Cell, Vol. 116, 281–297, January 23, 2004, Copyright 2004 by Cell Press
ReviewMicroRNAs: Genomics,
Biogenesis, Mechanism, and Function
ulation of hematopoietic lineage differentiation in mam-
mals (Chen et al., 2004), and control of leaf and flower
development in plants (Aukerman and Sakai, 2003;
David P. Bartel
1,2,
*
1
Whitehead Institute for Biomedical Research
9 Cambridge Center
Cambridge, Massachusetts 02142 Chen, 2003; Emery et al., 2003; Palatnik et al., 2003).
Computational approaches for finding messages con-
2
Department of Biology
Massachusetts Institute of Technology trolled by miRNAs indicate that these examples repre-
sent a very small fraction of the total (Rhoades et al.,Cambridge, Massachusetts 02139
2002; Enright et al., 2003; Lewis et al., 2003; Stark et
al., 2003).
This review highlights what has been learned aboutMicroRNAs (miRNAs) are endogenous
22 nt RNAs
that can play important regulatory roles in animals and miRNAs in the decade since the report of the lin-4 RNA
and its regulation of lin-14. The major topics discussedplants by targeting mRNAs for cleavage or transla-
tional repression. Although they escaped notice until are miRNA genomics, miRNA biogenesis, miRNA regula-
tory mechanisms, and the roles of miRNAs in gene regu-relatively recently, miRNAs comprise one of the more
abundant classes of gene regulatory molecules in latory pathways.
multicellular organisms and likely influence the output
of many protein-coding genes.
Genomics: The miRNA Genes
For seven years after the discovery of the lin-4 RNA, the
genomics of this type of tiny regulatory RNA appeared
In an investigation inspiring for both its perseverance
simple: there was no evidence for lin-4-like RNAs be-
and its scientific insight, Victor Ambros and colleagues,
yond nematodes and no sign of any similar noncoding
Rosalind Lee and Rhonda Feinbaum, discovered that
RNAs within nematodes. This all changed upon the dis-
lin-4, a gene known to control the timing of C. elegans
covery that let-7, another gene in the C. elegans hetero-
larval development, does not code for a protein but
chronic pathway, encoded a second
22 nt regulatory
instead produces a pair of small RNAs (Lee et al., 1993).
RNA. The let-7 RNA acts to promote the transition from
One RNA is approximately 22 nt in length, and the other
late-larval to adult cell fates in the same way that the
is approximately 61 nt; the longer one was predicted to
lin-4 RNA acts earlier in development to promote the
fold into a stem loop proposed to be the precursor of
progression from the first larval stage to the second
the shorter one. The Ambros and Ruvkun labs then no-
(Reinhart et al., 2000; Slack et al., 2000). Furthermore,
ticed that these lin-4 RNAs had antisense complemen-
homologs of the let-7 gene were soon identified in the
tarity to multiple sites in the 3
UTR of the lin-14 gene
human and fly genomes, and let-7 RNA itself was de-
(Lee et al., 1993; Wightman et al., 1993). This comple-
tected in human, Drosophila, and eleven other bilateral
mentarity fell in a region of the 3
UTR previously pro-
animals (Pasquinelli et al., 2000).
posed to mediate the repression of lin-14 by the lin-4
Because of their common roles in controlling the tim-
gene product (Wightman et al., 1991). The Ruvkun lab
ing of developmental transitions, the lin-4 and let-7
went on to demonstrate the importance of these com-
RNAs were dubbed small temporal RNAs (stRNAs), with
plementary sites for regulation of lin-14by lin-4, showing
anticipation that additional regulatory RNAs of this type
also that this regulation substantially reduces the
would be discovered (Pasquinelli et al., 2000). Indeed,
amount of LIN-14 protein without noticeable change
less than one year later, three labs cloning small RNAs
in levels of lin-14 mRNA. Together, these discoveries
from flies, worms, and human cells reported a total of
supported a model in which the lin-4 RNAs pair to the
over one hundred additional genes for tiny noncoding
lin-14 3
UTR to specify translational repression of the
RNAs, approximately 20 new genes in Drosophila, ap-
lin-14 message as part of the regulatory pathway that
proximately 30 in human, and approximately 60 in
triggers the transition from cell divisions of the first larval
worms (Lagos-Quintana et al., 2001; Lau et al., 2001;
stage to those of the second (Lee et al., 1993; Wightman
Lee and Ambros, 2001). The RNA products of these
et al., 1993).
genes resembled the lin-4 and let-7 stRNAs in that they
The shorter lin-4RNA is now recognized as the found-
were
22 nt endogenously expressed RNAs, potentially
ing member of an abundant class of tiny regulatory RNAs
processed from one arm of a stem loop precursor (Figure
called microRNAs or miRNAs (Lagos-Quintana et al.,
1), and they were generally conserved in evolution—
2001; Lau et al., 2001; Lee and Ambros, 2001). The
some quite broadly, others only in more closely related
breadth and importance of miRNA-directed gene regula-
species such as C. elegans and C. briggsae. But unlike
tion are coming into focus as more miRNAs and their
lin-4 and let-7 RNAs, many of the newly identified
22
regulatory targets and functions are discovered. Re-
nt RNAs were not expressed in distinct stages of devel-
cently discovered miRNA functions include control of
opment and instead were more likely to be expressed
cell proliferation, cell death, and fat metabolism in flies
in particular cell types. Thus the term microRNA was
(Brennecke et al., 2003; Xu et al., 2003), neuronal pat-
used to refer to the stRNAs and all the other tiny RNAs
terning in nematodes (Johnston and Hobert, 2003), mod-
with similar features but unknown functions (Lagos-
Quintana et al., 2001; Lau et al., 2001; Lee and Ambros,
2001). Intensified cloning efforts have revealed numer-*Correspondence: dbartel@wi.mit.edu

Cell
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Figure 1. Examples of Metazoan miRNAs
Shown are predicted stem loops involving the
mature miRNAs (red) and flanking sequence.
The miRNAs* (blue) are also shown in cases
where they have been experimentally identi-
fied (Lim et al., 2003a).
(A) Predicted stem loops of the founding
miRNAs, lin-4 and let-7RNAs (Lee et al., 1993;
Reinhart et al., 2000). The precise sequences
of the mature miRNAs were defined by clon-
ing (Lau et al., 2001). Shown are the C. ele-
gans stem loops, but close homologs of both
have been found in flies and mammals (Pas-
quinelli et al., 2000; Lagos-Quintana et al.,
2001, 2002).
(B) Examples of miRNAs from other metazoan
genes, mir-1, mir-34, and mir-124. Shown are
the C. elegans stem loops, but close homo-
logs of these miRNAs have been found in flies
and mammals (Lagos-Quintana et al., 2001,
2002; Lau et al., 2001; Lee and Ambros, 2001).
(C) Examples of miRNAs from plant genes,
MIR165a, MIR172a2, and JAW. Shown are
Arabidopsis stem loops, but close homologs
of these miRNAs have been found in rice and
other plants (Park et al., 2002; Reinhart et al.,
2002; Palatnik et al., 2003).
ous additional miRNA genes in mammals, fish, worms, latory scenarios are easy to imagine in which such coor-
dinate expression could be useful, which would explainand flies (Lagos-Quintana et al., 2002, 2003; Mourelatos
et al., 2002; Ambros et al., 2003b; Aravin et al., 2003; the conserved relationships between miRNAs and host
mRNAs. A striking example of this conservation involvesDostie et al., 2003; Houbaviy et al., 2003; Kim et al.,
2003; Lim et al., 2003a, 2003b; Michael et al., 2003). A mir-7, found in the intron of hnRNP K in both insects
and mammals (Aravin et al., 2003).registry has been set up to catalog the miRNAs and
facilitate the naming of newly identified genes (Griffiths- Other miRNA genes are clustered in the genome with
an arrangement and expression pattern implying tran-Jones, 2004).
Like C. elegans lin-4 and let-7, most miRNA genes scription as a multi-cistronic primary transcript (Lagos-
Quintana et al., 2001; Lau et al., 2001). Although thecome from regions of the genome quite distant from
previously annotated genes, implying that they derive majority of worm and human miRNA genes are isolated
and not clustered (Lim et al., 2003a, 2003b), over halffrom independent transcription units (Lagos-Quintana
et al., 2001; Lau et al., 2001; Lee and Ambros, 2001). of the known Drosophila miRNAs are clustered (Aravin
et al., 2003). The miRNAs within a genomic cluster areNonetheless, a sizable minority (e.g., about a quarter of
the human miRNA genes) are in the introns of pre- often, though not always, related to each other; and
related miRNAs are sometimes but not always clusteredmRNAs. These are preferentially in the same orientation
as the predicted mRNAs, suggesting that most of these (Lagos-Quintana et al., 2001; Lau et al., 2001). Orthologs
of C. elegans lin-4 and let-7 are clustered in the fly andmiRNAs are not transcribed from their own promoters
but are instead processed from the introns, as seen also human genomes and are coexpressed, sometimes from
the same primary transcript, leading to the idea that thefor many snoRNAs (Aravin et al., 2003; Lagos-Quintana
et al., 2003; Lai et al., 2003; Lim et al., 2003a). This genomic separation of lin-4 from let-7 in nematodes
might be unique to the worm lineage (Aravin et al., 2003;arrangement provides a convenient mechanism for the
coordinated expression of a miRNA and a protein. Regu- Bashirullah et al., 2003; Sempere et al., 2003). This exam-