Position Effect Variegation of Penicillium rubens Genome and Application in Heterologous Production of the Plant-Derived Anthraquinone Physcion.

The filamentous fungus Penicillium rubens is well known as an industrial host for producing penicillin and also used as a promising chassis for secondary metabolite biosynthesis. Chromosomal position effect significantly influences the functional implementation of exogenous genetic modules, but it remains underexplored in filamentous fungi such as P. rubens. In this study, a PiggyBac (PB) transposon-mediated mutagenesis library was constructed in P. rubens. High-throughput sequencing confirmed a broadly distributed with moderate local hopping of PB insertions across this fungal genome. To characterize the genome position effect, two genomic loci libraries including low-expression (P4) and high-expression (P7) positions were obtained respectively by flow cytometry. Heterologous biosynthesis of the plant-derived anthraquinone physcion in P. rubens, via targeted integration of its biosynthetic gene cluster (BGC) into the P7 genomic loci, substantially boosted the titre to a level significantly higher than that achieved via integration at the P4 loci, demonstrating a direct correlation between the integration locus and the secondary metabolite output. To our knowledge, this is the first systematic elucidation of chromosomal position effect on heterologous biosynthesis of secondary metabolites in P. rubens and it also provides an insight for rational selection of integration sites in filamentous fungi.