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Most Fusarium species produce bikaverin and aurofusarin for mycelium pigmentation and fusarubins for perithecial pigmentation [1]. However, Fusarium solani produces fusarubins during mycelial growth and another unknown pigment during sexual reproduction. This unknown pigment is predicted to be biosynthesized by a non-reducing polyketide synthase (PKS35 = PKSN [2]). PKS35 encodes a 2106 aa protein and share 52% identity to PhnA from P. herquei, which catalyzes the first step of the herqueinone biosynthetic pathway [3]. Genetic analyses of the PKS35 gene cluster suggests that it contains eleven genes, of which four are present in the herqueinone cluster. Herqueinone contains tricyclic phenalenone core ring structure cyclized by a FAD-dependent monooxygenase. To unravel the biosynthetic products in the PKS35 pathway the intron stripped PKS was cloned and put under control of a galactose inducible promoter in a 2µ vector, which was transformed into a Saccharomyces cerevisiae strain co-expressing a Sfp-Type 4′-Phosphopantetheinyl Transferase (Ppt1). The transformed yeast strain was cultivated under induced conditions in liquid cultures for five days and before production of secondary metaboliteswas analyzed by high-resolution mass spectrometry (HRMS). The results showed that the yeast strain was able to produce prephenalenone, which is also the first step of the herqueinone pathway. We also detected dehydroxyprephenalenone, which is formed through spontaneous dehydration [3]. Through heterologous expression of the other genes in the cluster, we will follow the biosynthetic route and ultimately unravel the entire pathway of the perithecial pigment of F. solani.[1] Studt L, Wiemann P, Kleigrewe K et al (2012): Appl Environ Microbiol, 78: 4468-4480.[2] Graziani S, Vasnier C, Daboussi MJ (2004): Appl Environ Microbiol, 70: 2984-2988.[3] Gao SS, Duan A, Xu W et al (2016): J Am Chem Soc, 138: 4249-4259.

Fungicide application remains a vital component in the management of Fusarium-induced plant infections, but environmental concerns and resistance toward some of the commonly used fungicides has spurred efforts into developing new specific fungicides. Phenamacril, a cyanoacrylate fungicide, represents a novel class of seemingly Fusarium-specific and environmentally benign fungicides that could hold great potential in our future efforts toward minimizing the occurrence and severity of Fusarium-induced plant infections. Phenamacril targets the motor domain of Fusarium class I myosin [1-2], an ubiquitous molecular motor which facilitates numerous ATP-driven and actin-associated processes within the cell. The myosin superfamiliy is divided into approximately 20 different classes (classes I, II, V and XVII are present in fungi), each potentially having several isoforms. While the taskspecific C-terminal tail domains are highly variable in size and function, the N-terminal motor domains remain highly conservered, both structurally and functionally. Interestingly, Phenamacril-resistance has been shown to correlate with single amino acid mutations in a highly conserved region of the motor domain [1], a region involved in the allosteric communication between the actin- and nucleotide binding-site [3]. To gain further insight into the specificity, the inhibitory potential and nature of the Phenamacril-mediated inhibitionof Fusarium myosin, we overexpressed Fusarium class I myosins in Sf9 insect cells. Through a combination of NADH-coupled and in vitro motility assays, we demonstrated that Phenamacril is a very potent but functionally reversible inhibitor of Fusarium class I myosin. Similarly, we demonstrated that Phenamacril had no or only minimal inhibitory effect on human myosin 1c, Dictyostelium discoideum class I and II myosins, but that the motor domain of the Phenamacril-resistant F. solani f. sp. pisi was highly inhibited by Phenamacril in vitro. This suggests thatdespite of the high degree of structural conservation in the myosin super-family, subtle differences render Phenamacril a highly specific and potent reversible inhibitor of Fusarium class I myosin.[1] Zheng Z, Yiping H, Yiqiang C et al (2015): Sci Rep, 5 (2021): 8248.[2] Zhang C, Yun C, Yanni Y et al (2015): Environ Microbiol, 17 (8): 2735–46.[3] Krishna C, Taft MH, Martin R et al (2011): J Biol Chem, 286 (34): 29700–708.