| 1. |
- Abdian, Patricia L., et al.
(author)
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RapA2 Is a Calcium-binding Lectin Composed of Two Highly Conserved Cadherin-like Domains That Specifically Recognize Rhizobium leguminosarum Acidic Exopolysaccharides
- 2013
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In: Journal of Biological Chemistry. - 1083-351X. ; 288:4, s. 2893-2904
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Journal article (peer-reviewed)abstract
- In silico analyses have revealed a conserved protein domain (CHDL) widely present in bacteria that has significant structural similarity to eukaryotic cadherins. A CHDL domain was shown to be present in RapA, a protein that is involved in autoaggregation of Rhizobium cells, biofilm formation, and adhesion to plant roots as shown by us and others. Structural similarity to cadherins suggested calcium-dependent oligomerization of CHDL domains as a mechanistic basis for RapA action. Here we show by circular dichroism spectroscopy, light scattering, isothermal titration calorimetry, and other methods that RapA2 from Rhizobium leguminosarum indeed exhibits a cadherin-like beta-sheet conformation and that its proper folding and stability are dependent on the binding of one calcium ion per protein molecule. By further in silico analysis we also reveal that RapA2 consists of two CHDL domains and expand the range of CHDL-containing proteins in bacteria and archaea. However, light scattering assays at various concentrations of added calcium revealed that RapA2 formed neither homo-oligomers nor hetero-oligomers with RapB (a distinct CHDL protein), indicating that RapA2 does not mediate cellular interactions through a cadherin-like mechanism. Instead, we demonstrate that RapA2 interacts specifically with the acidic exopolysaccharides (EPSs) produced by R. leguminosarum in a calcium-dependent manner, sustaining a role of these proteins in the development of the biofilm matrix made of EPS. Because EPS binding by RapA2 can only be attributed to its two CHDL domains, we propose that RapA2 is a calcium-dependent lectin and that CHDL domains in various bacterial and archaeal proteins confer carbohydrate binding activity to these proteins.
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| 2. |
- Ausmees, Nora
(author)
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Coiled coil cytoskeletons collaborate in polar growth of Streptomyces.
- 2013
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In: Bioarchitecture. - : Informa UK Limited. - 1949-0992 .- 1949-100X. ; 3:4, s. 110-112
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Journal article (peer-reviewed)abstract
- Streptomyces is a multicellular mycelial bacterium, which exhibits pronounced cell polarity and grows by extension of the hyphal tips. Similarly to other polarly growing walled cells, such as filamentous fungi or pollen tubes, Streptomyces hyphae face an intrinsic problem: addition of new cell wall material causes structural weakness of the elongating tip. Cellular strategies employed by walled cells to cope with this problem are not well understood. We have identified a coiled coil protein FilP, with properties similar to those of animal intermediate filament (IF) proteins, which somehow confers rigidity and elasticity to the Streptomyces hyphae. In a recent publication we showed that FilP forms extensive cis-interconnected networks, which likely explain its biological function in determining the mechanical properties of the cells. Surprisingly, the intrinsically non-dynamic cytoskeletal network of FilP exhibits a dynamic behavior in vivo and assembles into growth-dependent polar gradients. We show that apical accumulation of FilP is dependent on its interaction with the main component of the Streptomyces polarisome, DivIVA. Thus, the same polarisome complex that orchestrates cell elongation, also recruits an additional stress-bearing structure to the growing tips with an intrinsically weak cell wall. Similar strategy might be used by all polarly growing walled cells.
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| 3. |
- Ausmees, Nora
(author)
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Intermediate filament-like cytoskeleton of Caulobacter crescentus
- 2006
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In: Journal of Molecular Microbiology and Biotechnology. - : S. Karger AG. - 1464-1801. ; 11:3-5, s. 152-158
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Journal article (peer-reviewed)abstract
- Eukaryotic cytoskeleton consists of three main types of filaments: actin microfilaments, microtubules and intermediate filaments (IFs). Actin and tubulin-like proteins are also found in bacteria where they perform diverse cytoskeletal functions. IFs, however, are considered to be a characteristic constituent of metazoan cells only, where they (among other functions) are involved in determination and maintenance of cell shape and cellular integrity. Surprisingly, a coiled coil-rich protein called crescentin was recently shown to play a key role in determining the complex curved and helical cell shapes of the bacterium Caulobacter crescentus, and to exhibit several characteristic properties of animal IF proteins. First, the arrangement of the coiled coil domains of crescentin closely resembles the tripartite molecular architecture of IF proteins. Second, crescentin also possesses the defining biochemical property of IF proteins to assemble into 10-nm-wide filaments in vitro without cofactors. Furthermore, crescentin forms a higher-order helical structure in vivo, which is localized asymmetrically along the concave side of the cell. In close association with the cell membrane, the crescentin structure promotes the helical growth of the cell and thereby determines a curved or a helical shape, depending on the length of the cell. The unexpected finding of an IF-like element in a bacterium raises several interesting questions concerning, for example, the molecular mechanisms whereby complex and asymmetric cell shapes are generated by different bacteria, or the functional and evolutionary relatedness of crescentin to animal IF proteins.
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| 4. |
- Ausmees, Nora, et al.
(author)
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SmeA, a small membrane protein with multiple functions in Streptomyces sporulation including targeting of a SpoIIIE/FtsK-like protein to cell division septa
- 2007
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In: Molecular Microbiology. - : Wiley. - 0950-382X .- 1365-2958. ; 65:6, s. 1458-1473
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Journal article (peer-reviewed)abstract
- Sporulation in aerial hyphae of Streptomyces coelicolor involves profound changes in regulation of fundamental morphogenetic and cell cycle processes to convert the filamentous and multinucleoid cells to small unigenomic spores. Here, a novel sporulation locus consisting of smeA (encoding a small putative membrane protein) and sffA (encoding a SpoIIIE/FtsK-family protein) is characterized. Deletion of smeA-sffA gave rise to pleiotropic effects on spore maturation, and influenced the segregation of chromosomes and placement of septa during sporulation. Both smeA and sffA were expressed specifically in apical cells of sporogenic aerial hyphae simultaneously with or slightly after Z-ring assembly. The presence of smeA-like genes in streptomycete chromosomes, plasmids and transposons, often paired with a gene for a SpoIIIE/FtsK- or Tra-like protein, indicates that SmeA and SffA functions might be related to DNA transfer. During spore development SffA accumulated specifically at sporulation septa where it colocalized with FtsK. However, sffA did not show redundancy with ftsK, and SffA function appeared distinct from the DNA translocase activity displayed by FtsK during closure of sporulation septa. The septal localization of SffA was dependent on SmeA, suggesting that SmeA may act as an assembly factor for SffA and possibly other proteins required during spore maturation.
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| 5. |
- Ausmees, Nora, et al.
(author)
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Structural and putative regulatory genes involved in cellulose synthesis in Rhizobium leguminosarum bv. trifolii
- 1999
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In: Microbiology. - : Microbiology Society. - 1350-0872 .- 1465-2080. ; 145, s. 1253-1262
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Journal article (peer-reviewed)abstract
- Six genes involved in cellulose synthesis in Rhizobium leguminosarum bv. trifolii were identified using Tn5 mutagenesis. Four of them displayed homology to the previously cloned and sequenced Agrobacterium tumefaciens cellulose genes celA, celB, celC and celE. These genes are organized similarly in R. leguminosarum bv. trifolii. In addition, there were strong indications that two tandemly located genes, celR1 and celR2, probably organized as one operon, are involved in the regulation of cellulose synthesis. The deduced amino acid sequences of these genes displayed a high degree of similarity to the Caulobacter crescentus DivK and PleD proteins that belong to the family of two-component response regulators. This is to our knowledge the first report of genes involved in the regulation of cellulose synthesis. Results from attachment assays and electron microscopic studies indicated that cellulose synthesis in R. leguminosarum bv. trifolii is induced upon close contact with plant roots during the attachment process.
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| 6. |
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| 7. |
- Bagchi, Sonchita, 1978-
(author)
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Coiled coil Cytoskeleton in Bacterial Cell Architecture : Studies of Growth and Development in Streptomyces
- 2011
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Doctoral thesis (other academic/artistic)abstract
- Bacterial cytoskeleton is an exciting and relatively new field of research. Recent findings have proven that microbes are well-organized and neatly structured organisms. In this study we have shown that intermediate filament-like proteins with a characteristic rod domain architecture of coiled coil segments separated by non-coiled coil linkers, are widely spread among bacteria. We identified and characterized an intermediate filament-like protein (named FilP after filamentous protein) in Streptomyces coelicolor. It shares the characteristic biochemical property of eukaryotic intermediate filaments of formation of spontaneous filaments in vitro without requiring any energy or co-factor. We have provided here a preliminary model of its assembly in vitro. FilP also forms in vivo filaments in S. coelicolor hyphae, which are strongest at the sub-apical location of growing vegetative hyphae. We have proposed that FilP cytoskeletal network provides rigidity to the hyphae, especially at the growing tips, by interacting with an essential coiled coil protein DivIVA and possibly other partner elements, yet to be found. S. coelicolor is a well-studied model organism with a complicated life cycle. It germinates from a spore and spreads by forming branched vegetative hyphae. Lack of nutrients in the environment initiates formation of aerial hyphae in the air, perpendicular to the vegetative ones. The aerial hyphae differentiate into spore chains and eventually grey-pigmented dispersed individual spores are released. The signals involved in sporulation including cell division and chromosome segregation are not clear yet. We characterized here a novel locus consisting of two genes: a small putative membrane protein with no defined function, named SmeA and a member of the SpoIIIE/FtsK family, called SffA. The expression of this locus appears to be dependent on whiA and whiG-whiH-whiI pathways. This finding is intriguing as it can provide insight to the relationship between two apparently unrelated pathways, both leading to the same function of septation and maturation during sporulation.
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| 8. |
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| 9. |
- Cabeen, Matthew T., et al.
(author)
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Bacterial cell curvature through mechanical control of cell growth
- 2009
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In: EMBO Journal. - : Wiley. - 0261-4189 .- 1460-2075. ; 28:9, s. 1208-1219
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Journal article (peer-reviewed)abstract
- The cytoskeleton is a key regulator of cell morphogenesis. Crescentin, a bacterial intermediate filament-like protein, is required for the curved shape of Caulobacter crescentus and localizes to the inner cell curvature. Here, we show that crescentin forms a single filamentous structure that collapses into a helix when detached from the cell membrane, suggesting that it is normally maintained in a stretched configuration. Crescentin causes an elongation rate gradient around the circumference of the sidewall, creating a longitudinal cell length differential and hence curvature. Such curvature can be produced by physical force alone when cells are grown in circular microchambers. Production of crescentin in Escherichia coli is sufficient to generate cell curvature. Our data argue for a model in which physical strain borne by the crescentin structure anisotropically alters the kinetics of cell wall insertion to produce curved growth. Our study suggests that bacteria may use the cytoskeleton for mechanical control of growth to alter morphology. The EMBO Journal (2009) 28, 1208-1219. doi: 10.1038/emboj.2009.61; Published online 12 March 2009
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| 10. |
- Cabeen, Matthew T., et al.
(author)
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Mutations in the Lipopolysaccharide Biosynthesis Pathway Interfere with Crescentin-Mediated Cell Curvature in Caulobacter crescentus
- 2010
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In: Journal of Bacteriology. - 0021-9193 .- 1098-5530. ; 192:13, s. 3368-3378
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Journal article (peer-reviewed)abstract
- Bacterial cell morphogenesis requires coordination among multiple cellular systems, including the bacterial cytoskeleton and the cell wall. In the vibrioid bacterium Caulobacter crescentus, the intermediate filament-like protein crescentin forms a cell envelope-associated cytoskeletal structure that controls cell wall growth to generate cell curvature. We undertook a genetic screen to find other cellular components important for cell curvature. Here we report that deletion of a gene (wbqL) involved in the lipopolysaccharide (LPS) biosynthesis pathway abolishes cell curvature. Loss of WbqL function leads to the accumulation of an aberrant O-polysaccharide species and to the release of the S layer in the culture medium. Epistasis and microscopy experiments show that neither S-layer nor O-polysaccharide production is required for curved cell morphology per se but that production of the altered O-polysaccharide species abolishes cell curvature by apparently interfering with the ability of the crescentin structure to associate with the cell envelope. Our data suggest that perturbations in a cellular pathway that is itself fully dispensable for cell curvature can cause a disruption of cell morphogenesis, highlighting the delicate harmony among unrelated cellular systems. Using the wbqL mutant, we also show that the normal assembly and growth properties of the crescentin structure are independent of its association with the cell envelope. However, this envelope association is important for facilitating the local disruption of the stable crescentin structure at the division site during cytokinesis.
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