amphotericin B (Richie et al. 2009), and resistance towards voriconazole was further
reduced by the parallel deletion of derA (Richie
et al. 2011a). Interestingly, hrdA deletion
mutants show reduced voriconazole sensitivity
(Krishnan et al. 2013). Deletion of ormA,
encoding a protein involved in sphingolipid
synthesis, results in increased susceptibility to
azoles, whereas overexpression reduced azole
sensitivity. Expression of ormA is induced by
TM in a hacA-dependent manner, further corroborating the requirement of an intact ER
stress response for antifungal drug resistance
(Zhai et al. 2019).
2. Cryptococcus neoformans
Cryptococcus neoformans is a basidiomycetous
yeast and an opportunistic pathogen of immunosuppressed patients. Basidiospores serve as
predominant infectious propagules. After initially colonizing lung tissue C. neoformans can
subsequently spread and cause cryptococcal
meningitis, often as a secondary infection of
HIV-positive individuals. In C. neoformans,
the UPR has been studied with respect to
virulence-associated traits, such as stress resistance and thermotolerance, but also concerning
antifungal drug resistance and sexual mating.
While the ortholog of Ire1 in C. neoformans
was identified by straightforward BLAST
search, the identification of the first Hac1
ortholog in basidiomycetes required experimental validation (Cheon et al. 2011). Monitoring of potential unconventional splicing events
of five candidate genes encoding bZIP transcription factors with significant homology of
their bZIP domains to Hac1 identified Hxl1
(Hac1and XBP1-like gene 1) as the key UPR
regulator in C. neoformans. Splicing of the
unprocessed HXL1 mRNA results in the Ire1dependent removal of a 56 nt intron and
expression of Hxl1. Expression of the intronless
HXL1 cDNA fully restores ER stress resistance
of IRE1 deletion mutants, confirming Hxl1 as
the functional Hac1 ortholog. Interestingly,
expression of Hxl1 in S. cerevisiae DHAC1
mutants does not restore ER stress resistance,
indicating divergent transcriptional machineries involved in Hac1- and Hxl1-dependent
ER stress responses in either organism as causal
for the functional incompatibility. Besides their
requirement for ER stress resistance, IRE1 and
HXL1 deletion mutants are strongly suppressed
in thermotolerance and fail to grow at mammalian body temperature, resulting in avirulence
in mice infection experiments (Cheon et al.
2011). Resolution of ER stress during growth
at host temperature involves the major mRNA
deadenylase Ccr4 (Carbon Catabolite Repression 4), which is a central component of
CCR4-NOT complex mediating mRNA decay
(Havel et al. 2011). In addition, the mRNA
binding protein Puf4 (Pumilio homology
domain Family 4) destabilizes HXL1 mRNA
and therefore contributes to UPR dynamics
and ER stress resistance. Deletion of PUF4 alleviates growth at host temperature but does not
affect virulence (Glazier et al. 2015). Overexpression of the central ER chaperone Kar2/
Bip1 partly rescues ER stress resistance and
thermotolerance of HXL1 and IRE1 deletion
mutants (Jung et al. 2013). Ire1 and Hxl1 play
conserved roles in ER stress resistance in Cryptococcus species. Interestingly, antifungal drug
resistance towards azole derivatives and fludioxonil is in general decreased but differentially affected in DIRE1 and DHXL1 mutants,
with divergent effects in the C. neoformans
(H99), C. deneoformans (JEC21), and C. deutrogattii (R265) (Jung et al. 2018) backgrounds. In
addition, IRE1 but not HXL1 deletion mutants
are unable to undergo sexual mating, as
reflected by reduced cell fusion and conjugation tube formation. This defect is connected to
increased expression of the pheromone precursor gene MFa1 and an aberrant localization of
the Ste6 pheromone transporter and in part
suppressed by Kar2 overexpression in the
DIRE1 mutant background (Jung et al. 2016).
3. Candida albicans
The ascomycetous yeast Candida albicans is a
commensal of the human mucosa and opportunistic pathogen in immunosuppressed individuals. The ability to colonize the host is
connected to a yeast-to-hypha dimorphic tran3 Genetics of the Unfolded Protein Response in Fungi
57
reduced by the parallel deletion of derA (Richie
et al. 2011a). Interestingly, hrdA deletion
mutants show reduced voriconazole sensitivity
(Krishnan et al. 2013). Deletion of ormA,
encoding a protein involved in sphingolipid
synthesis, results in increased susceptibility to
azoles, whereas overexpression reduced azole
sensitivity. Expression of ormA is induced by
TM in a hacA-dependent manner, further corroborating the requirement of an intact ER
stress response for antifungal drug resistance
(Zhai et al. 2019).
2. Cryptococcus neoformans
Cryptococcus neoformans is a basidiomycetous
yeast and an opportunistic pathogen of immunosuppressed patients. Basidiospores serve as
predominant infectious propagules. After initially colonizing lung tissue C. neoformans can
subsequently spread and cause cryptococcal
meningitis, often as a secondary infection of
HIV-positive individuals. In C. neoformans,
the UPR has been studied with respect to
virulence-associated traits, such as stress resistance and thermotolerance, but also concerning
antifungal drug resistance and sexual mating.
While the ortholog of Ire1 in C. neoformans
was identified by straightforward BLAST
search, the identification of the first Hac1
ortholog in basidiomycetes required experimental validation (Cheon et al. 2011). Monitoring of potential unconventional splicing events
of five candidate genes encoding bZIP transcription factors with significant homology of
their bZIP domains to Hac1 identified Hxl1
(Hac1and XBP1-like gene 1) as the key UPR
regulator in C. neoformans. Splicing of the
unprocessed HXL1 mRNA results in the Ire1dependent removal of a 56 nt intron and
expression of Hxl1. Expression of the intronless
HXL1 cDNA fully restores ER stress resistance
of IRE1 deletion mutants, confirming Hxl1 as
the functional Hac1 ortholog. Interestingly,
expression of Hxl1 in S. cerevisiae DHAC1
mutants does not restore ER stress resistance,
indicating divergent transcriptional machineries involved in Hac1- and Hxl1-dependent
ER stress responses in either organism as causal
for the functional incompatibility. Besides their
requirement for ER stress resistance, IRE1 and
HXL1 deletion mutants are strongly suppressed
in thermotolerance and fail to grow at mammalian body temperature, resulting in avirulence
in mice infection experiments (Cheon et al.
2011). Resolution of ER stress during growth
at host temperature involves the major mRNA
deadenylase Ccr4 (Carbon Catabolite Repression 4), which is a central component of
CCR4-NOT complex mediating mRNA decay
(Havel et al. 2011). In addition, the mRNA
binding protein Puf4 (Pumilio homology
domain Family 4) destabilizes HXL1 mRNA
and therefore contributes to UPR dynamics
and ER stress resistance. Deletion of PUF4 alleviates growth at host temperature but does not
affect virulence (Glazier et al. 2015). Overexpression of the central ER chaperone Kar2/
Bip1 partly rescues ER stress resistance and
thermotolerance of HXL1 and IRE1 deletion
mutants (Jung et al. 2013). Ire1 and Hxl1 play
conserved roles in ER stress resistance in Cryptococcus species. Interestingly, antifungal drug
resistance towards azole derivatives and fludioxonil is in general decreased but differentially affected in DIRE1 and DHXL1 mutants,
with divergent effects in the C. neoformans
(H99), C. deneoformans (JEC21), and C. deutrogattii (R265) (Jung et al. 2018) backgrounds. In
addition, IRE1 but not HXL1 deletion mutants
are unable to undergo sexual mating, as
reflected by reduced cell fusion and conjugation tube formation. This defect is connected to
increased expression of the pheromone precursor gene MFa1 and an aberrant localization of
the Ste6 pheromone transporter and in part
suppressed by Kar2 overexpression in the
DIRE1 mutant background (Jung et al. 2016).
3. Candida albicans
The ascomycetous yeast Candida albicans is a
commensal of the human mucosa and opportunistic pathogen in immunosuppressed individuals. The ability to colonize the host is
connected to a yeast-to-hypha dimorphic tran3 Genetics of the Unfolded Protein Response in Fungi
57