Biology Reference
In-Depth Information
4.3. Rescue of miRNA mutants by target heterozygosity
Another general way to test for genetic interactions is to ask whether the
mutant phenotype associated with one locus can be suppressed by modula-
tion of another locus. In particular, cases of strong modification of one
mutant by heterozygosity of another mutation can serve as compelling
evidence to link the respective gene functions. This is especially the case
as relatively few genes exhibit obvious phenotypes when only one allele is
lost. This test, either by heterozygosity of classical mutant alleles of potential
targets or by RNAi-mediated knockdown, has proven to be one of the
cornerstones of linking specific target genes to miRNAs in
Drosophila
(
Fig. 8.3
A). As mentioned, finding “the” target gene of a miRNA is a
challenging if not potentially futile task, given that most conserved miRNAs
have tens if not hundreds of conserved targets. If the biological role of a
miRNA is to slightly repress 100 equivalent targets to tune the transcrip-
tome of a given cell, then one would not expect to observe genetic
interactions with any individual target. However, if there are specific targets
of particular genetic importance, this situation may be fulfilled. Especially
compelling would be the rescue of a miRNA mutant by heterozygosity of a
given target, which would suggest that derepression of that target plays an
important role in the etiology of the miRNA mutant phenotype.
Although it might be imagined that this is an exceptional genetic situa-
tion, in fact it has proven to be rather common in
Drosophila
(
Smibert and
Lai, 2010
). We have already mentioned how rescue of
mir-9a
and
mir-279
mutant phenotypes was achieved by target heterozygosity. Beyond these,
metabolic defects in
mir-278
mutants, including insulin resistance concom-
itant with elevated circulating sugar, could be suppressed by heterozygosity
for the FERM domain protein encoded by
expanded
(
Teleman
et al
., 2006
).
NMJ morphology defects observed in a
let-7
,
mir-125
double mutants could
be rescued by heterozygosity for the direct let-7 target
abrupt
, encoding a
transcription factor that regulates NMJ development (
Caygill and Johnston,
2008
). Additionally, the enhanced neurotransmitter release seen in the NMJ
of deletion mutants of the
mir-310/311/312/313
cluster, all of which
encoded seed-related miRNAs, was rescued by heterozygosity for
khc-73
,
a neural-specific kinesin family member (
Tsurudome
et al
., 2010
).
These findings suggest that it may be fairly common for the derepression
of specific miRNA target genes to be of disproportionate phenotypic
impact. We certainly do not wish to imply that these particular targets are
the “only” targets of these miRNAs, and phenotypic rescue by target
reduction is still compatible with the notion that miRNAs might com-
monly have hundreds of target genes. It merely highlights that these are not
necessarily of equal functional regulatory consequence. This is actually
reminiscent of the situation of transcription factor target genes. With
modern molecular profiling methods, it is clear that typical transcription
