Transcriptional Profiling ofCaulobacter crescentusduring Growth on Complex and Minimal Media

Abstract

Microarray analysis was used to examine gene expression in the freshwater oligotrophic bacterium Caulobacter crescentus during growth on three standard laboratory media, including peptone-yeast extract medium (PYE) and minimal salts medium with glucose or xylose as the carbon source. Nearly 400 genes (approximately 10% of the genome) varied significantly in expression between at least two of these media. The differentially expressed genes included many encoding transport systems, most notably diverse TonB-dependent outer membrane channels of unknown substrate specificity. Amino acid degradation pathways constituted the largest class of genes induced in PYE. In contrast, many of the genes upregulated in minimal media encoded enzymes for synthesis of amino acids, including incorporation of ammonia and sulfate into glutamate and cysteine. Glucose availability induced expression of genes encoding enzymes of the Entner-Doudoroff pathway, which was demonstrated here through mutational analysis to be essential in C. crescentus for growth on glucose. Xylose induced expression of genes encoding several hydrolytic exoenzymes as well as an operon that may encode a novel pathway for xylose catabolism. A conserved DNA motif upstream of many xylose-induced genes was identified and shown to confer xylose-specific expression. Xylose is an abundant component of xylan in plant cell walls, and the microarray data suggest that in addition to serving as a carbon source for growth of C. crescentus, this pentose may be interpreted as a signal to produce enzymes associated with plant polymer degradation.Caulobacter species are ubiquitous inhabitants of freshwater, marine, and subsurface environments and are sufficiently adaptable to low nutrient conditions that tapwater and even distilled water are ready sources for their isolation (19,48,49). The physiological properties that enable Caulobacter and other oligotrophic species to survive and reproduce in such environments are not well understood. In this work, we used wholegenome transcriptional profiling and genetic analysis to characterize metabolic pathways and transport systems that are differentially expressed by Caulobacter crescentus during growth in three standard laboratory media: a defined "minimal" inorganic salts medium supplemented with either glucose or xylose as the sole carbon source and a complex peptone-yeast extract medium (PYE), in which amino acids serve as the primary carbon source. These observations provide important baseline data on the genetic coordination of metabolism in this model organism.The dimorphic life cycle of C. crescentus has served as a model for understanding prokaryotic development, cell cycle regulation, and asymmetric cell division (24). At each division, C. crescentus divides into two morphologically and physiologically distinct cells: motile swarmer progeny with a single polar flagellum, and sessile stalked progeny displaying an adhesive stalk. The swarmer progeny, in which DNA replication is initially inhibited, is specialized for dispersal ...

Genetic Analysis of a Novel Pathway for d -Xylose Metabolism in Caulobacter crescentus

“…Six genes yielding the xyl mutant phenotype were found in multiple independent isolates, suggesting that the mutagenesis was approaching saturation and that these represent most, if not all, of the genes required specifically for xylose metabolism. None of the xyl mutant strains had a mutation in a putative transcriptional activator, consistent with previous suggestions (12,18) that C. crescentus xylose metabolism genes are controlled by an as-yet-unidentified repressor. In addition, no genes resembling transporters were identified in this screen.…”

“…The gene in which this insertion is located, designated "xylX" by Meisenzahl et al (18) and later "CC0823" in the C. crescentus genome annotation (19), does not closely resemble any gene of known function. xylX is the first gene in a xylose-inducible operon (CC0823-CC0819) (12), referred to here as the xyl operon. We show that all of the genes in this operon are involved in xylose metabolism and propose a metabolic pathway employing these gene products.…”

“…Using chromosomal DNA as the template and primers derived from the Tn5 sequence, mutants with growth defects were analyzed by cycle sequencing to determine the location of the transposon insertion in comparison with that of the C. crescentus genome sequence (19). Strains unable to use glucose but unaffected in xylose utilization had mutations in genes previously implicated in glucose catabolism (12), including components of the Entner-Doudoroff (E-D) pathway ( Fig. 1) (12, 22).…”

“…Excessive uptake of a nonmetabolized sugar, or the effects of xylose on gene expression, could also result in metabolic alterations that are harmful in the absence of metabolite flux through the xylose catabolic pathway. For example, xylose increases isocitrate lyase expression in C. crescentus (12). During growth on glucose in the absence of productive xylose metabolism, an increase in isocitrate lyase activity could excessively channel isocitrate into the glyoxylate bypass at the expense of critical TCA cycle intermediates, such as ␣-ketoglutarate, that are no longer being generated (directly or indirectly) from D-xylose.…”

Diversity and antagonistic potential of bacteria isolated from marine grass Halodule uninervis

“…From soil, maximum numbers of bacteria were cultivable on ZMA, while more bacterial isolates were counted from roots and leaves on ½ R2A. Complex media contain more nutrients that allow fastidious bacteria to grow, while minimal media contain lower concentration of nutrients hence allow slow growing bacteria to grow (Hottes et al 2004;Majzlik et al 2011). While some slow growing bacteria need prolong period for growth on minimal media (Connon and Giovannoni 2002;Alain and Querellou 2009).…”