Cells of all living organisms are programmed to self-destruct under certain conditions. The most well known form of programmed cell death is apoptosis, which is essential for proper development in higher eukaryotes. In fungi, apoptotic-like cell death occurs naturally during aging and reproduction, and can be induced by environmental stresses and exposure to toxic metabolites. The core apoptotic machinery in fungi is similar to that in mammals, but the apoptotic network is less complex and of more ancient origin. Only some of the mammalian apoptosis-regulating proteins have fungal homologs, and the number of protein families is drastically reduced. Expression in fungi of animal proteins that do not have fungal homologs often affects apoptosis, suggesting functional conservation of these components despite the absence of protein-sequence similarity. Functional analysis of Saccharomyces cerevisiae apoptotic genes, and more recently of those in some filamentous species, has revealed partial conservation, along with substantial differences in function and mode of action between fungal and human proteins. It has been suggested that apoptotic proteins might be suitable targets for novel antifungal treatments. However, implementation of this approach requires a better understanding of fungal apoptotic networks and identification of the key proteins regulating apoptotic-like cell death in fungi.

Necrotrophic fungi are unable to occupy living plant cells. How such pathogens survive first contact with living host tissue and initiate infection is therefore unclear. Here, we show that the necrotrophic grey mold fungus Botrytis cinerea undergoes massive apoptotic-like programmed cell death (PCD) following germination on the host plant. Manipulation of an anti-apoptotic gene BcBIR1 modified fungal response to PCD-inducing conditions. As a consequence, strains with reduced sensitivity to PCD were hyper virulent, while strains in which PCD was over-stimulated showed reduced pathogenicity. Similarly, reduced levels of PCD in the fungus were recorded following infection of Arabidopsis mutants that show enhanced susceptibility to B. cinerea. When considered together, these results suggest that Botrytis PCD machinery is targeted by plant defense molecules, and that the fungal anti-apoptotic machinery is essential for overcoming this host-induced PCD and hence, for establishment of infection. As such, fungal PCD machinery represents a novel target for fungicides and antifungal drugs.

Studies conducted in the early 1990s showed for the first time that Saccharomyces cerevisiae can undergo cell death with hallmarks of animal apoptosis. These findings came as a surprise, since suicide machinery was unexpected in unicellular organisms. Today, apoptosis in yeast is well-documented. Apoptotic death of yeast cells has been described under various conditions and S. cerevisiae homologs of human apoptotic genes have been identified and characterized. These studies also revealed fundamental differences between yeast and animal apoptosis; in S. cerevisiae apoptosis is mainly associated with aging and stress adaptation, unlike animal apoptosis, which is essential for proper development. Further, many apoptosis regulatory genes are either missing, or highly divergent in S. cerevisiae. Therefore, in this review we will use the term apoptosis-like programed cell death (PCD) instead of apoptosis. Despite these significant differences, S. cerevisiae has been instrumental in promoting the study of heterologous apoptotic proteins, particularly from human. Work in fungi other than S. cerevisiae revealed differences in the manifestation of PCD in single cell (yeasts) and multicellular (filamentous) species. Such differences may reflect the higher complexity level of filamentous species, and hence the involvement of PCD in a wider range of processes and life styles. It is also expected that differences might be found in the apoptosis apparatus of yeast and filamentous species. In this review we focus on aspects of PCD that are unique or can be better studied in filamentous species. We will highlight the similarities and differences of the PCD machinery between yeast and filamentous species and show the value of using S. cerevisiae along with filamentous species to study apoptosis.

A considerable number of fungal homologues of human apoptotic genes have been identified in recent years. Nevertheless, we are far from being able to connect the different pieces and construct a primary structure of the fungal apoptotic regulatory network. To get a better picture of the available fungal components, we generated an automatic search protocol that is based on protein sequences together with a domain-centred approach. We used this protocol to search all the available fungal databases for domains and homologues of human apoptotic proteins. Among all known apoptotic domains, only the BIR [baculovirus IAP (inhibitor of apoptosis protein) repeat] domain was found in fungi. A single protein with one or two BIR domains is present in most (but not all) fungal species. We isolated the BIR-containing protein from the grey mould fungus Botrytis cinerea and determined its role in apoptosis and pathogenicity. We also isolated and analysed BcNMA, a homologue of the yeast NMA11 gene. Partial knockout or overexpression strains of BcBIR1 confirmed that BcBir1 is anti-apoptotic and this activity was assigned to the N′-terminal part of the protein. Plant infection assays showed that the fungus undergoes massive PCD (programmed cell death) during early stages of infection. Further studies showed that fungal virulence was fully correlated with the ability of the fungus to cope with plant-induced PCD. Together, our result show that BcBir1 is a major regulator of PCD in B. cinerea and that proper regulation of the host-induced PCD is essential for pathogenesis in this and other similar fungal pathogens.

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