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We found that DCL1 and, by implication, miRNAs were required for multiple embryonic patterning events beginning as early as the eight-cell stage.
In fact, the only cell type that appeared to differentiate correctly in dcl1 embryos was the outermost protoderm layer. Furthermore, protoderm markers that are typically restricted to the embryo proper
were ectopically expressed in dcl1 suspensors at the late globular and early heart stages (Fig. 1B; Supplemental Fig. 2), which helps explain a previously reported function of DCL1/SUS1 in maintaining the extraembryonic cell fate of the suspensor (Schwartz et al. 1994).
The requirement of miRNAs for proper embryonic patterning and cell differentiation might have suggested that embryonic miRNAs
act to trigger developmental transitions, as was first reported for metazoan miRNAs (Lee et al. 1993; Wightman et al. 1993; Moss et al. 1997; Reinhart et al. 2000), where induced miRNAs act to down-regulate genes that contribute to the differentiation state of the precursor cells. Or
they might act to sharpen developmental transitions, as observed in fish, frog, and, perhaps, mammalian embryos, where miR-430
or its orthologs clears maternally expressed messages to sharpen the transition from maternal to zygotic expression programs
(Farh et al. 2005; Giraldez et al. 2006; Lund et al. 2009). Indeed, an analogous function in sharpening developmental transitions was proposed and has been observed for plant miRNAs
at later developmental stages (Rhoades et al. 2002; Aukerman and Sakai 2003; Lauter et al. 2005; Wu et al. 2009). However, our molecular characterization of the Dcl1 phenotype shows that the earliest known developmental roles of miRNAs
in plants is not to trigger or sharpen developmental transitions by repressing genes with developmental functions in precursor
cell types, but instead is the opposite (Fig. 7). In eight-cell embryos, miRNAs repress genes that function in daughter cell types later in embryo development. Instead of
attenuating pre-existing function, they prevent precocious function. Dicer and other miRNA biogenesis proteins are required
for formation of the mammalian mesoderm and differentiation of murine embryonic stem cells (Bernstein et al. 2003; Kanellopoulou et al. 2005; O'Rourke et al. 2007; Wang et al. 2007). Perhaps as the roles of individual vertebrate miRNAs are examined in more detail, analogous functions in preventing precocious
embryonic expression will help explain the requirement of miRNAs for mammalian embryonic development. Indeed, recent studies
show that the extraembryonic lineage of mouse embryos uses miRNAs to prevent premature differentiation and maintain extraembryonic
stem cell multipotency (Spruce et al. 2010).
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