PERSPECTIVES
while resisting brittle fracture. As advancesin the understanding of brittle fracture andthe control of microstructure feed into thedesign of new structural steels, more exciting
results of this kind can be expected.
References and Notes
1. Y. Kimura, T. Inoue, F. Yin, K. Tsuzaki, Science 320, 1057
(2008).
2. A. J. McEvily, Metal Failures (Wiley-Interscience, NewYork, 2001), chap. 1.
3. G. Thomas, Metall. Trans. A 2, 2373 (1971).
4. J. W. Morris Jr., C. S. Lee, Z. Guo, ISIJ Int. 43, 410 (2003).
5. The effective yield stress in the Yoffee diagram is thestress required to plastically deform the severely constrained material at the tip of a sharp crack. Unless theloaded piece is very thin, geometric constraints at thecrack tip have the consequence that plastic deformation is difficult there, and requires a stress σY that isthree to five times the conventional yield stress mea-

sured in standard tensile tests using unconstrained
samples and tabulated in tables of engineering properties (4, 11).
6. S. Takaki, T. Maki, Eds., Ultrafine Grained Steels (Iron andSteel Institute of Japan, Tokyo, 2001).
7. Z. Guo, C. S. Lee, J. W. Morris Jr., Acta Mater. 54, 5511
(2004).
8. T. Maki, in Fundamentals of Martensite and Bainite:
Toward Future Steels with High Performance, T. Furuhara,
K. Tsuzaki, Eds. (Iron and Steel Institute of Japan, Tokyo,2007), pp. 1–10.
9. The Development of High Performance Steels for the 21stCentury (Pohang Iron and Steel Company Ltd., Pohang,Korea, 2002).
10. K. T. Venkateswara Rao, W. Yu, R. O. Ritchie, Metall.
Trans. A 20A, 485 (1989).
11. R. M McMeeking, G. M. Parks, in Elastic-Plastic Fracture(American Society for Testing and Materials, Philadelphia,1979), pp. 175–194.
12. Supported by NSF grant DMR 0304629.
10.1126/science.1158994

MOLECULAR BIOLOGY

Slicing and Dicing for Small RNAsJames A. Birchler and Harsh H. Kavi
A new type of small RNA and mode of gene regulation is discovered in fly and mammals.

early two decades after gene silencing
was first described (1), the field continues to reveal new and diverse roles
for the small RNA moieties involved. These
functions include modulating the translation ofmessenger RNA (mRNA) into protein, establishing chromosomal architecture, regulatingstem cell renewal, and providing defense
against viruses and mobile genetic elements(transposons) that could cause deleterious
mutations (2). Three major classes of small
RNAs have been defined in plants and animals:
microRNAs (miRNAs), small interfering
RNAs (siRNAs), and Piwi-interacting RNAs
(piRNAs). Previous work has suggested that
dedicated pathways generate each class ofsmall RNA. Now, six recent studies, including
one by Ghildiyal et al. on page 1077 of this issue,
reveal an interplay of these canonical pathways
in generating a new class of endogenous
siRNAs (endo-siRNA) (3–8) (see the figure).
miRNAs are encoded by the genome andprocessed to 22–base pair products that interact with homologous mRNAs to modulate
their translation and affect developmental
processes. siRNAs are 21 base pairs long andgenerated from double-stranded RNA precursors such as those from viruses and endogenous transposons. Recently, piRNAs haveN

Division of Biological Sciences, University of Missouri,
Columbia, MO 65211, USA. E-mail: birchlerj@missouri.
eduwww.sciencemag.org

SCIENCEVOL 320

Published by AAAS

been described in the germ line of flies andmammals (9). They are approximately 25 basepairs long with homology to transposable
elements and other sequences.
In the fly Drosophila melanogaster, theenzyme Dicer-1 cleaves double-stranded
miRNA precursors, and the resulting products bind to the protein Argonaute1 (Ago1).
siRNAs are generated by Dicer-2 from double-stranded RNA precursors, and bind toArgonaute2 (Ago2), whose “slicer” activity
cleaves homologous RNAs. piRNAs are generated by a dicer-independent mechanismthat relies on the slicer function of a separate
clade of argonaute proteins consisting ofPiwi, Aubergine, and Argonaute3 (9). These
proteins use antisense transcripts (complementary to mRNA) encoded by piRNA clusters in the genome that harbor transposable
element fragments, and target the destruction of transposon sense transcripts. New
studies report sequencing of extensive poolsof small RNAs from various somatic andgermline sources in flies and mice (3, 4, 6–8)
and identify a new type of siRNA that ishomologous to transposons and proteinencoding gene sequences.
Ghildiyal et al. found that although mostsmall RNAs in fly somatic cells aremiRNAs, some have characteristics ofsiRNAs—equal quantities of sense and antisense orientations, a 21–base pair length, and2'-O-methyl modifications at the 3' end. Most

23 MAY 2008

Downloaded from www.sciencemag.org on May 22, 2008

other crystallographic discontinuity forces it
to deviate or stop. Processes to do so include
thermomechanical treatments during hotrolling from slab to plate, sheet, or bar (6),thermal cycling treatments that manipulate
the transformations that occur during quenching (7), and tempering treatments that interpose fine distributions of nanosized secondphase particles to disrupt the intragranular
crystallography (7).
The complexity of the many possible
microstructures of steel often makes it difficult to decipher what the effective grain size
really is. Recently, electron backscatter diffraction (EBSD) has been used to map crystallographic patterns in the microstructure ofsteels and to identify the effective grain sizes
in several important microstructures (4, 8).
This information is being systematically
exploited in steel research, particularly in various “supersteel” projects in Asia (9).
The second generic approach to lowering
TB is to decrease the effective yield stress at
the crack tip. A simple way to do this is toreplace thick beams with a laminate of thinsheets, a technique used by 19th-century engineers in early steels. A modern alternative is todesign the microstructure so that it spontaneously delaminates in the stress field at thecrack tip, effectively dividing itself into a laminate of thin sheets and lowering the effective
stress at the crack tip by a factor of 2 or more
(10). This approach was used (but, to myknowledge, never published) by manufacturers of steel line pipe more than 20 years ago. Itlowers TB but also lowers the toughness in theductile mode, presumably because the weakerof the delaminated sheets fracture under smallloads. Particularly in the case of ultrahighstrength steels, high toughness is required forstructural reliability in service.
Kimura et al. now describe a clever method
for combining grain refinement and delamination to obtain a particularly promising combination of properties. The steel is processedto high strength through a combination ofgrain refinement and precipitation hardening.
The grain refinement is accomplished bybar-rolling, producing a fibrous texture thatdelaminates ahead of and perpendicular to the
propagating crack tip, thereby relaxing thecrack-tip stress concentration without producing planes of weakness to lower toughness inthe ductile mode.
Defeating the ductile-brittle transition insteel is always difficult, and as the figure
makes clear, this becomes particularly challenging for an ultrahigh-strength steel. As
Kimura et al. show, this challenge can be
met with low-alloy steel by tailoring themicrostructure to achieve high strength1023

PERSPECTIVES
line–accentuated mechanism to ameliorate
the mobility of transposable elements, andthus reduce the occurrence of mutations.
However, the finding that endo-siRNAs arepresent with homology to complementary
pseudogenes and homologous genes as a
newly recognized mode of gene regulation(8), and that long hpRNA genes produce
siRNAs that can affect gene expression (5),might suggest that these roles for siRNAs
have maintained the Dicer-dependent machinery at some level in both somatic andgermline cells of animals. ThisSMALL RNAsmachinery could still metabolize
Germ line
double-stranded viral and transLengthposon RNAs.
TypeMechanism of generation
Biological role(nucleotides)
Dual small RNA–processing
miRNA
21–23
Dicer dependent
Regulation of translation
mechanisms in animals might
piRNA
24–27
Dicer independent, Piwi clade dependent
Transposon regulation and unknown functions
reflect a response to transposons
endo-siRNA 21–23
Dicer dependent
Transposon regulationthat evolve to evade silencing by
one or the other mechanism in theSoma
never-ending arms race between
miRNA
21–23Dicer-1 dependent (in Drosophila)
Regulation of translationtransposable elements and thepiRNA like
24–27
In Ago2 mutants (in Drosophila)
Transposon regulation
host genome. Transposons in mouseendo-siRNA 21–23
Dicer-2/Ago2 dependent (in Drosophila)
Regulation of transposons, mRNAs, and heterochromatin
oocytes have somewhat distinct
hpsiRNA
21–23
Dicer-2/Ago2/Loqs dependent (in Drosophila)
Regulation of mRNAs
profiles of siRNAs or piRNAs,
Small RNAs. A summary of small RNAs found in germline and somatic cells and their identified functions.
indicating that certain elements
can avoid one of the mechanisms
ily of argonautes. Mutations in dcr-2 or ago2 ing that of transposons and certain mRNAs. of silencing. Any transposon that escapes bothincreased the expression of certain trans- Because there is only one Dicer gene in mam- systems will likely proliferate into newposons, thus linking endo-siRNAs to the con- mals, these results demonstrate that this genomic positions and produce substrates thattrol of transposable elements in the soma.
enzyme functions in both miRNA and siRNA could be subject to one or the other type ofSimilarly, Czech et al. (4) found endo- production. The piRNAs found in oocytes silencing. The piRNA loci are graveyards ofsiRNAs in somatic and germline cells of flies were bound to MILI, a mammalian counter- transposable elements whose ghosts in theby analyzing sequences from three sources: part of Piwi. In this case, piRNAs and siRNAs form of small RNAs commit fratricide against
those bound to Ago1 and Ago2, those present in sometimes correspond to distinct transposons, their homologous brethren. From a selfish
small RNA libraries, and those from flies with indicating preferred mechanisms for their gen- DNA point of view, it is interesting to conmutations in the canonical miRNA and siRNA eration, although transposon-rich loci could template how the transposons are “tricked” bypathways. They observed that the protein also give rise to both siRNAs and piRNAs.
the host to insert into piRNA loci and in turnLoquacious (Loqs), which associates with
It is not unexpected that endo-siRNAs to whether classes of transposons evolve mechaDicer-1 for miRNA production, also interacts transposable elements would be found in nisms to avoid these locations for insertion.
with Dicer-2 and is required for some endo- animals, given their presence in plants (10)References
siRNA formation. Okamura et al. (5) analyzed and fungi (11). Based on the sophisticated
1. M. A. Matzke et al., EMBO J. 8, 643 (1989).
the processing of products of endogenous long immune systems of vertebrates, early thinking
2. P. J. Paddison, P. K. Vogt, Eds., RNA Interference, Current
RNA hairpin (hp) genes in flies. These posited that the small RNA–processing
Topics in Microbiology and Immunology (Springer,Berlin, 2008), vol. 320.
hpRNAs are processed by Dicer-2, in conjunc- machinery involved Dicer primarily for the
3. M. Ghildiyal et al., Science 320, 1077 (2008); publishedtion with Loqs, and then bind to Ago2. The maturation of miRNAs. The discovery of aonline 10 April 2008 (10.1126/science.1157396).
siRNAs generated from these genes can silence piRNA system that does not involve Dicer,
4. B. Czech et al. Nature 10.1038/nature07007 (2008).
5. K. Okamura et al., Nature 10.1038/nature07015
selected mRNAs using the slicer function of but protects the germ line from transposon(2008).
Ago2 rather than by translational repression. expression, reinforced this view. Although it is
6. Y. Kawamura et al., Nature 10.1038/nature06938 (2008).
Collectively, these new fly studies illustrate the difficult to compare the strength of gene
7. T. Watanabe et al., Nature 10.1038/nature06908 (2008).
interplay among mechanisms generating vari- silencing across species, Drosophila somatic
8. O. H. Tam et al., Nature 10.1038/nature06904 (2008).
9. A. A. Aravin, G. J. Hannon, J. Brennecke, Science 318,ous classes of small RNAs.
cells require multiple copies of a transgene to761 (2007).
In a complementary study, Kawamura reduce the expression of homologous tran- 10. A. Hamilton, O. Voinnet, L. Chappell, D. Baulcombe,
et al. (6) sequenced small RNAs bound by scripts (12), and the effect is not as great as
EMBO J. 21, 4671 (2002).
Ago2 in cultured Drosophila cells. The endo- seen with a few copies of many transgenes in 11. A. Chicas, C. Cogoni, G. Macino, Nucleic Acids Res. 32,4237 (2004).
siRNAs recovered showed homology to a sub- plants (2). Perhaps the evolution of robust 12. M.
Pal Bahdra, U. Bhadra, J. A. Birchler, Mol. Cell 9, 315
set of transposons. Depletion of Dicer-2 and immune systems in animals relaxed the(2002).
Ago2 in these cells reduced the quantities of selection pressure for a strong response of
Published online 8 May 2008;
endo-siRNAs and increased the expression of the siRNA defense against viruses. Thus, 10.1126/science.1159018
the corresponding transposons. Endo-siRNAs the piRNA system might represent a germ Include this information when citing this paper.

1024

with homology to transposons were also
found in fly ovaries, and their function depended on Dicer-2 and Ago2, revealing thatthe germ line has both siRNA- and piRNAgenerating machineries.
The production of small RNAs by slicing
and dicing has also been reported in mouse
germline cells. Watanabe et al. (7) and Tam etal. (8) sequenced small RNAs from mouse
oocytes and found both siRNAs and piRNAs.
Depletion of Dicer and Ago2 in these cells
reduced the number of siRNAs while increas-

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Downloaded from www.sciencemag.org on May 22, 2008of these endo-siRNAs are homologous totransposable elements, and many originate
from piRNA clusters. Curiously, mutations ofthe genes encoding Dicer-2 (dcr-2) and Ago2(ago2) did not eliminate all of these small
endo-siRNAs, suggesting that Dicer-1 andAgo1 can generate and bind, respectively,
some siRNAs under these circumstances.
Moreover, somatic pi-like RNAs, which
exhibit characteristics of the germline namesake, were found in ago2 mutants, raising thepossibility of somatic activity of the Piwi fam-