Dataset Information

Transposon sequencing uncovers an essential regulatory function of phosphoribulokinase for methylotrophy

ABSTRACT: Methylotrophy is the ability of organisms to grow at the expense of reduced one-carbon compounds such as methanol or methane. Here, we used transposon sequencing combining hyper-saturated transposon mutagenesis with high-throughput sequencing to define the essential methylotrophy genome of Methylobacterium extorquens PA1, a model methylotroph. To distinguish genomic regions required for growth only on methanol from general required genes, we contrasted growth on methanol with growth on succinate, a non-methylotrophic reference substrate. About 500'000 insertions were mapped for each condition, resulting in a median insertion distance of 5 base pairs. We identified 147 genes and 76 genes as specific for growth on methanol and succinate, respectively, and a set of 590 genes as required under both growth conditions. For the integration of metabolic functions, we reconstructed a genome-scale metabolic model and performed in silico essentiality analysis. In total, the approach uncovered 95 previously not described genes as crucial for methylotrophy, including genes involved in respiration, carbon metabolism, transport, and regulation. Strikingly, regardless of the absence of the Calvin cycle in the methylotroph, the screen led to the identification of the gene for phosphoribulokinase as essential during growth on methanol but not on succinate. Genetic experiments in addition to metabolomics and proteomics revealed that phosphoribulokinase serves a key regulatory function. Our data support a model according to which ribulose-1,5 bisphosphate is an essential metabolite that induces a transcriptional regulator driving one-carbon assimilation.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Methylobacterium Extorquens Pa1

SUBMITTER: Lucas Hemmerle

LAB HEAD: Julia A. Vorholt

PROVIDER: PXD006834 | Pride | 2017-07-11

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	20161212_06_AO_Sample_9_3.mgf	Mgf
	20161212_06_AO_Sample_9_3.raw	Raw
	20161212_06_AO_Sample_9_3_3ul.mgf	Mgf
	20161212_06_AO_Sample_9_3_3ul.raw	Raw
	20161212_07_AO_Sample_4_2.mgf	Mgf

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Transposon Sequencing Uncovers an Essential Regulatory Function of Phosphoribulokinase for Methylotrophy.

Ochsner Andrea M AM Christen Matthias M Hemmerle Lucas L Peyraud Rémi R Christen Beat B Vorholt Julia A JA

Current biology : CB 20170817 17

Methylotrophy is the ability of organisms to grow at the expense of reduced one-carbon compounds, such as methanol or methane. Here, we used transposon sequencing combining hyper-saturated transposon mutagenesis with high-throughput sequencing to define the essential methylotrophy genome of Methylobacterium extorquens PA1, a model methylotroph. To distinguish genomic regions required for growth only on methanol from general required genes, we contrasted growth on methanol with growth on succinat ...[more]

PMID: 28823675

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