Analysis of interactions between nutrient germinant receptors and SpoVA proteins of<i>Bacillus subtilis</i>spores

Vepachedu, Venkata Ramana; Setlow, Peter

doi:10.1111/j.1574-6968.2007.00807.x

Cited by 39 publications

(39 citation statements)

References 22 publications

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“…However, this degradation step happens after Ca-DPA release during germination and thus is not likely to be the ratelimiting step. Finally, the various SpoVA proteins thought to comprise the channel for Ca-DPA release in spore germination (1,25) are present in spores at ϳ10 3 ϫ higher levels than the GRs (26), and fluorescence microscopy of spores carrying SpoVA-GFP showed relatively constant fluorescence intensity in spores (data not shown), consistent with a relatively uniform expression level of SpoVA proteins. Unfortunately, the overall molecular mechanism of the nutrient-induced germination process is still not completely clear.…”

Section: Figmentioning

confidence: 57%

Expression Level of Bacillus subtilis Germinant Receptors Determines the Average Rate but Not the Heterogeneity of Spore Germination

et al. 2013

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Germination of Bacillus subtilis spores can be triggered by the binding of specific nutrients, called germinants, to germinant receptors (GRs) in the spore's inner membrane. This interaction eventually initiates, with variable time delays, the release of dipicolinic acid and cations from the spore core-a key step in spore germination. The kinetics of this process are highly heterogeneous for individual spores. In this work, we sought to investigate how the germination heterogeneity was controlled. In particular, we tested whether the rates of germination were determined by GR levels, which vary from spore to spore due to stochastic gene expression. Both the expression levels of GRs and the germination rate were measured in single spores, and the experimental results were compared to theoretical predictions. Our results indicated that the variation in the expression levels of GRs was not the primary factor that controls spore germination heterogeneity. Two alternative hypotheses are discussed in light of this experimental discovery.

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Section: Figmentioning

confidence: 57%

Expression Level of Bacillus subtilis Germinant Receptors Determines the Average Rate but Not the Heterogeneity of Spore Germination

et al. 2013

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“…Evidence that the receptor comprises a complex of at least three different proteins, for example, is provided by the conserved tricistronic receptor operon structure observed in all sporeformer genomes, suggesting that genes within the operon have coevolved, and, where tested, by the absolute requirement for all three receptor components for functionality (20,23). Indirect genetic evidence for physical interaction between the respective A, B, and C proteins that comprise the receptor (13) has been complemented recently by yeast two-hybrid experiments (34), which, in addition to providing evidence for the interaction between receptor subunits, also suggest that some receptor proteins interact with SpoVA proteins thought to be involved in the release of calcium dipicolinate during germination. Similarly, while some germinant receptors can function independently to trigger the spore germination response, for example, the GerA-mediated L-alanine response in Bacillus subtilis, most germinant receptors appear to work in concert to initiate germination, either in response to single germinants (e.g., the GerQ and GerI response to inosine in Bacillus cereus [4]) or to mixtures of germinants (e.g., the GerB-and GerK-mediated response to AGFK in B. subtilis [9,15]).…”

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confidence: 96%

Functional Consequences of Amino Acid Substitutions to GerVB, a Component of the Bacillus megaterium Spore Germinant Receptor

2008

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The extreme metabolic dormancy and resistance properties of spores formed by members of the Bacillus and Clostridium genera are lost upon exposure to a variety of small-molecule germinants. Germinants are known to interact in an as yet undefined manner with cognate receptor complexes that reside in the inner membrane that surrounds the spore protoplast. The receptor itself is a complex of at least three proteins, and in this study we identify amino acid residues, predicted to lie in loop regions of GerVB on the exterior aspect of the membrane, that influence the Bacillus megaterium spore germination response. Three consecutive residues adjacent to putative transmembrane domain 10 (TM10) were demonstrated to mediate to various degrees the proline germinative response while also influencing germination in response to leucine, glucose, and inorganic salts, suggesting that this region may be part of a ligand binding pocket. Alternatively, substitutions in this region may affect the conformation of associated functionally important TM regions. Leucine-and KBrmediated germination was also influenced by substitutions in other outer loop regions. These observations, when considered with accompanying kinetic analyses that demonstrate cooperativity between germinants, suggest that binding sites for the respective germinants are in close spatial proximity in the receptor but do not overlap. Additionally, proline recognition was conferred to a chimeric receptor when TM regions associated with the putative binding loop were present, indicating that residues in TM9 and/or TM10 of GerVB are also of functional importance in the proline-induced germinative response.Endospores formed in response to nutrient starvation by members of the genera Bacillus and Clostridium display remarkable properties of resistance and dormancy that permit their survival in the environment for extended periods. Despite this extreme dormancy, spores retain the ability to initiate vegetative metabolism rapidly upon exposure to appropriate nutrients via the process of germination (20, 30). Significant advances have been made in recent years through the application of mainly genetic techniques to characterize the molecular apparatus involved in spore germination, identifying structural genes that encode germinant receptors (4,8,14,23,37), ion channels (31, 35), and hydrolytic enzymes involved in the degradation of the cortical peptidoglycan that surrounds the spore protoplast (6,11,16,21). Cytological observations on spore coat degradation during germination (29) have also recently been extended by the application of atomic force microscopy to provide insights to the structural basis for the disassembly of the outer layers that provide the primary barrier to environmental insult (26).Despite these advances, however, little is known of the molecular mechanism of the primary event of spore germination-the interaction of the germinant, typically an amino acid, sugar, or riboside, with its cognate receptor-and how this interaction triggers the subsequent casc...

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“…Structural genes encoding the respective A, B, and C receptor subunits are arranged typically in tricistronic operons, the cotranscription of which suggests that the receptor is a complex of all three proteins. This view has been reinforced by molecular genetic (12,18) and biochemical (30) approaches that suggest physical interaction between at least some of the receptor subunits. Genetic evidence that proteins from different receptors can physically interact has also been presented (3,5).…”

mentioning

confidence: 99%

“…Precise mechanisms that permit rapid movement of these molecules across the membrane have not been elucidated, although SpoVA proteins that are involved in uptake of DPA in the developing forespore during sporulation seem also to be involved in its release during germination, perhaps by forming a pore or channel in the spore inner membrane (27,31,32). Biochemical evidence for physical interaction between B. subtilis GerA receptor subunits and two different SpoVA proteins suggests a route to signal transduction between the receptor and putative DPA channel (30).…”

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confidence: 99%

Amino Acid Substitutions in Transmembrane Domains 9 and 10 of GerVB That Affect the Germination Properties of Bacillus megaterium Spores

Christie

Lowe

2008

J Bacteriol

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The molecular basis for differences in germinant recognition of Bacillus megaterium QM B1551 spores containing the GerVB and/or GerUB receptor proteins has been examined by site-directed mutagenesis and the construction of cross-homologue chimeras. Focusing on nonconserved residues predicted to reside in transmembrane domains 9 and 10, we demonstrate that GerVB residues Ser319 and Leu345 are of particular importance in defining the specificity and apparent affinity of the receptor for germinants. Kinetic analyses of mutants with different amino acid substitutions at these positions indicate that Ser319 and Leu345 are not involved directly in the binding of germinants, but probably reside in regions of the receptor where structural perturbations can affect the conformation of, or access to, germinant binding sites. Position 345 is also shown to be of importance in GerUB, where the F345A mutation severely impairs receptor function. Functionality is restored in the GerUB Ala345 background by substituting putative outer-loop residues adjacent to TM10 for the corresponding residues in GerVB, indicating that a degree of structural coordination between these regions is important to receptor function.

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Analysis of interactions between nutrient germinant receptors and SpoVA proteins ofBacillus subtilisspores

Cited by 39 publications

References 22 publications

Expression Level of Bacillus subtilis Germinant Receptors Determines the Average Rate but Not the Heterogeneity of Spore Germination

Expression Level of Bacillus subtilis Germinant Receptors Determines the Average Rate but Not the Heterogeneity of Spore Germination

Functional Consequences of Amino Acid Substitutions to GerVB, a Component of the Bacillus megaterium Spore Germinant Receptor

Amino Acid Substitutions in Transmembrane Domains 9 and 10 of GerVB That Affect the Germination Properties of Bacillus megaterium Spores

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