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14.

15.

pattern of B lymphocyte induced
maturation protein-1 (Blimp-1) during
mouse embryonic development. Mech.
Dev. 117, 305–309.
de Souza, F.S., Gawantka, V., Gomez,
A.P., Delius, H., Ang, S.L., and Niehrs, C.
(1999). The zinc finger gene Xblimp1
controls anterior endomesodermal cellfate in Spemann’s organizer. EMBO J.
18, 6062–6072.
Baxendale, S., Davison, C., Muxworthy,
C., Wolff, C., Ingham, P.W., and Roy, S.
(2004). The B-cell maturation factor
Blimp-1 specifies vertebrate slow-twitch

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muscle fiber identity in response to
Hedgehog signaling. Nat. Genet. 36,
88–93.
Anderson, R., Copeland, T.K., Scholer,
H., Heasman, J., and Wylie, C. (2000).
The onset of germ cell migration in themouse embryo. Mech. Dev. 91, 61–68.
Starz-Gaiano, M., and Lehmann, R.
(2001). Moving towards the next
generation. Mech. Dev. 105, 5–18.
Raz, E. (2004). Guidance of primordial
germ cell migration. Curr. Opin. Cell Biol.
16, 169–173.
Balkwill, F. (2004). Cancer and theMicroRNAs: Loquacious Speaksout
In Drosophila, Dicer-2 requires the double-stranded RNA binding
protein R2D2, to mediate the assembly of short interfering RNAs into
the RNA-induced silencing complex. New data show that Dicer-1 also
requires a double-stranded RNA binding protein called Loquacious for
efficient microRNA-mediated gene silencing.
Philipp J. F. Leuschner,
Gregor Obernosterer andJavier Martinez
A major breakthrough in molecularbiology was the finding thateukaryotic cells harness
mechanisms of RNA interference(RNAi) to regulate the expressionof endogenous genes [1,2]. Just
as in the ‘canonical’ RNAi
pathway, short single-stranded
RNA molecules called microRNAs(miRNAs) serve as sequencespecific guides to target silencing
complexes containing an
Argonaute (Ago) protein tocognate sequences — typically inthe 3′′-untranslated region oftarget messenger RNAs [3].
miRNAs originate from longprimary transcripts (pri-miRNAs)[4], which are processed in the
nucleus by the RNase III-likeenzyme Drosha and its cofactor
Pasha/DCGR8 into ~65 nt, hairpin
shaped precursors, or pre-miRNAs[5–8]. Pre-miRNAs are exported tothe cytoplasm, where another
RNase III-like enzyme, Dicer,
liberates a ~22 nt long miRNA
duplex from the hairpin. One
strand of the duplex is integrated
into an active RNA-induced
silencing complex (RISC) [9].
While human Dicer is able toprocess long double-stranded
RNA (dsRNA) as well as premiRNAs, the two pathways are
separated in Drosophila (Figure1). Here, Dicer-2 does not play arole in miRNA biogenesis, but isrequired to cleave long dsRNAs
into short interfering RNAs(siRNAs), which are thenassembled into siRISCs (Figure1A). Dicer-1 processes premiRNAs and loads the resulting
miRNAs into miRISC containing
Ago-1 (Figure 1B). However,Dicer-1 also seems to berequired downstream of siRNAproduction in siRISC assembly[10–12]. It has been shown thatthe activities of Drosha andDicer-2 absolutely depend on theauxiliary dsRNA binding domain(dsRBD) proteins, Pasha andR2D2, respectively [5–8,13]. Now
Saito et al. [14] and Förstemannet al. [15] provide a missing piecein the RNAi versus miRNA puzzle:
they conclusively demonstratethat Dicer-1 also requires a
dsRBD protein to efficiently
process pre-miRNAs into miRNA
duplexes. Saito et al. [14] reliedon an RNAi-based functional
screen for Drosophila dsRBD
proteins that affect miRNAbiogenesis, while Förstemann etal. [15] searched a database forconserved dsRBDs containing
proteins. Both laboratories
identified the same candidate —a paralogue of Drosophila R2D2,
featuring two canonical and one
non-canonical dsRBDs. This
candidate was baptizedloquacious (loqs), as endogenous

20.

chemokine network. Nat. Rev. Cancer 4,
540–550.
Lapidot, T., Dar, A., and Kollet, O. (2005).
How do stem cells find their way home?
Blood, DOI: 10.1182/blood-2005-041417.

Max-Planck-Institute for Biophysical
Chemistry, Am Fassberg 11 37070,
Goettingen, Germany.
DOI: 10.1016/j.cub.2005.07.043

RNA-mediated silencing is lost in
mutant flies.
Both groups show that reduced
levels of Loqs result in theaccumulation of endogenous premiRNAs. The same phenotype isobserved when cells are depletedof Dicer-1, but not Dicer-2 or
R2D2. The physical association ofLoqs and Dicer-1 was confirmedby reciprocal coimmunoprecipitation, and did notdepend on a pre-miRNA
substrate. Saito et al. [14]demonstrate that both proteins
also co-immunoprecipitate with
Ago1, providing further evidence
that miRNA processing may bedirectly linked to the assembly of
miRISC. If assayed for processing
activity in vitro,
immunoprecipitates of Dicer-1 orLoqs readily generated mature
miRNAs from synthetic
precursors and were also foundto associate with pre- and mature
miRNAs in vivo.
So what is the actual function ofLoqs in this complex? It is certainly
not simply the stabilization of
Dicer-1, because Dicer-1 protein
levels did not decreasesignificantly in the absence ofLoqs. According to Saito et al. [14],Loqs confers substrate specificity
for pre-miRNAs to Dicer-1.
Surprisingly, Dicer-1 processes
long dsRNA as well as pre-miRNA
substrates, if Loqs is removed fromthe complex. Re-addition of Loqs
inhibited dsRNA processing andenhanced pre-miRNA processing.
But how is this achieved? Three
splice variants of loqs are known,of which only two isoforms interact
with Dicer-1. Interestingly, the third
isoform lacks the non-canonical
dsRBD, suggesting that thisdomain may be essential for
association with Dicer-1. Loqs
could enhance the bindingCurrent Biology Vol 15 No 15
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A

B

Dcr-1

Dcr-2

long dsRNA

pre-miRNAs
Loqs
Ago1

R2D2

Dcr-2

Dcr-1

‘holo-RISC’
Loqs

Ago2

Ago1
Loqs
Dcr-1

siRISC

Ago2

Ago1

m 7G

miRISC

AAAAA
Ago2
m 7G

AAAAA
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Current Biology

Figure 1. Model of the Dicer-1 and Dicer-2 pathways in Drosophila.
(A) The RNase III-like enzyme Dicer-2 (Dcr-2) is needed to process long dsRNAs into
duplex siRNAs. R2D2, a dsRBD protein, helps to sense duplex asymmetry and, together
with Dicer-2, facilitates the assembly of an 80S ‘holo-RISC’ [16], a multi-protein
complex. Both Dicer-1 and Loqs play a role downstream of siRNA production, probably as components of the putative ‘holo-RISC’ complex. Upon binding to complementary sequences in target RNAs, RISC cleaves target mRNAs. (B) A protein complex
consisting of the RNase III-like enzyme Dicer-1 (Dcr-1), the dsRBD protein Loqs andAgo1 is able to convert pre-miRNA hairpins into ~22 nt miRNA duplexes, which are thenloaded into the miRISC, composed of a single stranded miRNA bound to Ago1. Thissilencing complex inhibits translation by binding to complementary sequences in the 3′′
untranslated region of cognate messenger RNAs.

efficiency of Dicer-1 to premiRNAs, thus stabilizing thisinteraction. Alternatively, it could
position Dicer-1 in such a way thatallows efficient processing, or
introduce conformational changes
in Dicer-1 leading to an
enhancement of pre-miRNAcleavage.
In order to assess the in vivorole of loqs in greater detail,Förstemann et al. [15] took
advantage of a hypomorphic loqs
mutant. These flies showed
reduced silencing of a miRNAcontrolled transgene as well as
defects in silencing triggered bylong dsRNA. Thus, Loqs is not
only required for efficient miRNAmediated silencing, but also
cooperates with Dicer-1 in orderto enhance silencing by siRNAs,
emphasising the idea of acrosstalk between the Dicer-1 andDicer-2 pathways [10,16](Figure 1A). Furthermore, theauthors show that loqs plays acritical role in the maintenance ofgerm-line stem cells. While thetwo Loqs isoforms interacting with
Dicer-1 are present throughout
the adult fly, they show sexspecific expression in the gonads.
Mutant female flies are sterile andlack germ-line stem cells.
Mutations in piwi, a member ofthe Argonaute protein family,
result in a similar phenotype [17],implying that dicer-1 and loqs
might function together with piwi.
With the discovery of loqs, it
now seems a general feature of
RNase III enzymes to cooperatewith dsRBD-proteins in RNA
silencing. This raises a couple ofexciting questions. Both groups
highlight the homology of Loqs
with the human dsRBD protein
HIV TAR RNA binding protein(TRBP), which binds to a

hairpin-shaped TAR RNA,
implicated in the response to HIV
infection [18]. Moreover, TAR RNA
has recently been suspected to bea viral pre-miRNA [19]. The
homology between Loqs and
TRBP is especially intriguing, as
there is no clear human homolog
for R2D2. Furthermore, humanDicer is more closely related to
Drosophila’s Dicer-1. As Dicer-1
and Loqs are required for miRNAand siRNA-mediated genesilencing, and Dicer-1 even shows
weak siRNA-generating activity inthe absence of Loqs, it seems
plausible that human Dicer could
function in a fashion similar to
Dicer-1. Dicer-2 may play aunique role in Drosophila. Thus,
findings in Drosophila siRISC
biogenesis should be applied
carefully to the human system.
Beyond that, Loqs may be
involved in processing other typesof non-coding RNAs. Further
studies on other binding partners
will certainly help to characterize
its function in greater detail. There
are more than 70 miRNAs known
in Drosophila to date [20], of
which all pre-miRNAs fold into
precursor hairpins which do notshare common sequence motifs.
So how does the Dicer-1–Loqs
complex specifically recognize its
substrates? By now, this question
cannot be answered. Analyses ofthe double-stranded stem as wellas the terminal loop structure ofpre-miRNAs will make it possible
to elucidate the precise molecularmechanism of substrate
recognition.
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Microprocessor complex. Nature 432,
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Cell Division: SIN, Cytokinesis andEthanol Dependency
A novel mutant screen in fission yeast has identified the ‘ethanol
dependent’ protein etd1p as a potential link between the septation
initiation network (SIN), which initiates cytokinesis, and the actomyosin
contractile ring that drives separation of the two daughter cells at theend of mitosis.
Andrea Krapp andViesturs Simanis
Cells of the fission yeast
Schizosaccharomyces pombe arerod-shaped, grow mainly byelongation at their tips and divide
by means of a medially placed
division septum. The site ofdivision is established early inmitosis by the formation of anactomyosin-based contractile ringat the cell cortex. The ringassembles throughout mitosis andcontracts at the end of anaphase,coincident with breakdown of themitotic spindle [1].
The initiation of septum
formation is signaled from thespindle pole body by theseptation initiation network, or
SIN. The core components of the
SIN are three protein kinases —cdc7p, sid1p, sid2p, in presumed
order of action — and theirassociated regulatory/targeting
subunits — spg1p, cdc14p andmob1p, respectively. SIN
signaling also requires the protein
kinase plo1p, which stands at thehead of the signaling network. The
proteins assemble at the spindle
pole body on a scaffold
composed of the coiled-coilproteins sid4p and cdc11p. The
nucleotide status of the GTPasespg1p is regulated by thebyr4p–cdc16p GTPase activating
protein (GAP) complex.
Constitutive signaling gives rise tomultiseptated cells; in contrast,
loss-of-function mutants for anyof the core components of the SIN
or the anchoring scaffold, orfunction-specific mutants of plo1,
fail to signal septum formation,
becoming multinucleated as
growth and the nuclear cycle
continue in the absence ofcytokinesis (for reviews see [2,3]).
How do the SIN and thecontractile ring communicate? Of
the known core SIN proteins andregulators, only plo1p,
sid2p–mob1p and the negative
regulator dma1p associate withboth the ring and the spindle polebody. Association ofsid2p–mob1p with the ring isthought to be part of the trigger

17.

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RNAi in Drosophila. Cell 117, 83–94.
Cox, D.N., Chao, A., Baker, J., Chang, L.,
Qiao, D., and Lin, H. (1998). A novel classof evolutionarily conserved genes
defined by piwi are essential for stem
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Gatignol, A., Buckler-White, A.,
Berkhout, B., and Jeang, K.T. (1991).
Characterization of a human TAR RNAbinding protein that activates the HIV-1
LTR. Science 251, 1597–1600.
Bennasser, Y., Le, S.Y., Yeung, M.L., andJeang, K.T. (2004). HIV-1 encodedcandidate micro-RNAs and their cellulartargets. Retrovirology 1, 43.
Griffiths-Jones, S. (2004). The microRNA
registry. Nucleic Acids Res. 32,
D109–D111.

Institute of Molecular Biotechnology of
the Austrian Academy of Sciences,
IMBA, Vienna, Austria.
E-mail: javier.martinez@imba.oeaw.ac.at
DOI: 10.1016/j.cub.2005.07.044

for its contraction. However, theidentity of the ring component(s)that might link the SIN and thering has remained elusive.
A new study by Daga et al. [4]has identified a novel protein,
etd1p, that may link the SIN andthe contractile ring. The ‘ethanol
dependent’ etd1 gene was
identified in a screen for S. pombemutants that were either
hypersensitive to, or dependent
upon, ethanol in the medium [5].
Interestingly, etd1 has not been
identified in any screen forthermosensitive mutants,suggesting that this novel
selection scheme may allow
isolation of mutants in proteins
with structures that do notaccommodate changes conferring
thermosensitivity.
Etd1 is an essential gene: boththe etd1 mutation and a null giverise to elongated, multinucleatedcells which resemble mutants withdefects of the SIN or contractile
ring. Unfortunately, etd1p’s
sequence gives no clue to its
biochemical function, and there
are no obvious homologs in othereukaryotes.
GFP-tagged etd1p localises tothe cell cortex in interphase, witha bias for the growing cell tips.
Though it lacks any obvious
membrane-targeting sequences,the protein seems to associatewith sterol-rich membrane
domains. At the onset of mitosis it
forms a broad band overlying the