Genome-Guided Discovery and Heterologous Biosynthesis of Alkylresorcinols by Collaborating Highly Reducing and Type III Polyketide Synthases

The rice sheath rot pathogens <i>Sarocladium attenuatum</i> and <i>Sarocladium oryzae</i> share a conserved biosynthetic gene cluster (<i>sarc</i>), which encodes colocalized highly reducing polyketide synthases (HR-PKSs) and type III polyketide synthases (T3PKSs). Heterologous expression of the <i>sarc</i> cluster in the <i>Aspergillus nidulans</i> strain LO8030 led to the production of six previously unreported alkylresorcinols, sarocladones A-D (<b>1</b>-<b>4</b>) and H-I (<b>5</b>-<b>6</b>), along with three putative artifacts arising from <b>4</b>, sarocladones E-G (<b>4a</b>-<b>4c</b>). Biological screening revealed that <b>1</b> and <b>4c</b> both exhibit mild cytotoxicity against murine NS-1 myeloma cells, with IC₅₀ values of 13 μM and 9 μM, respectively. In addition, <b>1</b>, <b>3</b>, <b>6</b> and <b>7</b> displayed antiphagocytotic activity against THP1 macrophages. Subsequent bioinformatic analysis identified a homologous biosynthetic gene cluster (<i>col</i>) in the genome of the fungal plant pathogen <i>Colletotrichum fructicola</i>. To expand the structural diversity of alkylresorcinols, we employed a gene-mixing strategy, coexpressing the HR-PKS gene <i>colA</i> with the T3PKS gene <i>sarcB</i> in <i>A. nidulans</i>. This resulted in the production of two new sarocladone analogues, collecladones A (<b>7</b>) and B (<b>8</b>), lacking the C-2-C-3 double bond present in the sarocladones. These findings establish HR-PKS-T3PKS collaboration as an underexplored source of fungal chemical diversity.