An early A-signal-dependent gene in Myxococcus xanthus has a sigma 54-like promoter

Abstract

A-signaling plays an essential role in the early stages of Myxococcus xanthus fruiting body development. Expression of the 4521 gene, which is regulated at the level of RNA accumulation, depends on starvation and on A-signaling. To identify the cis-acting regulatory elements which allow gene 4521 to respond to the nutritional and A-factor signals, the 4521 transcription start site was mapped. The region just upstream of the start site showed sequence similarity to the 54 family of promoters and to the developmentally regulated mbhA promoter of M. xanthus. A mutational analysis of this region established that the bases which were conserved between the 54 consensus, mbhA, and 4521 promoters were also important for 4521 promoter activity. Changes which altered the spacing between two conserved regions centered around positions ؊14 and ؊24 abolished promoter activity. In contrast, mutations in a putative ؊10 region for a 70 -like promoter had little effect on expression of 4521. Despite their similar promoter regions, the regulation of the 4521 and mbhA genes was shown to differ with respect to timing of expression and requirement for a solid surface and extracellular signals. This suggests a model in which different activator proteins may be responsible for regulating expression of these two genes.In response to nutrient limitation in its environment, Myxococcus xanthus enters a developmental cycle in which about 100,000 cells aggregate to build a multicellular structure, the fruiting body. Once inside the fruiting body, the individual rod-shaped cells are able to undergo morphological and physiological differentiation to form ovoid, environmentally resistant, and metabolically dormant spores (39). Progress through this developmental program is highly coordinated and involves intercellular communication. Mutational analysis has shown that at least five extracellular signals are required for progress through different stages of the developmental program (6, 9, 26). One of the extracellular signals required early in development is the A-signal, which consists of a mixture of amino acids or small peptides. These signaling molecules have to be present in a narrow concentration range at 1 to 2 h after the onset of starvation and serve as a cell density signal (24). In the absence of extracellular A-signal, as in the asg class of mutants, development arrests prior to aggregation of the cells into mounds (22).To begin to understand how the information conveyed by the A-signal is transduced inside the cell and exerts its normal function in facilitating progress through development, a gene whose expression is dependent on the A-signaling process has been studied. This gene, subsequently called 4521, was identified by a transcriptional fusion of a promoterless lacZ gene to its promoter. The fusion was generated by transposition of Tn5lac into the transcribed region (20). In an otherwise wildtype cell, the ⍀4521 Tn5lac fusion begins to be expressed early in the developmental process, around 1.5 h after the onset of starvation...

Mutations of the Act Promoter in Myxococcus xanthus

“…This defect exemplifies a requirement for proper spacing between the Ϫ12 and Ϫ24 elements, as has been observed elsewhere (3,29,(43)(44)(45). Strict spacing between promoter elements helps to distinguish sigma-54 promoters from sigma-70 promoters (2,18).…”

“…This resembles the extensively studied M. xanthus sigma-54 pilA promoter (71). It also resembles the M. xanthus ⍀4521 promoter, for which sequence mutations have indicated sigma-54 specificity (29). The sigma-54 gene (rpoN), which is in single copy, is essential for M. xanthus development.…”

Nutrient regime regulates complex transcriptional start site usage within a Pseudoalteromonas chitinase gene cluster

“…Both s 70 - (Table 4a) and s 54 - (Table 4b) dependent promoter sequences were identified, each with homology to their respective consensus sequences (Hawley and McClure, 1983;Barrios et al, 1999) and those reported earlier in E. coli (Dombroski et al, 1996), Myxococcus xanthus (Keseler and Kaiser, 1995) and Pseudomonas aeruginosa (Medina et al, 2003). In addition, a putative in-frame translation initiation codon and corresponding potential ribosome-binding site were identified for each transcript arising from a potential TSP (Table 4).…”

Social complementation and growth advantages promote socially defective bacterial isolates

“…We selected for spontaneous rifampicin-resistant mutants and kanamycin-resistant transformants carrying the plasmid pREG1727 [44]. Only marked clones that exhibited no significant marker effects on focal traits relative to their unmarked parent were used for further experiments (data available upon request).…”