occur in the same stem cell population, the
two events are temporally separated, with
WUS being completely repressed ~2 days
after AG induction. The precise timing of
WUS repression allows the correct number of
cells to be produced for reproductive organ
formation. Too early and flowers have fewer
stamens and no carpel; one day too late and
flowers become partially indeterminate with
additional and abnormal organs (3). Sun et al.
now explain how the 2-day delay is achieved,
through linking cycles of cell division to transcriptional derepression.
A key component in the WUS-AG pathway is the transcriptional repressor
KNUCKLES (KNU), which represses WUS in stage
6 flowers (2). KNU is repressed by histone
H3 lysine 27 trimethylation (H3K27me3),
a repressive modification established by the
Polycomb repressive complex 2 (PRC2). In
developing flowers, AG activates KNU by
binding to its promoter, but KNU expression
only becomes detectable ~2 days after the
initial AG binding, resulting in a substantial
decrease in WUS expression. The delay in
KNU activation is likely caused by the gradual loss of the repressive H3K27me3 mark at
the KNU gene (3).
Sun et al. observed that the ~2-day delay
depends on cell-cycle progression, suggest-
ing that the “timer” may be a “counter” that
monitors the number of cell divisions. The
authors hypothesized that AG binding inter-
feres with the maintenance of H3K27me3 at
KNU: Incorporation of unmodified H3 after
roughly two rounds of cell divisions suffi-
ciently dilutes the preexisting H3K27me3 to
allow KNU expression. Indeed, a short region
in the KNU promoter immediately adjacent to
the AG binding site was found to be necessary
and sufficient for recruiting a core compo-
nent of the PRC2, FERTILIZATION-INDE-
PENDENT ENDOSPERM (FIE). Critically,
Sun et al. show that AG binding leads to the
immediate eviction of FIE.
AG also directly binds to the WUS gene
to repress WUS expression in an H3K27me3-
dependent manner (4) (see the figure). The
two independent modes of WUS repression
by AG likely have different consequences.
Whereas the direct repression of WUS by
AG is relatively mild but immediate (in
stage 4 to 6 flowers), the repression of WUS
through KNU is more complete but tempo-
rally delayed (at stage 6) (4). How AG recruits
PRC2 to repress some genes but evicts the
same complex to activate other genes remains
an open question.
Despite the importance of H3K27me3
in regulating plant development (targeting
~4000 to 5000 Arabidopsis genes) (5), relatively little is known about the mechanisms
responsible for the establishment, mainte-
nance, and removal of H3K27me3. In the
fruit fly Drosophila, PRC2 is targeted to
specific sites called Polycomb response elements (PREs) by the Pho-repressive complex
(PhoRC) (6). Genome-wide analysis of FIE
binding sites in Arabidopsis identified several motifs, including a GA-repeat motif frequently found in Drosophila PREs (7). However, because plants do not have Rho homologs, whether these motifs are involved in targeting PRC2 is not fully understood. The FIE
binding site at KNU adds to a short but growing list of functional PREs in plants (8). Notably, the KNU PRE also contains GA-repeats,
suggesting that similar motifs found in FIE
binding sites may indeed function as PREs.
However, several different motifs are present
at distinct subsets of FIE binding sites (7, 9),
suggesting that PRC2 recruitment in Arabi-
dopsis is likely carried out by many different
factors (8). Such a “recruiting by committee”
approach may provide more versatility dur-
ing the predominantly postembryonic devel-
opment in plants characterized by continued
growth and differentiation from small popu-
lations of stem cells.
The KNU PRE is located in the promoter,
but FIE binding and H3K27me3 extend
downstream to cover the entire transcribed
region of KNU. The absolute requirement of
the KNU PRE for FIE binding suggests that
PRC2 is initially recruited to the PRE and
then spreads unidirectionally to the remain-
der of KNU, establishing H3K27me3 along
the way. In addition, the cell-cycle–depen-
dent loss of H3K27me3 after FIE is displaced
by AG indicates that PRC2 needs to be con-
stantly recruited to the KNU PRE to maintain
H3K27me3 after cell divisions. It is not yet
clear whether additional factors facilitate the
spreading and maintenance of H3K27me3.
The repressive H3K27me3 mark can be
removed enzymically by the REF6 family
of JmjC-domain–containing H3K27me3
demethylases (10). The repressive state
associated with H3K27me3 can also be
overcome by strong transcription activators and/or chromatin-remodeling factors,
with H3K27me3-containing histones being
subsequently lost during active transcription. For example, AG itself is repressed
by H3K27me3 during vegetative growth,
and LFY recruits the SWI2/SNF2 chromatin-remodeling adenosine triphosphatases SPLAYED (SYD) and BRAHMA
(BRM) to AG for transcriptional activation
(11). LFY binding sites frequently colocalize with PRE-like sequences, indicating
that LFY binding may also displace PRC2
(12). Finally, when PRC2 is displaced from
the chromatin, H3K27me3 can be gradually
diluted by the incorporation of unmodified
histones during chromatin replication. Sun
et al. not only provide a clear example of this
scenario, but also reveal its biological function in specifying delayed and cell-cycle–
dependent transcriptional activation.
1. B. Sun et al., Science 343, 1248559 (2014); 10.1126/
2. J. U. Lohmann et al., Cell 105, 793 (2001).
3. B. Sun, Y. Xu, K. H. Ng, T. Ito, Genes Dev. 23, 1791
4. X. Liu et al., Plant Cell 23, 3654 (2011).
5. X. Zhang et al., PLOS Biol. 5, e129 (2007).
6. J. Müller, J. A. Kassis, Curr. Opin. Genet. Dev. 16, 476
7. W. Deng et al., BMC Genomics 14, 593 (2013).
8. C. He, H. Huang, L. Xu, Front. Plant Sci. 4, 454 (2013).
9. N. Berger, B. Dubreucq, F. Roudier, C. Dubos, L. Lepiniec,
Plant Cell 23, 4065 (2011).
10. F. Lu, X. Cui, S. Zhang, T. Jenuwein, X. Cao, Nat. Genet.
43, 715 (2011).
11. M. F. Wu et al., Proc. Natl. Acad. Sci. U.S.A. 109, 3576
12. C. M. Winter et al., Dev. Cell 20, 430 (2011).
Mild WUS repression by AG
AG evicts PRC2 from
Strong WUS repression by KNU
AG WUS KNU
A timed transcription circuit makes flowers. A network of transcription factors act in Arabidopsis flower
development to balance stem cell maintenance and cellular differentiation.