FUNCTIONAL STUDY OF LIPOXYGENASE-MEDIATED RESISTANCE AGAINST ASPERGILLUS FLAVUS AND FUSARIUM VERTICILLIOIDES INFECTION IN MAIZE

Mycotoxin contamination of maize kernels by fungal pathogens Aspergillus\r\nflavus (Af) and Fusarium verticillioides (Fv) is a chronic global challenge\r\nimpacting food security, health and trade. Current disease management\r\npractices are proven inadequate and strategies gearing towards hostmediated\r\nresistance can be an effective and sustainable approach to\r\nstrengthen efforts to control the pathogens. Lipid modification into a\r\ndiverse array of compounds, oxylipins, are recognized in response of plants\r\nto fungal pathogens. Lipoxygenase genes (LOXs) play a crucial role in the\r\nenzymatic oxidation of polyunsaturated fatty acids (PUFAs) into 9(S)- and\r\n13(S)-hydroperoxides that are further metabolized into different oxylipins\r\nincluding jasmonates, oxo- and keto-fatty acids and volatiles. The\r\ninvolvement of maize LOXs (ZmLOXs) in this respect has been a subject of\r\nstudies and their genetic manipulation resulted in the alteration of\r\nresistance or susceptibility to fungal pathogens in maize. However, the\r\nmaize genome encodes six 9-LOX and seven 13-LOX isoforms and the specific\r\nrole of each isoform remains elusive. The current study investigated the\r\nrole of ZmLOXs in host resistance against the fungi Af and Fv using in\r\nsilico and in planta approaches. The phylogenetic relationship, sequence\r\nsimilarity, protein domain structures, and transcript level structural\r\nvariations were explored by comparing publicly available maize pan-genomes.\r\nFurthermore, the role of ZmLOXs against Af and Fv infection was\r\ninvestigated through their expression analysis along with further key genes\r\ninvolved in oxylipin biosynthesis, mycotoxin accumulation and lipid\r\nprofiles in a ZmLOX4 mutant line (UFMulox4) together with W22, Mo17 and\r\nTzi18 inbred lines at 3- and 7-days post-inoculation (dpi). ZmLOX proteins\r\nshowed considerable variations in their sequences, functional domain\r\nstructure and transcript structural variations among the pan-genome\r\nmembers. Among the studied lines, Tzi18 showed the highest resistance to\r\nthe pathogens coupled with the lowest mycotoxin accumulation, while the\r\nmutant, UFMulox4, was highly susceptible to both pathogens with the most\r\nelevated content of mycotoxins. Fv inoculation determined a stronger\r\ninduction of ZmLOXs and maize allene oxide synthase genes as compared to Af\r\n. Higher constitutive levels of 9-LOXs genes, ZmLOX1 and ZmLOX2 and an\r\ninduction of ZmLOX4 were recorded in Tzi18, while the upregulation of ZmLOX1\r\nand ZmLOX4 observed in Mo17 might have larger active role in resistance\r\nagainst Fv. Liquid chromatography-mass spectrometry further revealed an\r\nincreased accumulation of the linoleic (18:2) derived 9-cyclopentenone, 10-\r\noxo-11-phytoenoic acid (10-OPEA), in Fv inoculated kernels of Tzi18 and\r\nMo17, which was previously identified to inhibit fungal growth in vitro.\r\nHence, the results confirm the pivotal role of ZmLOXs in controlling the\r\nresistance mechanisms against these two pathogens.