{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,1]],"date-time":"2026-06-01T18:08:32Z","timestamp":1780337312305,"version":"3.54.1"},"reference-count":31,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2026,6,1]],"date-time":"2026-06-01T00:00:00Z","timestamp":1780272000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"CAMS Innovation Fund for Medical Sciences","award":["2021-I2M-1-029"],"award-info":[{"award-number":["2021-I2M-1-029"]}]},{"name":"CAMS Innovation Fund for Medical Sciences","award":["2023-I2M-2-006"],"award-info":[{"award-number":["2023-I2M-2-006"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["JoF"],"abstract":"<jats:p>Aspergillus nidulans, a model filamentous fungus endowed with well-established genetic tools and a repertoire of cryptic secondary metabolite biosynthetic gene clusters (BGCs), is extensively exploited as a microbial chassis for heterologous biosynthesis. Mining of its secondary metabolites facilitates the discovery of novel bioactive compounds and the development and application of chassis cells. In the course of heterologous expression of exogenous genes in A. nidulans, we unexpectedly observed the activation of cryptic host BGCs, which resulted in substantial alterations to its secondary metabolic profile. Four previously undescribed compounds (1\u20134), together with six known analogs (5\u201310), were isolated from three recombinant A. nidulans strains. Notably, compounds 1\u20133 are the first naturally occurring examples of diketopiperazine\u2013isoindolinone hybrid alkaloids, while compound 4 is a previously unreported benzofuran carboxylic acid derivative. Their structures and absolute configurations were assigned by interpretation of a combination of spectroscopic data and electronic circular dichroism calculations. Compounds 4 and 5 exhibited potent DPPH radical scavenging activity (IC50, 6.01 and 7.00 \u03bcg\u00b7mL\u22121, respectively). This study uncovers a \u201cmetabolic perturbation\u201d effect on the host metabolic network during heterologous expression and offers a new strategy for activating silent gene clusters and discovering novel natural products through genetic manipulation.<\/jats:p>","DOI":"10.3390\/jof12060401","type":"journal-article","created":{"date-parts":[[2026,6,1]],"date-time":"2026-06-01T17:11:01Z","timestamp":1780333861000},"page":"401","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Heterologous Expression Unexpectedly Activates the Host Cryptic Genes in Aspergillus nidulans and Enables the Discovery of Novel Natural Products"],"prefix":"10.3390","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0009-0009-3753-3357","authenticated-orcid":false,"given":"Cong","family":"Liu","sequence":"first","affiliation":[{"name":"State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yinan","family":"Hao","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Siyuan","family":"Qi","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6080-7960","authenticated-orcid":false,"given":"Jian","family":"Bai","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,1]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"433","DOI":"10.1007\/s10295-013-1386-z","article-title":"Recent advances in genome mining of secondary metabolite biosynthetic gene clusters and the development of heterologous expression systems in Aspergillus nidulans","volume":"41","author":"Yaegashi","year":"2014","journal-title":"J. Ind. Microbiol. Biotechnol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"6","DOI":"10.1186\/s40694-018-0049-2","article-title":"CoIN: Co-inducible nitrate expression system for secondary metabolites in Aspergillus nidulans","volume":"5","author":"Wiemann","year":"2018","journal-title":"Fungal Biol. Biotechnol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"5908","DOI":"10.1128\/jb.172.10.5908-5914.1990","article-title":"Molecular characterization of the acyl-coenzyme A:isopenicillin N acyltransferase gene (penDE) from Penicillium chrysogenum and Aspergillus nidulans and activity of recombinant enzyme in Escherichia coli","volume":"172","author":"Tobin","year":"1990","journal-title":"J. Bacteriol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"8894215","DOI":"10.1155\/2020\/8894215","article-title":"Aspergillus nidulans: A potential resource of the production of the native and heterologous enzymes for industrial applications","volume":"2020","author":"Kumar","year":"2020","journal-title":"Int. J. Microbiol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"103477","DOI":"10.1016\/j.fgb.2020.103477","article-title":"In the fungus where it happens: History and future propelling Aspergillus nidulans as the archetype of natural products research","volume":"144","author":"Caesar","year":"2020","journal-title":"Fungal Genet. Biol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"8143","DOI":"10.1128\/AEM.00683-10","article-title":"Activation of a silent fungal polyketide biosynthesis pathway through regulatory cross talk with a cryptic nonribosomal peptide synthetase gene cluster","volume":"76","author":"Bergmann","year":"2010","journal-title":"Appl. Environ. Microbiol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1444","DOI":"10.1038\/s42003-025-08840-z","article-title":"Systematic over-expression of secondary metabolism transcription factors to reveal the pharmaceutical potential of Aspergillus nidulans","volume":"8","author":"Guo","year":"2025","journal-title":"Commun. Biol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"11022","DOI":"10.1039\/D3SC02226A","article-title":"A heterologous expression platform in Aspergillus nidulans for the elucidation of cryptic secondary metabolism biosynthetic gene clusters: Discovery of the Aspergillus fumigatus sartorypyrone biosynthetic pathway","volume":"14","author":"Lin","year":"2023","journal-title":"Chem. Sci."},{"key":"ref_9","first-page":"1001","article-title":"Full biosynthetic pathway of pyrrolobenzoxazines","volume":"36","author":"Wang","year":"2025","journal-title":"Chin. Chem. Lett."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"4782","DOI":"10.1002\/anie.201701547","article-title":"A Cascade of Redox Reactions Generates Complexity in the Biosynthesis of the Protein Phosphatase-2 Inhibitor Rubratoxin A","volume":"56","author":"Bai","year":"2017","journal-title":"Angew. Chem. Int. Ed."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"572","DOI":"10.1016\/j.apsb.2020.07.020","article-title":"Genome mining combined metabolic shunting and OSMAC strategy of an endophytic fungus leads to the production of diverse natural products","volume":"11","author":"Wei","year":"2021","journal-title":"Acta Pharm. Sin. B"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"687","DOI":"10.1016\/j.apsb.2017.12.012","article-title":"Epigenetic modification in histone deacetylase deletion strain of Calcarisporium arbuscula leads to diverse diterpenoids","volume":"8","author":"Bai","year":"2018","journal-title":"Acta Pharm. Sin. B"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"7592","DOI":"10.1002\/anie.201502452","article-title":"Epigenetic genome mining of an endophytic fungus leads to the pleiotropic biosynthesis of natural products","volume":"54","author":"Mao","year":"2015","journal-title":"Angew. Chem. Int. Ed."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1186","DOI":"10.1021\/acssynbio.1c00458","article-title":"Cre\/lox-Mediated Chromosomal Integration of Biosynthetic Gene Clusters for Heterologous Expression in Aspergillus nidulans","volume":"11","author":"Roux","year":"2022","journal-title":"ACS Synth. Biol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"8","DOI":"10.1186\/1758-2946-3-8","article-title":"Confab-systematic generation of diverse low-energy conformers","volume":"3","author":"Vandermeersch","year":"2011","journal-title":"J. Cheminform"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"27","DOI":"10.1021\/jf00049a007","article-title":"Antioxidant activity of various tea extracts in relation to their antimutagenicity","volume":"43","author":"Yen","year":"1995","journal-title":"J. Agric. Food Chem."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"997","DOI":"10.1039\/c2md20055d","article-title":"Identification and molecular genetic analysis of the cichorine gene cluster in Aspergillus nidulans","volume":"3","author":"Sanchez","year":"2012","journal-title":"MedChemComm"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"4249","DOI":"10.1021\/jacs.6b01528","article-title":"Phenalenone polyketide cyclization catalyzed by fungal polyketide synthase and flavin-dependent monooxygenase","volume":"138","author":"Gao","year":"2016","journal-title":"J. Am. Chem. Soc."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"648","DOI":"10.7164\/antibiotics.43.648","article-title":"Asperfuran, a novel antifungal metabolite from Aspergillus oryzae","volume":"43","author":"Pfefferle","year":"1990","journal-title":"J. Antibiot."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Orfali, R., and Perveen, S. (2019). Secondary metabolites from the Aspergillus sp. in the rhizosphere soil of Phoenix dactylifera (Palm tree). BMC Chem., 13.","DOI":"10.1186\/s13065-019-0624-5"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"7411","DOI":"10.1039\/C7OB01837A","article-title":"Volatiles from the fungal microbiome of the marine sponge Callyspongia cf. flammea","volume":"15","author":"Barra","year":"2017","journal-title":"Org. Biomol. Chem."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"677","DOI":"10.1007\/s10600-009-9433-8","article-title":"iso-\u03b1-Cyclopiazonic acid, a new natural product isolated from the marine-derived fungus Aspergillus flavus C-F-3","volume":"45","author":"Lin","year":"2009","journal-title":"Chem. Nat. Compd."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"7119","DOI":"10.1016\/j.tet.2010.06.092","article-title":"The Knight route to cyclopiazonic acid: Enantioselective synthesis of a key intermediate","volume":"66","author":"Beyer","year":"2010","journal-title":"Tetrahedron"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"550","DOI":"10.1248\/cpb.54.550","article-title":"Nine constituents including six xanthone-related compounds isolated from two ascomycetes, Gelasinospora santi-florii and Emericella quadrilineata, found in a screening study focused on immunomodulatory activity","volume":"54","author":"Fujimoto","year":"2006","journal-title":"Chem. Pharm. Bull."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1225","DOI":"10.1007\/s00343-020-0052-3","article-title":"A new acyclic peroxide from Aspergillus nidulans SD-531, a fungus obtained from deep-sea sediment of cold spring in the South China Sea","volume":"38","author":"Li","year":"2020","journal-title":"J. Oceanol. Limnol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"2305","DOI":"10.1039\/b504602e","article-title":"First total synthesis of cichorine and zinnimidine","volume":"3","author":"Moreau","year":"2005","journal-title":"Org. Biomol. Chem."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"eadk7416","DOI":"10.1126\/sciadv.adk7416","article-title":"Terpenoid balance in Aspergillus nidulans unveiled by heterologous squalene synthase expression","volume":"10","author":"Park","year":"2024","journal-title":"Sci. Adv."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"e00189-26","DOI":"10.1128\/mbio.00189-26","article-title":"ThLaeA functions as a conditional repressor to maintain metabolic homeostasis in Trichoderma hypoxylon","volume":"524","author":"Li","year":"2026","journal-title":"mBio"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"2311","DOI":"10.1016\/j.funbio.2024.11.006","article-title":"Could the transcription factor AtnN coordinating the aspercryptin secondary metabolite gene cluster in Aspergillus nidulans be a global regulator?","volume":"128","author":"Kocsis","year":"2024","journal-title":"Fungal Biol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1376","DOI":"10.1111\/j.1365-2958.2010.07051.x","article-title":"Heterochromatic marks are associated with the repression of secondary metabolism clusters in Aspergillus nidulans","volume":"76","author":"Bok","year":"2010","journal-title":"Mol. Microbiol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"e13092","DOI":"10.1111\/acel.13092","article-title":"Hdac3, Setdb1, and Kap1 mark H3K9me3\/H3K14ac bivalent regions in young and aged liver","volume":"19","author":"Price","year":"2020","journal-title":"Aging Cell"}],"container-title":["Journal of Fungi"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2309-608X\/12\/6\/401\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,1]],"date-time":"2026-06-01T17:32:38Z","timestamp":1780335158000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2309-608X\/12\/6\/401"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,6,1]]},"references-count":31,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2026,6]]}},"alternative-id":["jof12060401"],"URL":"https:\/\/doi.org\/10.3390\/jof12060401","relation":{},"ISSN":["2309-608X"],"issn-type":[{"value":"2309-608X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,6,1]]}}}