Growth, Differentiation and Sexuality

(Raju 1992). When male-sterile mutants are used
as fertilizing parent, perithecial development
is initiated, but then arrested at an early stage.
In N. crassa, numerous female-sterile mutants
that do not form functional protoperithecia or
display a reduced fertility have been described.
Because of this high frequency of sterility in
female strains, it was suggested that at least 400
genes are required for perithecium and ascospore
development in N. crassa (Johnson 1978; Leslie and
Raju 1985). However, many of these female-sterile
mutants show abnormal vegetative growth, and
thus this female sterility may be a consequence
of a morphological defect, rather than a mutation
in a gene specifically needed for fruiting-body
differentiation (Raju 1992).
Effective molecular techniques developed for
N. crassa, such as the transformation and creation
of cosmid genomic libraries involving, for example,
phenotypic complementation, have led to the
cloning and functional characterization of regulatorygenesthataffectmorphology(Baileyand
Ebbole 1998). Other molecular genetic approaches
used subtractive hybridization to isolate N. crassa
sexual developmental genes.
With this attempt, Nelson and Metzenberg
(1992) succeeded in identifying 14 genes transcribed
only under nitrogen-depleting growth
conditions. One of these, asd-1,hasbeenshownto
encode a putative rhamnogalacturonase necessary
for ascus development (Nelson et al. 1997b).
In large-scale analyses, fruiting body-specific
expressed sequence tags (ESTs) from N. crassa
were sequenced for further molecular characterization
(Nelson et al. 1997a). In such experimental
approaches, isolated candidate genes have to be
inactivated to obtain detailed information about
mutant phenotypes and gene function.
Inactivation of developmental genes in N.
crassa was achieved either by homologous recombination
or by gene silencing via repeat-induced
point mutation (RIP). The N. crassa RIP process
efficiently detects and mutates both copies of a sequence
duplication. RIP acts during the dikaryotic
stage of the sexual cycle, causing numerous C:Gto-T:A
transitions within duplicated sequences,
and is frequently used to inactivate genes in N.
crassa (Galagan and Selker 2004). Finally, the
whole genome sequence of N. crassa has become
available, opening the opportunity for analyzing
genes involved in fruiting-body development by
means of reverse genetic approaches (Galagan
et al. 2003; Borkovich et al. 2004).
Fruiting Bodies in Ascomycetes 331
2. Sordaria macrospora
In the heterothallic species N. crassa, mutations
conferring male and/or female sterility can be
detected directly because of their sterility effects
in heterozygous crosses. However, recessive mutations
that affect post-fertilization perithecial
development will remain undetected in heterothallic
species until the mutant allele is available in
both mating types, thus allowing homozygous
crosses. In contrast to N. crassa, the homothallic
pyrenomycete S. macrospora is self-fertile, which
means that recessive mutations can directly be
tested for defects in fruiting-body development.
Moreover, S. macrospora produces only meiotically
derived ascospores, whereas asexual spores,
such as conidia, are absent. Thus, there is no
interference between two different developmental
programs, which makes it easier, for example, to
analyze differentially expressed genes involved in
ascocarp development. Under laboratory conditions,
perithecia and ascospores reach maturity
within 7 days after ascospore germination. During
this development, distinct reproductive structures,
such as ascogonia, protoperithecia (young fruiting
bodies), and perithecia, can be distinguished
(Fig. 16.2). Because S. macrospora represents
such a favorable genetic system for scientists, this
homothallic pyrenomycete was used to generate
numerous mutants that are blocked at various
stages of perithecial development (Esser and
Straub 1958; Masloff et al. 1999). Wild-type strains
of S. macrospora are self-fertile and produce
perithecia. However, fertile perithecia are also
formed in crosses between sterile strains, when
nuclei are interchanged by hyphal anastomoses
at the contact zones of two sterile mycelia. These
crosses facilitate the analysis of epistatic relationships
among developmental mutants. The
establishment of molecular tools provides the
basis for studying fruiting-body development in S.
macrospora (Walz and Kück 1995; Pöggeler et al.
1997).
3. Aspergillus nidulans
The plectomycete Aspergillus nidulans (teleomorph:
Emericella nidulans) propagates by the
formation of spores that can be either asexual or
sexual, and has long served as a model system for
understanding the genetic regulation of asexual
development in ascomycetes (Adams et al. 1998).
The asexual cycle is characterized by the produc-