further illustrated by the Aspergillus oryzae sirtuin HstD, which controls the expression of the
velvet complex member LaeA and thus the kojic
acid production and conidiation (Kawauchi
et al. 2013).
C. Histone Phosphorylation, Ubiquitination,
and Sumoylation
In addition to methylation and acetylation, further post-translational histone modifications
comprise phosphorylation, ubiquitination, or
sumoylation. These modifications were rather
analyzed in yeast than in filamentous fungi.
Histone phosphorylations by kinases that
transfer the phosphate group from ATP to the
hydroxyl group of the modified amino acid take
place at serines, threonines, or tyrosines and
are reversed by phosphatases. Phosphorylation
of H3S10 is crucial for yeast chromosome condensation and cell cycle progression during
mitosis and meiosis (Nowak and Corces 2004).
Ubiquitination of histones by ubiquitin
ligases is the covalent attachment of the small
modifier ubiquitin to lysine residues and is
reversed by deubiquitinating enzymes (DUBs,
see Sect. IV.B). Methylation of H3K4 is
mediated by the ubiquitination of H2BK123 in
yeast (Sun and Allis 2002). These concerted
histone modifications on distinct histone tails
are referred to as trans-tail regulation (Zheng
et al. 2010). Similar to ubiquitination, sumoylation is the covalent attachment of the small
ubiquitin-like modifier (SUMO) to lysines of
proteins through the activity of an E1-E2-E3
enzyme cascade. Sumoylation has been found
for all four histones and is associated with transcriptional repression. SUMO, which is
required for multicellular fungal development
in A. nidulans, most likely blocks the histone
modification sites to prevent acetylation or ubiquitination (Nathan et al. 2006; Harting et al.
2013). Co-purification experiments identified
SUMO-associated proteins connected to the
ubiquitin network involved in A. nidulans histone modification. They include one subunit of
the COMPASS complex required for histone
methylation as well as subunits of the SAGA
complex for histone acetylation (Harting et al.
2013). Like other post-translational modifications, ubiquitin and ubiquitin-like modifications do not only modify histones but also
other proteins and thereby can influence their
activity, function, localization, or stability and
affect fungal secondary metabolism and development (see Sects. III and IV).
D. Epigenetic Tools to Activate Silenced Gene
Clusters
The crucial role of histone modifications such
as methylation or acetylation in the regulation
of secondary metabolism can be utilized in the
laboratory for the identification of new secondary metabolites. Epigenetically silenced clusters
can be reactivated by changes in chromatin
modifications. Deletion of the genes encoding
HDACs, which are generally associated with
transcriptional repression, can lead to derepression of secondary metabolite gene clusters.
Accordingly, penicillin and sterigmatocystin
gene clusters of A. nidulans are activated
when the HDAC gene hdaA had been deleted
(Shwab et al. 2007). These types of experiments
have resulted in chemical epigenetics as an
emerging new research field (Okada and Seyedsayamdost 2017). The addition of HDAC or
methyltransferase inhibitors to fungal cultures
alters the epigenetic status of the cells to activate, identify, and study new secondary metabolite gene clusters and their products. The
advantage of this method is that the fungus
does not have to be genetically modified, and
it can therefore be applied to any organism.
Examples for such inhibiting chemicals are the
HDAC inhibitors valproic acid, trichostatin A
(see Sect. III.B) and its synthetic derivative suberoylanilide hydroxamic acid (SAHA), or the
DNA methyltransferase inhibitor 5-azacytidine
(5-AZA). Successful chemical epigenetics
approaches were applied in many different
fungi, such as A. alternata and P. expansum
(Shwab et al. 2007), Cladosporium cladosporioides and Diatrype disciformis (Williams et al.
2008), A. niger (Fisch et al. 2009; Henrikson
et al. 2009), and A. fumigatus (Magotra et al.
2017).
188
J. Gerke et al.
velvet complex member LaeA and thus the kojic
acid production and conidiation (Kawauchi
et al. 2013).
C. Histone Phosphorylation, Ubiquitination,
and Sumoylation
In addition to methylation and acetylation, further post-translational histone modifications
comprise phosphorylation, ubiquitination, or
sumoylation. These modifications were rather
analyzed in yeast than in filamentous fungi.
Histone phosphorylations by kinases that
transfer the phosphate group from ATP to the
hydroxyl group of the modified amino acid take
place at serines, threonines, or tyrosines and
are reversed by phosphatases. Phosphorylation
of H3S10 is crucial for yeast chromosome condensation and cell cycle progression during
mitosis and meiosis (Nowak and Corces 2004).
Ubiquitination of histones by ubiquitin
ligases is the covalent attachment of the small
modifier ubiquitin to lysine residues and is
reversed by deubiquitinating enzymes (DUBs,
see Sect. IV.B). Methylation of H3K4 is
mediated by the ubiquitination of H2BK123 in
yeast (Sun and Allis 2002). These concerted
histone modifications on distinct histone tails
are referred to as trans-tail regulation (Zheng
et al. 2010). Similar to ubiquitination, sumoylation is the covalent attachment of the small
ubiquitin-like modifier (SUMO) to lysines of
proteins through the activity of an E1-E2-E3
enzyme cascade. Sumoylation has been found
for all four histones and is associated with transcriptional repression. SUMO, which is
required for multicellular fungal development
in A. nidulans, most likely blocks the histone
modification sites to prevent acetylation or ubiquitination (Nathan et al. 2006; Harting et al.
2013). Co-purification experiments identified
SUMO-associated proteins connected to the
ubiquitin network involved in A. nidulans histone modification. They include one subunit of
the COMPASS complex required for histone
methylation as well as subunits of the SAGA
complex for histone acetylation (Harting et al.
2013). Like other post-translational modifications, ubiquitin and ubiquitin-like modifications do not only modify histones but also
other proteins and thereby can influence their
activity, function, localization, or stability and
affect fungal secondary metabolism and development (see Sects. III and IV).
D. Epigenetic Tools to Activate Silenced Gene
Clusters
The crucial role of histone modifications such
as methylation or acetylation in the regulation
of secondary metabolism can be utilized in the
laboratory for the identification of new secondary metabolites. Epigenetically silenced clusters
can be reactivated by changes in chromatin
modifications. Deletion of the genes encoding
HDACs, which are generally associated with
transcriptional repression, can lead to derepression of secondary metabolite gene clusters.
Accordingly, penicillin and sterigmatocystin
gene clusters of A. nidulans are activated
when the HDAC gene hdaA had been deleted
(Shwab et al. 2007). These types of experiments
have resulted in chemical epigenetics as an
emerging new research field (Okada and Seyedsayamdost 2017). The addition of HDAC or
methyltransferase inhibitors to fungal cultures
alters the epigenetic status of the cells to activate, identify, and study new secondary metabolite gene clusters and their products. The
advantage of this method is that the fungus
does not have to be genetically modified, and
it can therefore be applied to any organism.
Examples for such inhibiting chemicals are the
HDAC inhibitors valproic acid, trichostatin A
(see Sect. III.B) and its synthetic derivative suberoylanilide hydroxamic acid (SAHA), or the
DNA methyltransferase inhibitor 5-azacytidine
(5-AZA). Successful chemical epigenetics
approaches were applied in many different
fungi, such as A. alternata and P. expansum
(Shwab et al. 2007), Cladosporium cladosporioides and Diatrype disciformis (Williams et al.
2008), A. niger (Fisch et al. 2009; Henrikson
et al. 2009), and A. fumigatus (Magotra et al.
2017).
188
J. Gerke et al.