Identification of three ubiquitin genes of the rice blast fungus Magnaporthe grisea, one of which is highly expressed during initial stages of plant colonisation

Abstract

Differential cDNA screening was used to identify genes expressed during the colonisation of rice leaves by the pathogenic fungus Magnaporthe grisea. This led to the identification of a gene, called UEP1, which encodes a ubiquitin extension protein. UEP1 was highly expressed 48 h after initial fungal infection of rice leaves when M. grisea is proliferating in the leaf epidermis but not yet causing disease symptoms. UEP1 appeared to be down-regulated after this time despite further extensive growth of the fungus throughout the leaf tissue. To investigate the potential role of ubiquitin in fungal pathogenesis we subsequently isolated UEP3 and PUB4, encoding a second ubiquitin extension protein and a polyubiquitin respectively. UEP1 was expressed abundantly during active growth of M. grisea in axenic culture but was down-regulated by starvation-stress. UEP3 showed a similar pattern of expression to UEP1 during the growth of M. grisea in culture and after environmental stress, but was not highly expressed during plant colonisation. PUB4 was highly expressed after environmental stress, but was not highly expressed during plant colonisation. UEP1 was found to be present in a much-higher copy number per haploid genome compared to UEP3 and PUB4. The restricted high-level expression of UEP1 suggests that M. grisea undergoes rapid ribosomal biogenesis and protein turnover during initial plant-tissue colonisation, which is regulated by a specific UEP1-encoded component of the M. grisea ubiquitin gene family.