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3 Initiation and Early Developmental Events
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Myxococcus xanthus is a rod-shaped, gram-negative soil bacterium that, when subjected to nutrient deprivation, undergoes a developmental process culminating in the formation of a multicellular fruiting body filled with spores. For the purposes of this chapter, early development can be defined as events occurring from initiation to the start of aggregation at approximately the first 6 h poststarvation. In the chapter, the following tenets are addressed: how individual M. xanthus cells recognize starvation; how these cells perceive population starvation; and how individual cells integrate this information to ultimately initiate fruiting body formation and cellular differentiation. In summary, the balance of SocE and CsgA proteins in the cell is critical for sustaining the developmental program past initiation and is just one example of the unique aspects of the stringent response in this organism. Based on the current data, the simplest model for nutrient sensing still focuses on the cell’s ability to utilize its translational capacity as an overall measurement of starvation. It should be noted that other members of the Group B signaling mutants remain unmapped and are yet to be extensively characterized. A section exclusively discusses the properties of the bsgA mutants.
Pictorial and photographic representations of the developmental process in M. xanthus DK1622. The diagram shows approximate times for each step in the process: starvation (0 h), aggregation (6 to 8 h), mound formation (12 h), fruiting body formation and sporulation (24 to 48 h). The first row represents development in an MC7 submerged culture system ( Kuner and Kaiser, 1982 ), and the second row represents development on TPM starvation agar plates at a magnification of × 40. This figure is adapted from Tzeng et al., 2006 .
Diagram of the E. coli stringent response. The enzymes involved in the (p)ppGpp metabolism are shown in bold. Ribosome-associated RelA or SpoT catalyzes the synthesis of pppGpp from ATP and GTP upon amino acid or carbon starvation, respectively. Gpp (or Ppx) dephosphorylates pppGpp to make ppGpp. (p)ppGpp accumulates in the cell and interacts with RNAP and DksA [which has been shown to play an important role in (p)ppGpp-dependent transcriptional regulation ( Paul et al., 2004 , 2005; Perederina et al., 2004 )] to positively and negatively control transcription to respond to starvation. (p)ppGpp levels are modulated by SpoT, and when nutrient conditions change, SpoT hydrolyzes ppGpp to GDP (ppG). Ppk is involved in ATP synthesis, and Ndk forms GTP from ATP and GDP (ppG). For reviews, consult Cashel et al., 1996, and Chatterji and Ojha, 2001.
Diagram of the M. xanthus (p)ppGpp response and genes involved in its activation. The enzymes involved in (p)ppGpp metabolism are shown in bold. The asterisk (*) represents uncharged tRNA in the acceptor site of the ribosome that triggers the associated RelA to catalyze the synthesis of (p)ppGpp from ATP and GTP. See Figure 2 legend for more details. In M. xanthus, (p)ppGpp levels are maintained by balancing the hydrolase activity of RelA with its biosynthetic activity. Diodati and Singer have postulated that Shd, a gene product with homology to the hydrolase domain of E. coli SpoT, may play a role in (p)ppGpp degradation. In addition, five proteins (SocE [Crawford and Shimkets, 2000b], Nsd [Brenner et al., 2004], Nla18 [Diodati et al., 2006], Nla4 [Diodati and Singer, personal communication; Ossa et al., unpublished], and CsgA [Crawford and Shimkets, 2000b]) have been shown to either inhibit or stimulate ppGpp accumulation in M. xanthus through as yet unknown mechanisms. For more details, consult text. With elevated (p)ppGpp levels, RNA polymerase (EσA) is predicted to interact with DksA, to modulate changes in RNA polymerase activity to alter gene expression. To date no secondary (p)ppGpp biosynthetic pathway has been identified.
Linear phosphorelay model of A-signaling in M. xanthus. Linear model of A-signal production by a phosphorelay (Kaplan and Plamann, 1996). In this model, AsgA recognizes a rise in (p)ppGpp levels, which leads to the phosphorylation of AsgA and initiates a phosphorelay that ultimately activates the production of the A-signal proteases, which in turn produce the A-signal amino acids, collectively known as A-signal. This model predicts the existence of a histidine phosphotransfer protein, designated AsgX as an intermediary between AsgA~P and AsgB. Phosphorylated AsgB in conjunction with SigA (AsgC) activates expression of the A-signal proteases. Dashed lines represent predicted, yet mechanistically unknown interactions.
The network model of A-signaling in M. xanthus. An alternative model whereby AsgA recognizes a rise in (p)ppGpp levels and sits on top of a hierarchy of genes that are required to activate a variety of signals that collectively make up the quorum-sensing system of M. xanthus. In the simplest model, AsgA is required for all components of the A-signaling system. Alternatively, there may be requirements for other starvation signals [in addition to (p)ppGpp] or additional hierarchical regulators (like AsgD) that either directly or indirectly activate the system. These inputs are represented by dashed lines.
Model for dual starvation in M. xanthus. Schematic of major players, identified to date, that modulate entrance into the developmental process by monitoring and responding to nutrient levels. Proteins are identified in boldface type, and genes are in italics; direct interactions are represented by solid lines, presumed indirect interactions are shown as dashed lines, and proposed interactions are indicated by a dotted line and a question mark (?). Arrowheads indicate a positive interaction, and a blunt head indicates a negative interaction. Note : for simplicity, some important components of this process are not included in this model; thus, it is not all-inclusive. For more details, see text.
Conditions known to initiate development and induce a stringent response a
List of stringent-response-related homologues in M. xanthusa
List of genes affecting (p)ppGpp accumulation in M. xanthus
Partial list of genes implicated in nutrient sensing and developmental timing
Names, MXAN and Mx numbers, N-terminal regulatory domains, gene knockout constructions, mutant phenotypes, and related references for all EBPs in M. xanthusa