Penicillium roqueforti PR toxin gene cluster characterization

Abstract

PR toxin is a well-known isoprenoid mycotoxin almost solely produced by Penicillium roqueforti after growth on food or animal feed. This mycotoxin has been described as the most toxic produced by this species. In this study, an in silico analysis allowed identifying for the first time a 22.4-kb biosynthetic gene cluster involved in PR toxin biosynthesis in P. roqueforti. The pathway contains 11 open reading frames encoding for ten putative proteins including the major fungal terpene cyclase, aristolochene synthase, involved in the first farnesyl-diphosphate cyclization step as well as an oxidoreductase, an oxidase, two P450 monooxygenases, a transferase, and two dehydrogenase enzymes. Gene silencing was used to study three genes (ORF5, ORF6, and ORF8 encoding for an acetyltransferase and two P450 monooxygenases, respectively) and resulted in 20 to 40% PR toxin production reductions in all transformants proving the involvement of these genes and the corresponding enzyme activities in PR toxin biosynthesis. According to the considered silenced gene target, eremofortin A and B productions were also affected suggesting their involvement as biosynthetic intermediates in this pathway. A PR toxin biosynthesis pathway is proposed based on the most recent and available data.

The Biosynthetic Gene Cluster for Andrastin A in Penicillium roqueforti

“…This technology has been widely used to demonstrate the functionality of several biosynthetic gene clusters in P. roqueforti (Kosalková et al, 2015; Del-Cid et al, 2016; Hidalgo et al, 2016). For this purpose, ten suitable plasmids were constructed and used to transform P. roqueforti CECT 2905 (see Material and Methods for details).…”

“…Penicillium roqueforti strain CECT 2905 produces several secondary metabolites and currently, the gene clusters responsible for the biosynthesis of three of them, namely roquefortine C, PR-toxin and mycophenolic acid, have been identified (Kosalková et al, 2015; Del-Cid et al, 2016; Hidalgo et al, 2016). Here we describe the gene cluster responsible for the biosynthesis of another secondary metabolite from this fungus, andrastin A, a promising antitumoral compound.…”

“…Thus, this species produces mycotoxins (such as roquefortine C and PR-toxin), and bioactive compounds such as mycophenolic acid and andrastin A (García-Estrada and Martín, 2016). In P. roqueforti , the gene clusters responsible for the biosynthesis of roquefortine C, PR-toxin and mycophenolic acid have been recently identified (Kosalková et al, 2015; Del-Cid et al, 2016; Hidalgo et al, 2016). However, to date the genes responsible for the biosynthesis of andrastin A in this fungus remain as unknown.…”

PR Toxin – Biosynthesis, Genetic Regulation, Toxicological Potential, Prevention and Control Measures: Overview and Challenges

“…Among these, only PR toxin is lethal and can cause a wide range of short-term as well as long-term ill-effects in organisms, ranging from immediate or delayed toxic response to potentially adverse long-term carcinogenic effects in animals such as rats, mice, and cats ( Wei et al, 1975 , 1976 ; Chen et al, 1982 ). It is worth mentioning that in spite of being well known for its acute toxicity, only a few toxicity data and its implicated mechanism are available in the literature ( Hidalgo et al, 2014 , 2017 ; Riclea and Dickschat, 2015 ; García-Estrada and Martín, 2016 ). Previous studies reveal that PR toxin is lethal to rodents after administered orally (p.o.…”

“…The PR toxin producing P. roqueforti was first isolated and partially characterized by Wei et al (1975) . The biosynthetic cluster involved in PR toxin production in P. roqueforti was recently elucidated ( Hidalgo et al, 2014 , 2017 ). The biosynthetic route, leading to the production of PR toxin have been confirmed through various precursors ( 14 C and 13 C) and revealed the isoprenoid route for biosynthesis of PR toxin ( Pedrosa and Griessler, 2010 ).…”