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miRNA Regulatory Ecosystem in Early Development
Yuka W. Iwasaki1 and Haruhiko Siomi1,*1Department of Molecular Biology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku, Tokyo 160-8582, Japan*Correspondence: awa403@z2.keio.jp
http://dx.doi.org/10.1016/j.molcel.2014.11.010

MicroRNAs (miRNAs) reshape spatiotemporal gene expression by both modulating the levels of activelytranscribed genes and accelerating the clearance of previously transcribed messages, thereby promoting
the transition from a preceding stage to subsequent processes during development. Lee et al. (2014) now
demonstrate that maternal miRNAs are adenylated by Wispy, which leads to clearing of maternal miRNAs
during early embryogenesis.
I knew the moment had arrived
For killing the past and comingback to life
—David Gilmour (The Division Bell)
Maternally deposited mRNAs are involved in early animal development.
Maternal mRNAs are translationally activated by egg activation following fertilization, which correlates with an increase in
the length of their poly(A) tails. Subsequently, maternal mRNAs are destroyed,
and genetic control is switched fromthe maternal genome to the zygotic
genome, in an early embryogenesis
process called the maternal-to-zygotic
transition (MZT) (Giraldez 2010). Two
degradation pathways function to eliminate maternal mRNA. First, maternal factors initiate the destruction of a subset ofmaternal mRNAs, which enables zygotictranscription. Second, de novo zygotic
transcription of microRNAs (miRNAs)
clears maternal mRNAs by promoting
their deadenylation. For example, inDrosophila, the maternally provided
RNA-binding protein Smaug directs thedegradation of a subset of maternal transcripts following egg activation. Smaug
does this by recruiting the CCR4/POP2/
NOT deadenylase complex to its target
transcripts, which induces poly(A) tailshortening and transcript destruction(Semotok et al., 2005; Benoit et al.,2009). This Smaug-mediated clearanceof maternal mRNAs activates the secondand more rapid phase of maternal mRNA
clearance by enabling zygotic expressionof miRNAs (Bushati et al., 2008). Zygotic
transcription may be activated following
the destruction of maternal mRNA by thesubsequent downregulation of maternal

proteins that repress the zygote genome(Figure 1).
miRNAs are typically produced fromlong primary transcripts by two sequential
processing reactions. The first processincludes the enzyme Drosha, which isfound in the nucleus, producing pre-miRNAs. The second process consists ofthe enzyme Dicer, which is found in thecytoplasm, generating mature miRNAs.
Following Dicer processing, miRNAs areloaded onto Argonaute proteins to form
miRISCs, which control mRNA expression
by promoting degradation or repressing
translation. Thus, miRNAs play an important role in establishing specific cellularstates by shaping specific transcriptomes
and proteomes. A large fraction of miRNAs
exhibit developmental, stage-specific,
and tissue-specific expression patterns,
which often involves posttranscriptional
regulation of miRNA production and turnover (Ha and Kim, 2014). One pathwaythat controls the abundance and/or activity of some miRNAs involves the nontemplated addition of nucleotides (i.e., notfound in genomic sequences) to their 30ends. This is performed by members of
the Pol b superfamily, including poly(A)polymerases (PAPs), noncanonical PAPs,
and terminal uridylyl transferases (TUTs)
with distinct nucleotidyl transferase activities and biological functions. The consequences of nucleotide addition to the 30
ends of miRNAs varies and depends on
substrates, numbers (mono or oligo) andmodifications (adenylation or uridylation)of added nucleotides. For example, oligouridylation of pre-let-7 by TUT4 induces
its degradation in embryonic stem cells,
whereas monouridylation by the same
enzyme promotes processing by Dicer insomatic cells.

The nontemplated 30 nucleotide additions occur only on specific miRNAs andare cell-type and developmental-stage
specific. It has been shown that in early
fly embryos, a large number of miRNAs
exhibit nontemplate 30 end additions ofadenosines (Fernandez-Valverde et al.,2010). Deep sequencing performed by
Lee and colleagues (2014) also showedthat over 30% of miRNAs from
Drosophila-activated eggs exhibited nontemplate 30 end adenosine additions.
However, adenylation was found to be
reduced in early embryos, which suggests the adenylation is restricted tomaternal miRNAs. This dynamic adenylation pattern during early embryogenesis
was also evident in sea urchin and mice.
To identify the enzyme responsible formaternal miRNA adenylation, Lee et al.
(2014) examined expression patterns ofseveral PAPs in Drosophila and foundthat Wispy (Wisp), a noncanonical PAP,
is highly expressed in egg cells and early
embryos and decreases following zygotic
genome activation. Wisp mutant eggs
have decreased adenylation of miRNAs
and subsequently increased miRNAlevels. Upregulating Wisp in cultured S2cells results in increased adenylation
with a concomitant reduction of mature
miRNAs, without modification or accumulation of pre-miRNAs. Consistent with
this, close examination of individual
sequencing reads show that both the 5pand 3p strands of miRINAs are adenylated. This indicates that Wisp acts downstream of Dicer processing, which exposes the 30 end of the 5p strand.
Previous studies showed that Wisp isrequired for poly(A) tail elongation ofseveral maternal mRNAs upon egg activation, which initiates the translation of

Molecular Cell 56, December 4, 2014 ª2014 Elsevier Inc. 615

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enylation leads to opposite effects indifferent cell types, how Wisp is regulated
to add long poly(A) tails on mRNAs andmono or oligo adenines on miRNAs, andhow specific miRNAs are selected for adenylation given that Wisp adenylates miRNAs through direct association withAGO1.
REFERENCES

Figure 1. Function of Wispy during Maternal-to-Zygotic Transition(A) General diagram of maternal and zygotic mRNA and miRNA levels during the MZT. Wispy plays a role inmaternal clearance of transcripts and activation of zygotic translation.
(B) miRNA and mRNA levels are regulated by Wispy-mediated adenylation. Wispy associates with AGO1,
leading to adenylation of miRNAs. Adenylation triggers degradation of miRNAs by an unknown nuclease,resulting in depression of miRNA target genes. AGO1-miRNA complexes (miRISCs), which are unrecognized by Wispy, are capable of silencing their target genes. Wispy, together with RNA-binding proteins(RBP), polyadenylates some population of maternal mRNAs and stabilizes them. However, some adenylated mRNAs are recognized by a translational repressor, Smaug, which results in deadenylation anddestabilization of the mRNA. Consequently, regulation of maternal mRNAs leads to activation of zygoticgenes.

these mRNAs (Cui et al., 2008). Because
Wisp lacks an RNA-recognition motif, itis recruited to target maternal mRNAs
through direct interaction with RNA-binding proteins, including Bicaudal-C. Leeet al. (2014) now demonstrate that Wisp
physically interacts with AGO1, on which
miRNAs are loaded, to form miRISCs after
Dicer processing (Okamura et al., 2004).
Wisp directly adenylates AGO1-associated miRNAs. These findings suggestthat Wisp has two functions in the activation of zygotic transcription: it triggers theclearance of specific maternal miRNA
subpopulations by adenylating them during MZT and stabilizes some maternalmRNA by poly(A) elongation. Together

with maternal mRNA clearance, thesefunctions activate zygotic genome transcription (Figure 1).
miRNA adenylation by GLD2, a possible Wisp ortholog, is required to stabilize
miRNAs in somatic cells (Katoh et al.,2009; D’Ambrogio et al., 2012). This function is the exact opposite to the proposedrole of Wisp. In addition to causing miRNA
destruction, Wisp is also required formaternal mRNA poly(A) tail elongation
upon egg activation. Therefore, if there isa limited amount of Wisp, competition between miRNA destruction and mRNApoly(A) elongation must occur. The study
by Lee et al. (2014) raises several important questions, including how miRNA ad-616 Molecular Cell 56, December 4, 2014 ª2014 Elsevier Inc.

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