The Role of MicroRNAs in Muscle Development - Current Topics in Developmental Biology - page 65

targeting the general RNA-binding proteins required for miRNA produc-
tion including Dicer and Drosha. In this section, I focus first on muscle
phenotypes displayed by dicer mutants and then discuss the phenotypic
analysis of individual miRNA mutants.
3.1. Muscle phenotypes of zebrafish and mouse
dicer mutants
Although dicer mutants provide a powerful approach to analyzing the global
role of miRNAs, some caveats exist. First, Dicer is required for the proces-
sing of not only miRNAs but other biologically active small RNA species
like endogenous siRNAs as well (for a recent review, see Suh and Blelloch,
2011 ). Second, Dicer depletion does not result in the complete elimination
of all miRNAs, since some stable miRNAs can still be detected well after
dicer depletion ( Giraldez et al ., 2005 ) and other miRNAs are processed by a
Dicer-independent mechanism ( Cheloufi et al ., 2010 ; Cifuentes et al ., 2010 ;
Giraldez et al ., 2005 ). Nevertheless, the expression of many miRNAs is
affected by Dicer depletion, which therefore provides a good starting point
for analyzing the general roles of miRNAs in developmental processes.
Below, I summarize the phenotypes displayed by zebrafish and mouse
dicer mutants. Depleting Dicer to identify the role of miRNAs during
nematode and fruit fly muscle development has not been possible, since
dicer mutant worms arrest at an early developmental stage that is difficult
to study ( Grishok et al ., 2001 ), and maternal fly dicer is required for germline
development, precluding its analysis in the early embryo ( Hatfield et al .,
2005 ).
The role of miRNAs during embryogenesis was explored in zebrafish,
through the analysis of embryos lacking both maternal and zygotic Dicer
( Giraldez et al ., 2005 ). These MZ dicer mutants undergo axis formation and
differentiate multiple cell types including muscles, indicating that miRNAs
are not required for the initial patterning and specification of the germ layers
including the mesoderm and at least some of its derivatives. MZ dicer mutants
display defects in subsequent embryonic events, however, including mor-
phogenesis of the somites and heart. Although somites form normally in
MZ dicer mutants, they do not acquire their normal chevron shape. Simi-
larly, a two-chamber heart does not form, even though contractile cardio-
myocytes are specified correctly in MZ dicer mutants. Instead, MZ dicer
mutants display a tubular heart and pericardial edema. These results indicate
that miRNAs are not absolutely required for myogenic cell fate specifica-
tion, but may play roles in modulating cell type-specific differentiation as
well as influencing the behavior of myogenic cells once differentiated.
To extend this phenotypic analysis to the molecular level, global gene
expression was profiled in muscle cells isolated from 24-h-old MZ dicer
mutants ( Mishima et al ., 2009 ). Hundreds of inappropriately expressed