title: Exploring and dissecting genome-wide gene expression responses of Penicillium chrysogenum to phenylacetic acid consumption and penicillinG production.
abstract: BACKGROUND
  Since the discovery of the antibacterial activity of penicillin by Fleming 80 years ago, improvements of penicillin titer were essentially achieved by classical strain improvement through mutagenesis and screening. The recent sequencing of Penicillium chrysogenum strain Wisconsin1255-54 and the availability of genomics tools such as DNA-microarray offer new perspective.
  
  
  RESULTS
  In studies on beta-lactam production by P. chrysogenum, addition and omission of a side-chain precursor is commonly used to generate producing and non-producing scenarios. To dissect effects of penicillinG production and of its side-chain precursor phenylacetic acid (PAA), a derivative of a penicillinG high-producing strain without a functional penicillin-biosynthesis gene cluster was constructed. In glucose-limited chemostat cultures of the high-producing and cluster-free strains, PAA addition caused a small reduction of the biomass yield, consistent with PAA acting as a weak-organic-acid uncoupler. Microarray-based analysis on chemostat cultures of the high-producing and cluster-free strains, grown in the presence and absence of PAA, showed that: (i) Absence of a penicillin gene cluster resulted in transcriptional upregulation of a gene cluster putatively involved in production of the secondary metabolite aristolochene and its derivatives, (ii) The homogentisate pathway for PAA catabolism is strongly transcriptionally upregulated in PAA-supplemented cultures (iii) Several genes involved in nitrogen and sulfur metabolism were transcriptionally upregulated under penicillinG producing conditions only, suggesting a drain of amino-acid precursor pools. Furthermore, the number of candidate genes for penicillin transporters was strongly reduced, thus enabling a focusing of functional analysis studies.
  
  
  CONCLUSIONS
  This study demonstrates the usefulness of combinatorial transcriptome analysis in chemostat cultures to dissect effects of biological and process parameters on gene expression regulation. This study provides for the first time clear-cut target genes for metabolic engineering, beyond the three genes of the beta-lactam pathway.
authors.affiliation: Department of Biotechnology, Delft University of Technology, Delft, The Netherlands. DHarris@cntnl.jnj.com
authors.forename: Diana M
authors.initials: DM
authors.lastname: Harris
authors.forename: Zita A
authors.initials: ZA
authors.lastname: van der Krogt
authors.forename: Paul
authors.initials: P
authors.lastname: Klaassen
authors.forename: Leonie M
authors.initials: LM
authors.lastname: Raamsdonk
authors.forename: Susanne
authors.initials: S
authors.lastname: Hage
authors.forename: Marco A
authors.initials: MA
authors.lastname: van den Berg
authors.forename: Roel A L
authors.initials: RA
authors.lastname: Bovenberg
authors.forename: Jack T
authors.initials: JT
authors.lastname: Pronk
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Daran
mesh_terms: D003470:Culture Media; D017353:Gene Deletion; D018628:Gene Dosage; D020869:Gene Expression Profiling; D015966:Gene Expression Regulation, Fungal; D016681:Genome, Fungal; D005810:Multigene Family; D020411:Oligonucleotide Array Sequence Analysis; D010400:Penicillin G / Q000378:metabolism*; D010408:Penicillium chrysogenum / Q000235:genetics* / Q000378:metabolism; D010648:Phenylacetates / Q000378:metabolism*; D012331:RNA, Fungal / Q000378:metabolism
chemical_list: D003470:Culture Media; D010648:Phenylacetates; D012331:RNA, Fungal; C025136:phenylacetic acid; D010400:Penicillin G