). Linear plasmids can recombine with the
chromosome. Because the Streptomyces chromosome is
normally linear (Lin et al., 1993), this results in the
exchange of the ends, creating plasmids that carry chromosomal DNA. These plasmids can be transferred by
conjugation to new Streptomyces species, where they can
replicate either autonomously or recombine again with
the chromosome.
But also circular plasmids have been reported to mobilize chromosomal fragments with high efficiency (Kieser
et al., 1982; Hopwood & Kieser, 1993). However, the
mechanism of chromosome mobilization differs fundamentally from the one reported for other bacteria. Classical High Frequency of Recombination strains (HFR) carry
the conjugative plasmid at a specific location in the
chromosome (Thomas & Nielsen, 2005). Plasmid integration normally occurred via homologous recombination
between IS elements. Initiation of rolling-circle replication
at the plasmid oriT by the conjugative relaxase creates a
linear single-stranded DNA molecule that contains plasmid sequences followed by the chromosomal loci next to
the integration site. This strand is guided by the covalently
bound relaxase to the recipient, where it can recombine
with the chromosome (de la Cruz et al., 2010).
Because the Streptomyces DNA-translocase TraB does
not have a relaxase activity and most probably does not
process the DNA (Reuther et al., 2006b) and because clt
is dispensable for the transfer of chromosomal markers
(Pettis & Cohen, 1994), the chromosome mobilization
mechanism in Streptomyces must be different (Fig. 2). An
explanation provides the finding that TraB recognizes
8-bp TRS motifs and that clt-like sequences containing
repeated TRS are frequently found in Streptomyces
chromosomes (Vogelmann et al., 2011a). Analysis of the
Streptomyces coelicolor genomic sequence for pSVH1 cltª 2012 Federation of European Microbiological Societies
Published by Blackwell Publishing Ltd. All rights reserved
86
like sequences (four copies of the TRS GACCCGGA with
a spacing of up to 13 bp, allowing one mismatch) identified 25 hits. These sequences are not part of integrated
plasmids or represent remnants of plasmids, but are often
located within genes without disrupting their coding
region. These insertions are only found in the respective
S. coelicolor genes but not in the corresponding homologues of Streptomyces avermitilis or those of other Streptomyces species, which carry clt-like sequences on other
locations (Sepulveda et al., 2011). This demonstrates that
these insertions have been acquired later and are probably
not involved in the respective enzymatic activities. It is
unclear how these insertions have been generated. But
with respect to the prevalence of plasmids in Streptomyces, one can speculate that there is an adaptive selection
for clt-like sequences in Streptomyces genomes to benefit
from the presence of conjugative plasmids.
Concluding remarks/open questions
Pettis & Cohen (1994) clearly demonstrated that TraB is
the only plasmid-encoded protein required for conjugative
transfer of pIJ101. Similarity of TraB to the chromosome
segregator proteins FtsK or SpoIIIE suggests a conjugative
DNA translocation mechanism for the transfer between a
donor and a recipient mycelium that resembles the intracellular segregation of chromosomal DNA during cell division and sporulation. TraB hexamers probably assemble at
the plasmid localized clt or, with lower efficiency, at chromosomal clt-like sequences. These hexamers form pore
structures in the membrane, which act as molecular
motors, energized by ATP hydrolysis and translocate double-stranded DNA to the recipient (Fig. 3).
However, this simplified model has drawbacks and
leaves several open questions. To fully understand TraBmediated conjugative DNA transfer, one has to propose
additional enzymatic activities for TraB waiting to be
uncovered. Alternatively, TraB might recruit other chromosomally encoded proteins for the transfer process.
(1) How to cross the PG barrier?
A TraB–eGFP fusion was localized at the hyphal tip, suggesting that the tips of the mycelium are involved in conjugation (Reuther et al., 2006b). Also, TraB was shown to
bind isolated PG (Vogelmann et al., 2011a). Because TraB
itself does not have a PG-lysing activity (Finger and Muth,
unpublished), it is possible that TraB interacts with chromosomally encoded PG hydrolases at the tip to direct
fusion of the PG layers of donor and recipient.
(2) How to cross membranes of donor and recipient?
In contrast to FtsK that is found in both compartments
during cell division, TraB is present only in the donor
mycelium. Therefore, the TraB pore has to traverse two
membranes (one from the donor, one from the recipient)
ª 2012 Federation of European Microbiological Societies
Published by Blackwell Publishing Ltd. All rights reserved
L. Thoma & G. Muth
or the two membranes have to fuse. For SpoIIIE that
mediates translocation of the chromosome into the forespore during Bacillus sporulation, a membrane fusing
activity has been reported (Sharp & Pogliano, 2003).
Therefore, it is tempting to speculate that also TraB
might have a membrane fusing activity allowing formation of a pore structure to the recipient.
(3) How to translocate a circular covalently closed
plasmid molecule?
During cell division or sporulation, the septum closes,
while chromosomal DNA is already present, allowing
FtsK to assemble at both chromosomal arms to translocate the DNA. DNA translocation causes topological
stress to the DNA, which has to be relieved by topoisomerases. The interaction of E. coli FtsK with topoisomerase IV has been reported (Espeli et al., 2003). However,
it is still unclear, how the remaining end of the circular
chromosome becomes translocated through the membrane and fusion of the two FtsK hexamer structures has
been postulated (Burton et al., 2007).
During Streptomyces conjugation, the situation is even
more complex. The translocase TraB is definitely present
only on the donor site of the mating hyphae, and a mechanism translocating a circular double-stranded DNA molecule is not very plausible. Because the plasmid DNA is not
processed during TraB binding at clt, one has to propose
involvement of an additional enzymatic activity, for example, a topoisomerase, which might produce a linear molecule that can be transported through the TraB pore.
(4) How to pass the septal cross-walls in the recipient
mycelium?
Crossing the septal cross-walls during intramycelial
plasmid spreading seems to be an even more challenging
task compared to the primary DNA transfer at the hyphal
tip. It involves, in addition to TraB, several Spd proteins.
The structure of the Streptomyces septal cross-walls has
not been elucidated, and it is not clear whether preexisting channel structures in the cross-walls connect the compartments of the substrate mycelium (Jakimowicz & van
Wezel, 2012). The Spd proteins might interact to build a
cross-wall traversing complex, allowing translocation of
the plasmid by the motor protein TraB.
(5) Is TraB able to promote intergeneric DNA transfer?
The capability of the T4SS conjugation system to transfer plasmids between distantly related bacteria, even across
kingdoms, is well documented (Bates et al., 1998; Thomas
& Nielsen, 2005). Although conjugative transfer of Streptomyces plasmids between different Streptomyces species has
been observed (Hopwood & Kieser, 1993), conjugative
transfer to other bacteria has not been reported. Therefore,
the relevance of the Streptomyces conjugative DNA transfer
system in the dissemination of the Streptomyces reservoir
of resistance genes is still concealed.
FEMS Microbiol Lett 337 (2012) 81–88
Conjugative DNA transfer in Streptomyces
Acknowledgements
We thank the DFG (SFB766) for financial support.
References
Allen HK, Donato J, Wang HH, Cloud-Hansen KA, Davies J
& Handelsman J (2010) Call of the wild: antibiotic
resistance genes in natural environments. Nat Rev Microbiol
8: 251–259.
Bates S, Cashmore AM & Wilkins BM (1998) IncP plasmids
are unusually effective in mediating conjugation of
Escherichia coli and Saccharomyces cerevisiae: involvement of
the tra2 mating system. J Bacteriol 180: 6538–6543.
Bath J, Wu LJ, Errington J & Wang JC (2000) Role of Bacillus
subtilis SpoIIIE in DNA transport across the mother
cell-prespore division septum. Science 290: 995–997.
Begg KJ, Dewar SJ & Donachie WD (1995) A new Escherichia
coli cell division gene, ftsK. J Bacteriol 177: 6211–6222.
Bentley SD, Chater KF, Cerdeno-Tarraga AM et al. (2002)
Complete genome sequence of the model actinomycete
Streptomyces coelicolor A3(2). Nature 417: 141–147.
Bigot S, Sivanathan V, Possoz C, Barre FX & Cornet F (2007)
FtsK, a literate chromosome segregation machine. Mol
Microbiol 64: 1434–1441.
Burton BM, Marquis KA, Sullivan NL, Rapoport TA & Rudner
DZ (2007) The ATPase SpoIIIE transports DNA across
fused septal membranes during sporulation in Bacillus
subtilis. Cell 131: 1301–1312.
Chen I, Christie PJ & Dubnau D (2005) The ins and outs of
DNA transfer in bacteria. Science 310: 1456–1460.
D’Costa VM, McGrann KM, Hughes DW & Wright GD (2006)
Sampling the antibiotic resistome. Science 311: 374–377.
de la Cruz F, Frost LS, Meyer RJ & Zechner EL (2010)
Conjugative DNA metabolism in Gram-negative bacteria.
FEMS Microbiol Rev 34: 18–40.
Dubarry N & Barre FX (2010) Fully efficient chromosome
dimer resolution in Escherichia coli cells lacking the integral
membrane domain of FtsK. EMBO J 29: 597–605.
Espeli O, Lee C & Marians KJ (2003) A physical and
functional interaction between Escherichia coli FtsK and
topoisomerase IV. J Biol Chem 278: 44639–44644.
Flardh K & Buttner MJ (2009) Streptomyces morphogenetics:
dissecting differentiation in a filamentous bacterium.
Nat Rev Microbiol 7: 36–49.
Franco B, Gonzalez-Ceron G & Servin-Gonzalez L (2003)
Direct repeat sequences are essential for function of the
cis-acting locus of transfer (clt) of Streptomyces
phaeochromogenes plasmid pJV1. Plasmid 50: 242–247.
Grohmann E, Muth G & Espinosa M (2003) Conjugative
plasmid transfer in gram-positive bacteria. Microbiol Mol
Biol Rev 67: 277–301.
Hagege J, Pernodet JL, Sezonov G, Gerbaud C, Friedmann A &
Guerineau M (1993) Transfer functions of the conjugative
integrating element pSAM2 from Streptomyces ambofaciens:
FEMS Microbiol Lett 337 (2012) 81–88
87
characterization of a kil-kor system associated with transfer.
J Bacteriol 175: 5529–5538.
Hempel AM, Wang SB, Letek M, Gil JA & Flardh K (2008)
Assemblies of DivIVA mark sites for hyphal branching and
can establish new zones of cell wall growth in Streptomyces
coelicolor. J Bacteriol 190: 7579–7583.
Hopwood DA & Kieser T (1993) Conjugative Plasmids of
Streptomyces. Bacterial conjugation (Clewell DB, ed),
pp. 293–311. Plenum Press, New York.
Hsu CC & Chen CW (2010) Linear plasmid SLP2 is
maintained by partitioning, intrahyphal spread, and
conjugal transfer in Streptomyces. J Bacteriol 192:
307–315.
Jakimowicz D & van Wezel GP (2012) Cell division and DNA
segregation in Streptomyces: how to build a septum in the
middle of nowhere? Mol Microbiol 85: 393–404.
Jyothikumar V, Klanbut K, Tiong J, Roxburgh JS, Hunter IS,
Smith TK & Herron PR (2012) Cardiolipin synthase is
required for Streptomyces coelicolor morphogenesis. Mol
Microbiol 84: 181–197.
Kataoka M, Seki T & Yoshida T (1991) Five genes involved in
self-transmission of pSN22, a Streptomyces plasmid.
J Bacteriol 173: 4220–4228.
Kataoka M, Kiyose YM, Michisuji Y, Horiguchi T, Seki T &
Yoshida T (1994) Complete nucleotide sequence of the
Streptomyces nigrifaciens plasmid, pSN22: genetic
organization and correlation with genetic properties.
Plasmid 32: 55–69.
Kendall KJ & Cohen SN (1987) Plasmid transfer in
Streptomyces lividans: identification of a kil-kor system
associated with the transfer region of pIJ101. J Bacteriol 169:
4177–4183.
Kendall KJ & Cohen SN (1988) Complete nucleotide sequence of
the Streptomyces lividans plasmid pIJ101 and correlation of the
sequence with genetic properties. J Bacteriol 170: 4634–4651.
Kieser T, Hopwood DA, Wright HM & Thompson CJ (1982)
pIJ101, a multi-copy broad host-range Streptomyces plasmid:
functional analysis and development of DNA cloning
vectors. Mol Gen Genet 185: 223–228.
Kosono S, Kataoka M, Seki T & Yoshida T (1996) The TraB
protein, which mediates the intermycelial transfer of the
Streptomyces plasmid pSN22, has functional NTP-binding
motifs and is localized to the cytoplasmic membrane. Mol
Microbiol 19: 397–405.
Lenarcic R, Halbedel S, Visser L et al. (2009) Localisation of
DivIVA by targeting to negatively curved membranes.
EMBO J 28: 2272–2282.
Lin YS, Kieser HM, Hopwood DA & Chen CW (1993) The
chromosomal DNA of Streptomyces lividans 66 is linear. Mol
Microbiol 14: 1103.
Massey TH, Mercogliano CP, Yates J, Sherratt DJ & Lowe J
(2006) Double-stranded DNA translocation: structure and
mechanism of hexameric FtsK. Mol Cell 23: 457–469.
Medema MH, Trefzer A, Kovalchuk A et al. (2010) The
sequence of a 1.8-mb bacterial linear plasmid reveals a rich
ª 2012 Federation of European Microbiological Societies
Published by Blackwell Publishing Ltd. All rights reserved
88
evolutionary reservoir of secondary metabolic pathways.
Genome Biol Evol 2: 212–224.
Medema MH, Blin K, Cimermancic P, de Jager V, Zakrzewski P,
Fischbach MA, Weber T, Takano E & Breitling R (2011)
antiSMASH: rapid identification, annotation and analysis
of secondary metabolite biosynthesis gene clusters in bacterial
and fungal genome sequences. Nucleic Acids Res 39: W339–
W346.
Pettis GS & Cohen SN (1994) Transfer of the plJ101 plasmid
in Streptomyces lividans requires a cis-acting function
dispensable for chromosomal gene transfer. Mol Microbiol
13: 955–964.
Pettis GS & Cohen SN (1996) Plasmid transfer and expression
of the transfer (tra) gene product of plasmid pIJ101 are
temporally regulated during the Streptomyces lividans life
cycle. Mol Microbiol 19: 1127–1135.
Possoz C, Ribard C, Gagnat J, Pernodet JL & Guerineau M (2001)
The integrative element pSAM2 from Streptomyces: kinetics
and mode of conjugal transfer. Mol Microbiol 42: 159–166.
Reuther J, Wohlleben W & Muth G (2006a) Modular
architecture of the conjugative plasmid pSVH1 from
Streptomyces venezuelae. Plasmid 55: 201–209.
Reuther J, Gekeler C, Tiffert Y, Wohlleben W & Muth G (2006b)
Unique conjugation mechanism in mycelial streptomycetes: a
DNA-binding ATPase translocates unprocessed plasmid DNA
at the hyphal tip. Mol Microbiol 61: 436–446.
Sepulveda E, Vogelmann J & Muth G (2011) A septal
chromosome segregator protein evolved into a conjugative
DNA-translocator protein. Mob Genet Elements 1: 225–229.
Servin-Gonzalez L, Sampieri AI, Cabello J, Galvan L, Juarez V
& Castro C (1995) Sequence and functional analysis of the
Streptomyces phaeochromogenes plasmid pJV1 reveals a
ª 2012 Federation of European Microbiological Societies
Published by Blackwell Publishing Ltd. All rights reserved
L. Thoma & G. Muth
modular organization of Streptomyces plasmids that replicate
by rolling circle. Microbiology 141: 2499–2510.
Sharp MD & Pogliano K (2003) The membrane domain of
SpoIIIE is required for membrane fusion during Bacillus
subtilis sporulation. J Bacteriol 185: 2005–2008.
Sherratt DJ, Arciszewska LK, Crozat E, Graham JE & Grainge I
(2010) The Escherichia coli DNA translocase FtsK. Biochem
Soc Trans 38: 395–398.
Thomas CM & Nielsen KM (2005) Mechanisms of, and
barriers to, horizontal gene transfer between bacteria. Nat
Rev Microbiol 3: 711–721.
Tiffert Y, Gotz B, Reuther J, Wohlleben W & Muth G (2007)
Conjugative DNA transfer in Streptomyces: SpdB2 involved
in the intramycelial spreading of plasmid pSVH1 is an
oligomeric integral membrane protein that binds to dsDNA.
Microbiology 153: 2976–2983.
Vogelmann J, Ammelburg M, Finger C et al. (2011a) Conjugal
plasmid transfer in Streptomyces resembles bacterial
chromosome segregation by FtsK/SpoIIIE. EMBO J 30:
2246–2254.
Vogelmann JW, Wohlleben W & Muth G (2011b) Streptomyces
conjugative genetic elements. Streptomyces – Molecular
Biology and Biotechnology (Dyson P, ed), pp. 27–42. Caister
Academic Press, Norfolk, UK.
Voss S & Skerra A (1997) Mutagenesis of a flexible loop in
streptavidin leads to higher affinity for the Strep-tag II
peptide and improved performance in recombinant protein
purification. Protein Eng 10: 975–982.
Wu LJ, Lewis PJ, Allmansberger R, Hauser PM & Errington J
(1995) A conjugation-like mechanism for prespore
chromosome partitioning during sporulation in Bacillus
subtilis. Genes Dev 9: 1316–1326.
FEMS Microbiol Lett 337 (2012) 81–88