Srisuksam Chettida, Punya Juntira, Wattanachaisaereekul Songsak, Toopaang Wachiraporn, Cheevadhanarak Supapon, Tanticharoen Morakot, Amnuaykanjanasin Alongkorn
National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA), 113 Thailand Science Park, Phayonyothin Rd., Tambon Khlong Nueng, Amphoe Khlong Luang, Pathum Thani 12120, Thailand.
School of Bioresources and Technology, King Mongkut's University of Technology Thonburi (KMUTT), Bangkok 10140 Thailand.
FEMS Microbiol Lett. 2018 Aug 1;365(15). doi: 10.1093/femsle/fny131.
The reducing clade IIb polyketide synthase gene, pks14, is preserved throughout the evolution of entomopathogenic fungi. We examined the functions of pks14 in Beauveria bassiana using targeted gene disruption, and pks14 disruption was verified by Southern blot and PCR analyses. The radial growth, cell dry weight and conidial germination of Δpks14 were comparable to that of the wild type. Our sequence and gene expression analyses of the pks14 biosynthetic cluster demonstrated: (i) cotranscription and constitutive expression of nearly all the genes of the aforementioned cluster including the C2H2 zinc finger transcription regulator gene, but not pks14 and the cytochrome P450 gene; (ii) expression of the pks14 gene in the insect-containing culture condition only; and (iii) a KAR9-like gene in direct proximity with pks14 is the only gene showing co-regulation. The Δpks14-infected Spodoptera exigua larvae survived significantly longer than those infected by the wild type, indicating a marked reduction in the virulence of Δpks14 against the insect. LT50 of Δpks14 was increased by 1.55 days. Hyphal body formation was decreased in the hemolymph of insects infected by Δpks14 as compared with those inoculated by the wild type. Our results suggest that PKS14-catalyzed polyketide enhances virulence and pathogenicity of B. bassiana on insects.
还原性进化枝IIb聚酮合酶基因pks14在昆虫病原真菌的整个进化过程中都得以保留。我们利用靶向基因敲除技术研究了球孢白僵菌中pks14的功能，并通过Southern杂交和PCR分析验证了pks14的敲除。Δpks14的径向生长、细胞干重和分生孢子萌发与野生型相当。我们对pks14生物合成簇的序列和基因表达分析表明：(i)上述簇中几乎所有基因（包括C2H2锌指转录调节基因）共转录且组成型表达，但pks14和细胞色素P450基因除外；(ii)pks14基因仅在含昆虫的培养条件下表达；(iii)紧邻pks14的一个类似KAR9的基因是唯一显示出共调节的基因。被Δpks14感染的甜菜夜蛾幼虫存活时间明显长于被野生型感染的幼虫，这表明Δpks14对昆虫的毒力显著降低。Δpks14的LT50增加了1.55天。与接种野生型的昆虫相比，被Δpks14感染的昆虫血淋巴中菌丝体的形成减少。我们的结果表明，PKS14催化的聚酮增强了球孢白僵菌对昆虫的毒力和致病性。
