Cloning and disruption of pgx4 encoding an in planta expressed exopolygalacturonase from Fusarium oxysporum

Fusarium oxysporum f. sp, lycopersici, the causal agent of tomato vascular wilt, produces an array of pectinolytic enzymes, including at least two exo -cll,4-polygalacturonases (exoPGs), A gene encoding an exoPG, pgx4, was iso lated with degenerate polymerase chain reaction primers derived from amino acid sequences conserved in two fungal exoPGs, pgx4 encodes a 454 amino aci d polypeptide with nine potential N-glycosylation sites and a putative 21 a mino acid N-terminal signal peptide. The deduced mature protein has a calcu lated molecular mass of 47.9 kDa, a pI of 8.0, and 51 and 49% identity with the exoPGs of Cochliobolus carbonum and Aspergillus tubingensis, respectiv ely, The gene is present in a single copy in different formae speciales of F. oxysporum. Expression of pgx4 was detected during in vitro growth on pec tin, polygalacturonic acid, and tomato vascular tissue and in roots and ste ms of tomato plants infected by F. oxysporum f. sp. lycopersici, Two mutant s of F. oxysporum f. sp. lycopersici with a copy of pgx4 inactivated by gen e replacement were as virulent on tomato plants as the wild-type strain.