Gillot Guillaume, Jany Jean-Luc, Dominguez-Santos Rebeca, Poirier Elisabeth, Debaets Stella, Hidalgo Pedro I, Ullán Ricardo V, Coton Emmanuel, Coton Monika
Université de Brest, EA 3882 Laboratoire Universitaire de Biodiversité et d'Ecologie Microbienne, IBSAM ESIAB, Technopôle Brest-Iroise, 29280 Plouzané, France.
Área de Microbiología, Facultad de Ciencias Biológicas y Ambientales, Universidad de León, 24071 León, Spain; Instituto de Biotecnología de León (INBIOTEC), Avenida Real n°1, Parque Científico de León, 24006 León, Spain.
Food Microbiol. 2017 Apr;62:239-250. doi: 10.1016/j.fm.2016.10.013. Epub 2016 Oct 4.
Mycophenolic acid (MPA) is a secondary metabolite produced by various Penicillium species including Penicillium roqueforti. The MPA biosynthetic pathway was recently described in Penicillium brevicompactum. In this study, an in silico analysis of the P. roqueforti FM164 genome sequence localized a 23.5-kb putative MPA gene cluster. The cluster contains seven genes putatively coding seven proteins (MpaA, MpaB, MpaC, MpaDE, MpaF, MpaG, MpaH) and is highly similar (i.e. gene synteny, sequence homology) to the P. brevicompactum cluster. To confirm the involvement of this gene cluster in MPA biosynthesis, gene silencing using RNA interference targeting mpaC, encoding a putative polyketide synthase, was performed in a high MPA-producing P. roqueforti strain (F43-1). In the obtained transformants, decreased MPA production (measured by LC-Q-TOF/MS) was correlated to reduced mpaC gene expression by Q-RT-PCR. In parallel, mycotoxin quantification on multiple P. roqueforti strains suggested strain-dependent MPA-production. Thus, the entire MPA cluster was sequenced for P. roqueforti strains with contrasted MPA production and a 174bp deletion in mpaC was observed in low MPA-producers. PCRs directed towards the deleted region among 55 strains showed an excellent correlation with MPA quantification. Our results indicated the clear involvement of mpaC gene as well as surrounding cluster in P. roqueforti MPA biosynthesis.
霉酚酸（MPA）是由包括罗克福青霉在内的多种青霉菌产生的次生代谢产物。最近在短密青霉中描述了MPA生物合成途径。在本研究中，对罗克福青霉FM164基因组序列进行的电子分析定位了一个23.5 kb的假定MPA基因簇。该基因簇包含七个基因，推测编码七种蛋白质（MpaA、MpaB、MpaC、MpaDE、MpaF、MpaG、MpaH），并且与短密青霉基因簇高度相似（即基因共线性、序列同源性）。为了证实该基因簇参与MPA生物合成，在高产MPA的罗克福青霉菌株（F43-1）中使用RNA干扰靶向编码假定聚酮合酶的mpaC进行基因沉默。在获得的转化体中，MPA产量降低（通过LC-Q-TOF/MS测量）与Q-RT-PCR检测到的mpaC基因表达降低相关。同时，对多个罗克福青霉菌株的霉菌毒素定量分析表明MPA产量具有菌株依赖性。因此，对MPA产量不同的罗克福青霉菌株的整个MPA基因簇进行了测序，在低MPA生产者中观察到mpaC中有174bp的缺失。针对55个菌株中缺失区域进行的PCR与MPA定量分析显示出极好的相关性。我们的结果表明mpaC基因以及周围的基因簇明显参与了罗克福青霉MPA的生物合成。
