Connell John, Bates Helen J, Geoghegan Ivey, Wilson Fiona, Harrison Richard J, Price R Jordan
NIAB, Cambridge, CB3 0LE, UK.
Marlow Ingredients, Billingham, TS23 4HA, UK.
Fungal Biol Biotechnol. 2025 Mar 24;12(1):3. doi: 10.1186/s40694-025-00195-8.
Quorn mycoprotein, a protein-rich meat alternative, is produced through large-scale fermentation of the fungus Fusarium venenatum. However, a major challenge during F. venenatum fermentation is the consistent appearance of mutants called colonial variants (C-variants). These C-variants have a highly branched morphology, which ultimately lead to a less desirable final product and early termination of the fermentation process. This study aimed to identify the genetic mutations responsible for C-variant morphology.
We first isolated both C-variant and wild-type strains from commercial fermentation samples and characterised radial growth rates on solid media. Whole genome sequencing facilitated the identification of mutations in a gene called jg4843 in 11 out of 12 C-variant isolates, which were not observed in the wild-type isolates. The jg4843 gene was identified as the ortholog of LRG1, a Rho-GTPase activating protein that regulates the Rho1 signalling pathway affecting fungal growth. Notably, the mutations in jg4843 were primarily located in the RhoGAP domain responsible for LRG1 activity. To confirm the role of these mutations, we used CRISPR/Cas9-mediated homology-directed recombination to introduce the C-variant mutations into the wild-type isolate, which successfully recapitulated the characteristic C-variant morphology.
This study identified mutations in the LRG1 ortholog jg4843 as the genetic cause of C-variant morphology in commercial fermentation F. venenatum isolates. Understanding this genetic basis paves the way for developing strategies to prevent C-variants arising, potentially leading to more efficient and sustainable production of Quorn mycoprotein.
阔恩素肉蛋白是一种富含蛋白质的肉类替代品，通过对产毒镰刀菌进行大规模发酵生产。然而，产毒镰刀菌发酵过程中的一个主要挑战是持续出现称为菌落变体（C变体）的突变体。这些C变体具有高度分支的形态，最终导致最终产品不太理想，并使发酵过程提前终止。本研究旨在确定导致C变体形态的基因突变。
我们首先从商业发酵样品中分离出C变体和野生型菌株，并对其在固体培养基上的径向生长速率进行了表征。全基因组测序有助于在12株C变体分离株中的11株中鉴定出一个名为jg4843的基因中的突变，而在野生型分离株中未观察到这些突变。jg4843基因被鉴定为LRG1的直系同源基因，LRG1是一种Rho-GTPase激活蛋白，可调节影响真菌生长的Rho1信号通路。值得注意的是，jg4843中的突变主要位于负责LRG1活性的RhoGAP结构域。为了证实这些突变的作用，我们使用CRISPR/Cas9介导的同源定向重组将C变体突变引入野生型分离株，成功重现了特征性的C变体形态。
本研究确定LRG1直系同源基因jg4843中的突变是商业发酵产毒镰刀菌分离株中C变体形态的遗传原因。了解这一遗传基础为制定防止C变体出现的策略铺平了道路，这可能会使阔恩素肉蛋白的生产更高效、更可持续。
