Introduction
Extracellular vesicles (EVs)
mediate cell-cell-dependent environmental responses (Rodrigues et al.,
2015; Anand et al., 2019, Baldrich et al., 2019). Numerous studies have
isolated and characterized fungal pathogen EVs and found that they
produce phytotoxic effects, among other things, when inoculated on plant
tissues such as leaves (Bleackley et al., 2020). This suggests that
cargo within these vesicles can be disease-promoting. How this cargo is
sorted is currently elusive in plant pathogenic fungi, which limits our
understanding of virulence factors transmitted through non-canonical
pathways. A repertoire of EV constituents, including nucleic acids,
proteins, and secondary metabolites often reflect the pathophysiological
state of the cell from which EVs are secreted. These cargos can be taken
up by other cells naked or enclosed in the EV membrane to yield the
different EV-mediated physiological states in the acceptor cells (Koch
et al., 2020). As EVs carry various bioactive molecules, many of which
facilitate cell-cell communication (Rodrigues et al., 2015; Anand et
al., 2019), they have also been implicated in plant-fungal interaction
and pathogenesis. For instance, EVs of several species of fungi have
been shown to traffic virulence factors, such as cell wall degrading
enzymes, protein effectors, and toxins with phytotoxic effects on their
plant host tissues (Bleackley et al., 2020; Costa et al., 2021; Regente
et al., 2017).
However,
there is still a lack of understanding about the regulatory mechanisms
involved in cargo sorting, packaging and trafficking of the respective
vesicles, especially in filamentous fungi.
The ESCRT (e ndosomal s orting c omplexr equired for t ransport) pathway is utilized by many
eukaryotes including fungi whereby it is involved in the sorting and
trafficking of molecules within the endosomal system. This system is
involved in the formation and sorting of endosomal vesicles, which are
part of the endocytic pathway responsible for internalizing molecules
from the cell surface and delivering them to specific destinations
within the cell. In filamentous fungi, the ESCRT pathway plays a role in
the biogenesis of filamentous growth and pathogenesis (Zheng et al.,
2018; Sun et al., 2022). Studies have shown that the ESCRT pathway is
required for the conventional organization of the fungal cytoskeleton
(An et al., 2006; Henne et al., 2013), which is essential for the
formation of hyphae. The ESCRT pathway also plays a role in the
splitting of daughter nuclei during cell division and the sorting of
proteins destined for the plasma membrane (Henne et al., 2013).
Additionally, the ESCRT pathway is involved in the formation of
endosomal vesicles that are important for nutrient uptake and stress
response (Fan et al., 2015; Wang et al., 2020). In pathogenic fungi, the
ESCRT pathway also plays a role in the secretion of virulence factors
and evasion of host immunity (Regente et al., 2017; Martínez-López et
al., 2022).
The ESCRT pathway is vital for endocytosis whereby it allows for the
packaging of extracellular materials and membrane proteins into EVs for
trafficking into the cytoplasm (Fig 1(A) , Ahmed et al., 2019).
Endocytosis is characterized by the presence of late endosomes or
multivesicular bodies (MVBs), which harbour intraluminal vesicles (ILVs)
that usually arise from the invagination and budding of the endosomal
membrane (Haag et al., 2015; Ahmed et al., 2019). The process of
endocytosis is fundamental to various cellular process ranging from
signal transduction to morphogenesis (Schimid et al., 2014). In certain
fungal species, endocytosis also plays a role during interaction with
plants including in the apical growth of hyphae (Toshima et al., 2006;
Bielska et al., 2014). In addition to growth and development,
ESRCT-based regulation of cellular function also appears to be crucial
for adaptation and response to both external and internal stimuli such
as biotic and abiotic stress factors (Mosesso et al., 2019; Rosa et al.,
2020). Consequently, many of the basic components of the ESCRT pathway
are conserved across eukaryotes, albeit with notable lineage-specific
adaptations in some taxa (Leung et al., 20008).
Most of the significant contributions to our understanding of ESCRT
derive strongly from studies on model organisms (Thompson et al., 2005;
Vaccari & Bilder, 2005; Herz et al., 2006; Spitzer et al., 2006).
Adding to these are several studies highlighting the key roles of ESCRT
proteins in mammals (Pornillos et al., 2002; Xu et al., 2003; Skibinski
et al., 2005; Parkinson et al., 2006; Saksena and Emr, 2009; Stuffers et
al., 2009; Hurley et al., 2015). The ESRCT pathway was originally
discovered in Archaeal cytokinesis (Lindas et al., 2008; Samson et al.,
2008) and in fungi it was first discovered in the model fungusSaccharomyces cerevisiae (Emr et al., 2001). In this yeast, the
ESCRT pathway’s constituents were named based on their functions in
sorting ubiquitinated membrane proteins into lysosome/vacuole lumens for
degradation (Katzmann et al., 2001; Babst et al., 2002a; Xie et al.,
2019b). The pathway represents a complex endomembrane system that
consists of five complexes, namely ESCRT-0, -I, -II -III and Vps4
(v acuolar p rotein s orting 4 ). Together with
various accessory proteins, the individual elements of the pathway act
in concert to form MVBs during diverse processes including cytokinesis,
membrane repair and autophagy (Roxrud et al., 2010; Henne et al., 2013;
Hurley et al., 2015).
As in other eukaryotes, fungal MVBs are critical for transporting
ubiquitinated membrane proteins to the vacuole with the aid of the ESCRT
machine, which both recognizes and packages these ubiquitin-modified
proteins into the ILVs contained inside MVBs (An et al., 2006; Hurley
and Hanson, 2010; Henne et al., 2011). Studies in yeast also showed that
ILVs form when MVB membranes evaginate and undergo fission (Ahmed et
al., 2019; Anand et al., 2019). Others have reported that impairment in
the ESCRT apparatus can lead to reduced formation of ILVs and aberrant
endosomal compartments (Raymond et al., 1992; Xie et al, 2019a). In the
filamentous fungal pathogen, Fusarium graminearum , such defects
can significantly impact cellular processes like deoxynivalenol
production, growth and pathogenicity, as well as sexual and asexual
reproduction (Xie et al. 2019a). However, details regarding this pathway
have been considered in only a few phytopathogens, most notablyMagnaporthe oryzae and F. graminearum (Oh et al., 2012;
Xie et al., 2016; Cheng et al., 2018; Xie et al., 2019a, b; Que et al.,
2019). Currently, most fungal work pertaining to the ESCRT pathway is
focused on human pathogenic yeasts including Cryptococcus
neoformans and Candida albicans (Godinho et al., 2014; Hu et
al., 2015; Zhang et al., 2015; Park et al., 2020). The elucidation of
the core functions of the ESCRT pathway in fungal plant pathogens
represents a critical aspect given the functional importance of this
pathway. Therefore, this review reflects on the recent ESCRT discoveries
as they relate to filamentous fungi and touch on the relevance of
released EVs, which are the end-products of ESCRT, in fungal biology and
fungal-host interactions.