Positive Selection is shaping the Evolution of two Aspartic Proteinases in phytopathogenic Botrytis species

REVUELTA MV; VAN KAN JAL; ARJEN TEN HAVE

Bahía Blanca

Congreso; 6to Congreso Argentino de Bioinformática y Biología Computacional; 2015

Asociacion Argentina de Bioinformática y Biología Computacional

p { margin-bottom: 0.08in; }a:link { }BackgroundAsignificantly higher nonsynonimous (dN) than synonimous (dS)substitution rate is a powerful landmark of positive Darwinianselection [1]. The rate ratio ω=dN/dSmeasures the magnitude and direction of selective pressure on aprotein or a site, with ω>1 indicating, positive selection [2]. It forms the basis fortests that are useful for the detection of genes related topathogenicity. Positive selection results in elevated geneticvariation and can play a significant role in the adaptation of apathogen to a new environment, among which the host species range[3]. Theobjective of this study was to identify the role of positiveselection in the evolution of Aspartic Proteinases (APs) from broadhost range phytopathogen Botrytiscinerea andrelated narrow host range Botrytisspecies.ResultsTwomembers of the Botrytissp AP family, constituting in between X and 15 members, are underpositive selection (AP8 and AP14). AP8 has been previously shown tobe highly expressed during early stages of fungal growth (23% of theearly secretome, [4]). BcAP8 knock-out mutants did show 80%extracellular reduced proteolytic activity but no differentialphenotype during infection [4]. The AP8 ortholog in Trichodermaharzianum,the major biocontrol agent of B.cinera, wassuggested to be involved in mycoparasitism [5] and a recent phylogenydid show all AP8 orthologs to be derived from fungi characterized byan important saprophytic phase in their lifecyle [6]. Residues24 and 340 are under positive selection and locate in the bindingcleft, which suggests a role in substrate binding / specificity. AP14clusters in a small subclade with fungal phytopathogen APs only.However, the BcAP14 knock out mutant shows wildtype virulence ontomato leaves at 72 hpi. All AP14 orthologs appear to have a normalsignal peptide but they lack the typical propeptide that is removedin order to activate the APs, suggesting they are active in thesecretory pathway. The lack of phenotype can be explained byredundancy. Residues 264and 298 show positive selection. 264 is located at the end of aninsert, which is particular to AP14 orthologs. 298 is located in thevicinity of the substrate binding cleft, in close contact withresidues 297, 293 and 287 that are part of the subsite.Further experiments are required to investigate the role of these APsduring the fungal life cycle. References[1]Li WH. 1997. Molecular evolution. Sunderland (MA): Sinauer AI.[2]Yang Z, Nielsen R. Estimatingsynonymous and nonsynonymous substitution rates under realisticevolutionary models.MolBiol Evol.2000;17(1):32-43.[3]Rowe HC, Kliebenstein DJ. Elevated genetic variation withinvirulence-associated Botrytis cinereapolygalacturonase loci. Mol Plant Microbe Interact.2007;20(9):1126-37.[4]ten Have A, Espino JJ, Dekkers E, Van Sluyter SC, Brito N, Kay J,González C, van Kan JA. The Botrytis cinereaaspartic proteinase family. Fungal Genet Biol. 2010Jan;47(1):53-65.[5]Suárez MB, Sanz L, Chamorro MI, Rey M, González FJ, Llobell A,Monte E. Proteomic analysis of secreted proteins from Trichodermaharzianum. Identification of a fungal cell wall-inducedaspartic protease. Fungal Genet Biol. 2005Nov;42(11):924-34.[6] Revuelta MV, van Kan J, Kay J, ten Have A. Extensive ExpansionofA1 Family Aspartic Proteinases in Fungi Revealed by EvolutionaryAnalyses of 107 Complete Eukaryotic Proteomes.GenomeBiol and Evol.2014 May;6(6):1480-1494.