A bacteriophage tubulin harnesses dynamic instability to center DNA in infected cells

Erb, Marcella L.; Kraemer, James A.; Coker, Joanna; Chaikeeratisak, Vorrapon; Nonejuie, Poochit; Agard, David A.; Pogliano, Joe

doi:10.7554/elife.03197

Cited by 70 publications

(136 citation statements)

References 38 publications

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“…PhuZ assembles a unique triple stranded polymer in which protofilaments twist around each other and the C-terminal tail plays a key role in making longitudinal contacts with adjacent subunits (Zehr et al, 2014). PhuZ filaments display dynamic instability both in vitro and in vivo (Erb et al, 2014). During phage infection, PhuZ forms a bipolar spindle composed of dynamically unstable filaments that position phage DNA at midcell (Erb et al, 2014).…”

Section: Introductionmentioning

confidence: 99%

“…PhuZ filaments display dynamic instability both in vitro and in vivo (Erb et al, 2014). During phage infection, PhuZ forms a bipolar spindle composed of dynamically unstable filaments that position phage DNA at midcell (Erb et al, 2014). Mutations in the catalytic domain that eliminate PhuZ GTPase activity disrupt both spindle dynamics and phage nucleus positioning and reduce phage burst size by 50% (Erb et al, 2014; Kraemer et al, 2012).…”

Section: Introductionmentioning

confidence: 99%

“…During phage infection, PhuZ forms a bipolar spindle composed of dynamically unstable filaments that position phage DNA at midcell (Erb et al, 2014). Mutations in the catalytic domain that eliminate PhuZ GTPase activity disrupt both spindle dynamics and phage nucleus positioning and reduce phage burst size by 50% (Erb et al, 2014; Kraemer et al, 2012). The PhuZ spindle is the only known example of a cytoskeletal structure in Bacteria or Archaea that shares three key properties with the eukaryotic spindle: dynamic instability, a bipolar array of filaments, and central positioning of DNA.…”

Section: Introductionmentioning

confidence: 99%

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The Phage Nucleus and Tubulin Spindle Are Conserved among Large Pseudomonas Phages

et al. 2017

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Summary We recently demonstrated that the large Pseudomonas chlororaphis bacteriophage 201φ2-1 assembles a nucleus-like structure that encloses phage DNA and segregates proteins according to function, with DNA processing proteins inside and metabolic enzymes and ribosomes outside the nucleus. Here we investigate the replication pathway of the Pseudomonas aeruginosa bacteriophages φKZ and φPA3. Bacteriophages φKZ and φPA3 encode a proteinaceous shell that assembles a nucleus-like structure that compartmentalizes proteins and DNA during viral infection. We show that the tubulin-like protein PhuZ encoded by each phage assembles a bipolar spindle that displays dynamic instability and positions the nucleus at midcell. Our results suggest that the phage spindle and nucleus play the same functional role in all three phages, 201φ2-1, φKZ and φPA3, demonstrating that these key structures are conserved among large Pseudomonas phages.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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The Phage Nucleus and Tubulin Spindle Are Conserved among Large Pseudomonas Phages

et al. 2017

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“…BtubA/B have more sequence and structure in common with eukaryotic tubulins than any other bacterial tubulin, and the presence of both btubA and btubB genes suggests that the horizontal transfer event that is thought to have occurred not long after the initial duplication event created the ancestors of the A and B and ␣ and ␤ homolog pairs (7). The more evolutionarily distant phage tubulin PhuZ has been demonstrated to exhibit dynamic instability (32) and to segregate DNA (34). We cannot exclude the possibility that filament dynamics evolved after the horizontal gene transfer to Prosthecobacter and that any similarities between the dynamics of BtubA/B filaments, phage tubulin filaments, and eukaryotic MTs are the result of convergent evolution.…”

Section: Discussionmentioning

confidence: 99%

Bacterial Tubulins A and B Exhibit Polarized Growth, Mixed-Polarity Bundling, and Destabilization by GTP Hydrolysis

et al. 2017

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Bacteria of the genus Prosthecobacter express homologs of eukaryotic ␣-and ␤-tubulin, called BtubA and BtubB (BtubA/B), that have been observed to assemble into filaments in the presence of GTP. BtubA/B polymers are proposed to be composed in vitro by two to six protofilaments in contrast to that in vivo, where they have been reported to form 5-protofilament tubes named bacterial microtubules (bMTs). The btubAB genes likely entered the Prosthecobacter lineage via horizontal gene transfer and may be derived from an early ancestor of the modern eukaryotic microtubule (MT). Previous biochemical studies revealed that BtubA/B polymerization is reversible and that BtubA/B folding does not require chaperones. To better understand BtubA/B filament behavior and gain insight into the evolution of microtubule dynamics, we characterized in vitro BtubA/B assembly using a combination of polymerization kinetics assays and microscopy. Like eukaryotic microtubules, BtubA/B filaments exhibit polarized growth with different assembly rates at each end. GTP hydrolysis stimulated by BtubA/B polymerization drives a stochastic mechanism of filament disassembly that occurs via polymer breakage and/or fast continuous depolymerization. We also observed treadmilling (continuous addition and loss of subunits at opposite ends) of BtubA/B filament fragments. Unlike MTs, polymerization of BtubA/B requires KCl, which reduces the critical concentration for BtubA/B assembly and induces it to form stable mixed-orientation bundles in the absence of any additional BtubA/B-binding proteins. The complex dynamics that we observe in stabilized and unstabilized BtubA/B filaments may reflect common properties of an ancestral eukaryotic tubulin polymer.IMPORTANCE Microtubules are polymers within all eukaryotic cells that perform critical functions; they segregate chromosomes, organize intracellular transport, and support the flagella. These functions rely on the remarkable range of tunable dynamic behaviors of microtubules. Bacterial tubulin A and B (BtubA/B) are evolutionarily related proteins that form polymers. They are proposed to be evolved from the ancestral eukaryotic tubulin, a missing link in microtubule evolution. Using microscopy and biochemical approaches to characterize BtubA/B assembly in vitro, we observed that they exhibit complex and structurally polarized dynamic behavior like eukaryotic microtubules but differ in how they self-associate into bundles and how this bundling affects their stability. Our results demonstrate the diversity of mechanisms through which tubulin homologs promote filament dynamics and monomer turnover.

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“…Finally, bacterial FtsZ proteins, tubulin-like homologs, form short cytomotive protofilaments that function in cell division, with structures very different from those of eukaryotic MTs. However, some bacteriophages encode tubulin-like PhuZ, which forms a filamentous, spindle-like array that positions phage DNA at the bacterial cell center for optimal virus replication (144)(145)(146). These examples serve to illustrate the broader importance of MTs or MT-like filaments in infection by diverse viruses across kingdoms and species.…”

Section: Mt Functions During Infection Of Plants Insects and Bacteriamentioning

confidence: 99%

Microtubule Regulation and Function during Virus Infection

Naghavi

Walsh

2017

J Virol

105

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Microtubules (MTs) form a rapidly adaptable network of filaments that radiate throughout the cell. These dynamic arrays facilitate a wide range of cellular processes, including the capture, transport, and spatial organization of cargos and organelles, as well as changes in cell shape, polarity, and motility. Nucleating from MT-organizing centers, including but by no means limited to the centrosome, MTs undergo rapid transitions through phases of growth, pause, and catastrophe, continuously exploring and adapting to the intracellular environment. Subsets of MTs can become stabilized in response to environmental cues, acquiring distinguishing posttranslational modifications and performing discrete functions as specialized tracks for cargo trafficking. The dynamic behavior and organization of the MT array is regulated by MT-associated proteins (MAPs), which include a subset of highly specialized plus-end-tracking proteins (ϩTIPs) that respond to signaling cues to alter MT behavior. As pathogenic cargos, viruses require MTs to transport to and from their intracellular sites of replication. While interactions with and functions for MT motor proteins are well characterized and extensively reviewed for many viruses, this review focuses on MT filaments themselves. Changes in the spatial organization and dynamics of the MT array, mediated by virus-or host-induced changes to MT regulatory proteins, not only play a central role in the intracellular transport of virus particles but also regulate a wider range of processes critical to the outcome of infection.KEYWORDS virus, cytoskeleton, microtubules, nucleation, microtubule-associated proteins, plus-end-tracking proteins T he dense cytosolic environment poses a major barrier to the free movement of macromolecules, making microtubule (MT)-based transport a critical aspect of virus replication. Indeed, almost as soon as these filaments were identified and characterized, virus particles were observed proximal to "microtubuli," with evidence that MTassociated proteins (MAPs) might mediate this association. Chemicals that disrupt MT networks, still commonly used today, were reported to suppress virus infection. Moreover, either infection or expression of viral proteins was observed to alter the organization of these networks, suggesting that MTs not only facilitated infection but were also likely to be actively manipulated by viruses. In subsequent decades an enormous body of work helped unravel how virus particles exploit MT motors to traffic within infected cells, and we direct readers to a recent review of this subject (1). Here, we discuss recent advances in our understanding of how MTs themselves are regulated and function during infection, limiting this minireview to some illustrative examples in each case. TWO ENDS TO THE STORY: THE BASICS OF MT POLARITY AND ORGANIZATIONMTs form through the polymerization of ␣/␤-tubulin heterodimers into polarized filaments that have minus and plus ends (Fig. 1A and B). MTs nucleate through minus-end seeding at MT-organizing cen...

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A bacteriophage tubulin harnesses dynamic instability to center DNA in infected cells

Cited by 70 publications

References 38 publications

The Phage Nucleus and Tubulin Spindle Are Conserved among Large Pseudomonas Phages

The Phage Nucleus and Tubulin Spindle Are Conserved among Large Pseudomonas Phages

Bacterial Tubulins A and B Exhibit Polarized Growth, Mixed-Polarity Bundling, and Destabilization by GTP Hydrolysis

Microtubule Regulation and Function during Virus Infection

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