|
|
|
|
|
|
|
||||
| Home Help Feedback Subscriptions Archive Search Table of Contents | ||||
First published online 11 February 2009
doi: 10.1242/dev.029363
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Departments of Pediatrics and Molecular Biology, Division of Pediatric Hematology-Oncology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390, USA.
* Author for correspondence (e-mail: scott.cameron{at}utsouthwestern.edu)
Accepted 9 January 2009
 |
SUMMARY |
|---|
TOP
SUMMARY
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
|---|
Key words: BMI-1, C. elegans, Histone modification, Polycomb, Neuronal migration, ring1B
|
INTRODUCTION |
|---|
|
TOP
SUMMARY
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
|---|
;
Lewis, 1978;
Nusslein-Volhard et al., 1985;
Simon et al., 1992).
Biochemical and molecular analyses have defined at least three protein
complexes, called Polycomb repressive complex 2 (PRC2), PhoRC and PRC1. PRC2
is a histone methyltransferase that methylates Histone 3 lysine 27, and
includes the proteins Enhancer of zeste, Extra sex combs, Su(z)12 and Nurf-55
(Cao et al., 2002;
Czermin et al., 2002;
Kuzmichev et al., 2002;
Muller et al., 2002). The
H3K27me3 histone mark is in turn recognized by the PRC1 complex
(Cao et al., 2002;
Czermin et al., 2002;
Kuzmichev et al., 2002;
Saurin et al., 2001), which
has as core components in human cells Ring1, Ring2/Ring1B, Bmi-1 and HPH2
(Wang et al., 2004a). PRC1 is
recruited to sites methylated by PRC2, where the Ring1B protein of PRC1
monoubiquitylates H2A at lysine 119 (Wang
et al., 2004a; Wang et al.,
2004b). In this complex, BMI-1 enhances the stability and
enzymatic activity of Ring1B (Cao et al.,
2005). PRC1 inhibits gene expression through mechanisms that are
as yet not entirely clear but which may involve repressing the initiation of
transcription, inhibiting nucleosome remodeling, regulating association of
linker histone H1 and/or chromatin compaction
(Francis et al., 2004;
King et al., 2002;
Levine et al., 2002;
Shao et al., 1999;
Zhu et al., 2007). Recent
genome-wide identification of Polycomb group target genes has also shown that
some targets are transcribed despite the presence of Polycomb group proteins
on the gene, suggesting that repression may not be a universal outcome at all
loci (Boyer et al., 2006;
Bracken et al., 2006;
Lee et al., 2006;
Tolhuis et al., 2006).
Caenorhabditis elegans orthologs of the PRC2 complex have been
identified and well characterized. These include the products of the
mes-2, mes-3 and mes-6 genes,
mutations in which result in maternal effect sterility
(Capowski et al., 1991). The
MES proteins form a complex in which the SET domain of MES-2 mediates di- and
trimethylation of H3K27 (Bender et al.,
2004; Holdeman et al.,
1998). The H3K27me3 mark is concentrated on the X chromosome of
wild-type animals, and transcription of X chromosome genes is normally
silenced in the germline (Fong et al.,
2002; Xu et al.,
2001b). In mes-2, mes-3 and
mes-6 mutants chromatin marks associated with active
chromatin are found on the X chromosome and, according to the current model,
inappropriate expression of X chromosome genes in the germline is responsible
for the degeneration of germ cells and the sterility observed in mes
mutants (Fong et al.,
2002).
In addition to their role in the germline, mes genes also act in
somatic cells. mes-2, mes-3 and
mes-6 mutants have weak but reproducible defects consistent
with abnormal Hox gene activity, in agreement with the classical role of
Polycomb group genes as repressors of Hox gene activity
(Ross and Zarkower, 2003). The
abnormalities include subtle defects in migration by specific neurons,
expansion of the domains of Hox gene expression, and mislocalization of
sensory rays in the male tail. The genetic screens that identified the
mes genes did not identify genes homologous to PRC1 components, and
the C. elegans genome does not encode obvious homologs of many of the
components of PRC1. These data suggest that either C. elegans lacks a
PRC1 complex, which would suggest that PRC2 function could be uncoupled from
PRC1 function, or that the function, composition or amino acid sequences of
the proteins in a PRC1-like complex are sufficiently divergent to make
recognizing them difficult.
Here we describe the genes mig-32 and spat-3, which encode homologs of the human PRC1 core components Bmi-1 and Ring1B, respectively. Consistent with MIG-32 and SPAT-3 being functionally analogous to PRC1, ubiquitylation of H2A is markedly reduced or absent in mig-32 and spat-3 mutants. Both mutants have defects in their nervous system that are similar those of mes mutants. Surprisingly, unlike mes mutants, mig-32 and spat-3 mutants are fertile, suggesting that H2A ubiquitylation by PRC1 is not required in the germline for MES/PRC2 function.
|
MATERIALS AND METHODS |
|---|
|
TOP
SUMMARY
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
|---|
).
Mutations and mapped integrated transgenes used were as follows. LGI:
mig-1(e1787); mes-3(bn35); unc-73(e936); unc-40(e271);
ced-1(e1735). LGII: mes-2(bn11); unc-4(e120); muIs32
[Pmec-7gfp]
(Pujol et al., 2000);
muIs16 [Pmab-5gfp]
(Hunter et al., 1999);
nIs128 [Ppkd-2gfp]
(Yu et al., 2003). LGIII:
pal-1(e2091); mig-10(ct41); unc-119(ed3). LGIV: plx-1(nc37);
ced-10(n1993); mes-6(bn38); dpy-20(e1282); mig-32(n4275) (kindly provided
by the Horvitz lab, MIT, Cambridge MA, USA), mig-32(tm1807),
mig-32(tm1684) (kindly provided by the Mitani lab, Tokyo Women's
University, Tokyo, Japan); unc-31(e928). LGV: him-5(e1490).
LGX: nIs106 [Plin-11gfp]
(Cameron et al., 2002),
kyIs4 [Pceh-23gfp]
(Zallen et al., 1998);
oxIs12 [Punc-47gfp]
(McIntire et al., 1997);
unc-6(ev400); mig-2(mu28); slt-1(eh15); lin-15(n765ts);
spat-3(gk22).
Integrated transgenes not mapped to a chromosome included: kyIs39
[Psra-6gfp]
(Troemel et al., 1995);
bxIs13 [Pegl-5gfp]
(Zhang and Emmons, 2001);
mxIs23 [Pmig-32mig-32:gfp].
Extra-chromosomal arrays included: rtEx238 [Pnlp-1gfp] (kindly provided by Leon Avery, University of Texas Southwestern Medical Center, Dallas TX, USA); mxEx53 [Pplx-1mig-32:cfp].
Imaging
Transgenic animals were imaged using a Zeiss Axiophot. To quantitate HSN
and ALM migrations, the distance along the body axis between the rectum and
the HSN or ALM nuclei was determined using Openlab software and compared to
the distance to the vulva.
Construction and analysis of mig-32/Df animals
sDf62 unc-31(e169)/nT1(IV); +/nT1(V) hermaphrodites were mated
with him-5(e1490) males and cross-progeny males were mated with
mig-32(n4275) unc-31(e928) hermaphrodites. The position of ray-1 in
Unc males was determined.
Plasmid constructs and Pmig-32mig-32:gfp construction
To construct Pmig-32 mig-32:gfp, an 8065 bp
BamHI/SphI fragment of the F11A10 cosmid was cloned into
pUC19. An XmaI fragment of pPD102.33, including the GFP coding
sequences, was cloned into the AgeI site in the first exon of
mig-32. This construct was injected into lin-15(765ts)
worms, integrated and backcrossed three times to mig-32(n4275);
him-5(e1490).
To construct Pplx-1mig-32::cfp, an
EcoRI-XbaI fragment of the full-length yk1321a11
mig-32 cDNA was cloned into pUC19. CFP coding sequences were
amplified by PCR and cloned into mig-32 at the AgeI site.
The mig-32 start codon was converted into an NsiI site using
site-directed mutagenesis, and 2.6 kb of the plx-1 promoter
(Dalpe et al., 2004) was PCR
amplified and cloned into the NsiI site. Constructs were injected
into unc-119(ed3); mig-32(n4275); him-5(e1490) worms using
unc-119 as the co-injection marker. We used the position of Ray 1 as
an assay for mig-32 function. Of transgenic mig-32(n4275)
mutants carrying the Pmig-32mig-32:gfp reporter
(n=50), 28±6% had anterior Ray 1s, compared with 64±5%
of mig-32 mutants (n=100).
To construct the spat-3a(RNAi) plasmid, 889 nucleotides of the spat-3a genomic region flanking the sequences that encode the RING domain were amplified by PCR and cloned into the L4440 plasmid as a HindIII-BglII restriction fragment.
Core histone extraction and western blotting
Packed L1-stage worms (0.5 ml) were harvested from freshly-starved plates.
Worms were sonicated in 5 ml NIB buffer [15 mM PIPES (pH 6.8), 5 mM
MgCl2, 60 mM KCl, 0.25 M sucrose, 15 mM NaCl, 1 mM
CaCl2, 0.8% Triton and protease inhibitors]
(Jackson et al., 2004) until
the cuticle was completely broken. Lysate was centrifuged at 10,000
g for 10 minutes. The pellet was resuspended in 2 ml of 0.4 N
H2SO4 and incubated at 4°C for 1 hour to overnight.
The soluble material was dialyzed with PBS or precipitated with 20% TCA.
TCA-precipitated histone extract was washed with cold acetone three times and
the final pellet dried and dissolved in water. For detection of H2A, rabbit
antiserum against histone H2A from Upstate (#07-146) was used at 1/1000
dilution. For detection of ubiquitylated histones, monoclonal anti-ubiquitin
(clone P4D1) from Cell Signaling Technology was used at 1/1000 dilution.
|
RESULTS |
|---|
|
TOP
SUMMARY
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
|---|
We first confirmed the gene structure predictions for mig-32 by
determining the DNA sequences of five full-length mig-32 cDNAs, and
found that all were identical in the predicted coding sequences and were
SL2-spliced, consistent with genome database predictions that mig-32
is the second gene in a three-gene operon (WormBase,
http://www.wormbase.org,
release WS193, July, 2008) (Spieth et al.,
1993). mig-32 is predicted to encode a 542 amino acid
protein with no close homologs in the completely sequenced C. elegans
genome, and with a predicted RING domain as the only domain recognized by Pfam
(Bateman et al., 2002). Using
BLAST searches we identified MIG-32 homologs in vertebrate genomes, including
six homologous proteins from humans (Fig.
1). Included in this group are the BMI-1 protein and the related
proteins MEL-18/RFP110, NSPc1/PcGRF1 and Pcgf5, which participate in
PRC1-related complexes (Alkema et al.,
1997; Sanchez et al.,
2007; Trimarchi et al.,
2001). The Drosophila melanogaster genome contains two
homologs: Posterior sex combs, a component of the Drosophila
PRC1 complex (Saurin et al.,
2001), and Lethal (3) 73 Ah, an essential gene
(Belote et al., 1990;
Irminger-Finger and Nothiger,
1995). We identified a single MIG-32 homolog in the genomic
sequences of each of the nematodes C. briggsae and C.
remanei (Fig. 1). These
data suggest that MIG-32 is a RING domain protein most similar to core
components of human and Drosophila PRC1 and related complexes.
We identified three deletion alleles of mig-32 (see Materials and methods). Each of the alleles deletes mig-32 genomic coding sequences and not coding sequences of the upstream or downstream genes in the operon. mig-32(n4275) mutants, and mutants carrying the tm1684 and tm1807 alleles, are homozygous viable and have defects in the male tail and other structures as outlined briefly here and in greater detail below.
The male tail has nine bilateral sensory ray structures
(Emmons, 2005). We used the
defect in the position of one of these rays to examine the consequences for
mig-32 function of the three deletion alleles. All three alleles
result in qualitatively and quantitatively similar defects in the position of
Ray 1, with 60-80% of the mutants having an anterior Ray 1; all three also
result in similar defects in migration of the HSN neurons (see below). The
alleles are recessive, and 75% of mutants of genotype n4275 over
sDf62, a chromosomal deficiency that deletes the mig-32
region, have defects in Ray 1 position similar to those of mig-32
homozygotes (data not shown).
Three observations suggest that the deletions specifically affect
mig-32 function and not the function of F11A10.8 or
F11A10.6, the upstream and downstream genes in the operon,
respectively. First, F11A10.8 encodes a very-well-conserved homolog
of human CPSF4, a splicing factor (WormBase). Inactivation of
F11A10.8 by RNAi is lethal, as is a deletion mutation,
ok844, which deletes parts of the F11A10.8 and mig-32 coding
regions (WormBase), suggesting that the mig-32(n4275), tm1684 and
tm1807 deletions do not severely impair F11A10.8 function.
Second, RNAi of mig-32, which primarily targets processed mRNA
(Fire et al., 1998), results
in an anterior position of Ray 1, as we observed in mig-32 mutants.
Third, expression using the plx-1 promoter
(Dalpe et al., 2004) of a
mig-32 cDNA in the male tail rescued the anterior Ray 1 defects of
mig-32(n4275) mutants. Specifically, of 50 mig-32(n4275)
mutants carrying a Pplx-1mig-32:cfp transgene,
12±5% (standard error of the proportion) had an anterior Ray 1 compared
with 64±5% of 100 uninjected controls, and in contrast to
mig-32 mutant males the transgenic males mated efficiently (see below
and data not shown). These data suggest that the n4275, tm1684 and
tm1807 alleles are strong loss-of-function or null alleles of
mig-32. Unless otherwise indicated, we used the
mig-32(n4275) allele for the experiments described here.
|
|
). spat-3 is predicted to encode two large protein
products, SPAT-3A and SPAT-3B, generated by alternative promoters and
supported by partial cDNA evidence (WormBase). The SPAT-3A protein is 2471
amino acids and contains a RING domain at the N-terminus that is most similar
to that of the Ring1B proteins of mammals, although the similarity is very
poor (Fig. 1). SPAT-3B lacks
the RING domain, and neither protein has additional recognized domains. Using
the BLAST algorithm, SPAT-3A is the RING domain protein in the C.
elegans genome most similar to Ring1B; the next most similar protein is
MIG-32 (Fig. 1). A deletion
allele of spat-3, gk22, deletes the genomic sequences common to the
spat-3a and spat-3b transcripts, and if transcribed and
translated is predicted to truncate the SPAT-3A protein after position 1620.
As such, it may represent a loss-of-function but not null allele, as the RING
domain is very near the N-terminus (Fig.
1). spat-3(gk22) mutants are viable and healthy, with a
brood size that is 90% of wild-type animals
(Table 1).
MIG-32 and SPAT-3A are required for ubiquitylation of histone H2A
The defined biochemical function of PRC1 is ubiquitylation of histone H2A
at position 119. In PRC1, the Ring1B protein serves as the E3 that catalyzes
H2A ubiquitylation (Wang et al.,
2004a). Other core components, especially BMI-1, stimulate the
catalytic activity of Ring1B (Cao et al.,
2005; Li et al.,
2006; Wei et al.,
2006), possibly by promoting folding and stability of Ring1B
(Ben-Saadon et al., 2006). We
therefore asked whether H2A ubiquitylation was abnormal in mig-32 and
spat-3 mutants. We analyzed histone modification using western blots
of acid-extracted histones from wild-type, mig-32 and spat-3
mutant C. elegans. Using an H2A-specific antibody and extracts from
wild-type animals, we detected H2A and a rare, higher molecular weight band
that migrated at the size expected for ubiquitin-modified H2A; this band was
not detected in extracts from mig-32 or spat-3 mutants
(Fig. 2). Using an antibody
that detects ubiquitin with histone extracts from wild-type animals, we
detected two bands that correspond to the predicted molecular weights of H2A
and H2B, both of which are modified by ubiquitylation
(Osley, 2006). H2A, the
smaller of these bands, is not detected in extracts from mig-32
mutants and is greatly reduced in spat-3 mutants
(Fig. 2). These data suggest
that MIG-32 and SPAT-3A are required for ubiquitylation of Histone H2A, the
defining biochemical function of PRC1.
mig-32 is broadly expressed, localized to nuclei and concentrated within nucleoli
In situ hybridization using a mig-32 cDNA suggested that the gene
is expressed prominently in the C. elegans germline (Y. Kohara,
personal communication). We constructed a rescuing GFP reporter to determine
the expression pattern in somatic cells (see Materials and methods).
Consistent with MIG-32 acting as a modifier of chromatin, the
Pmig-32mig-32:gfp reporter is expressed broadly in most or
all nuclei, beginning early in embryogenesis and continuing in larval
development and into adulthood of males and hermaphrodites
(Fig. 3 and data not shown).
Expression is predominantly nuclear, with relatively bright intranuclear areas
of fluorescence that correspond with nucleoli evident as seen with Nomarski
optics, particularly in some hypodermal cells where nucleoli are easily
identified (Fig. 3).
Transfection of mammalian cells with a FLAG-epitope-tagged MIG-32 showed
nuclear expression concentrated in a shell around nucleoli, suggesting that
the interactions that determine subcellular localization may be evolutionarily
conserved (Fig. 3). We
considered the possibility that localization of MIG-32 might depend upon
mes gene activity. To test this, we examined the expression and
localization of the Pmig-32mig-32:gfp reporter in
mes-2 mutants and observed that expression of the reporter is
brighter in the mes-2 mutant background but that the fusion protein
remains localized to nuclei and nucleoli (data not shown).
|
;
Ross and Zarkower, 2003). We
constructed double mutants between mig-32 and putative null alleles
of each of the mes genes. Strikingly, the Ray 1 position defects of
these double mutants were not enhanced
(Fig. 4), suggesting that
mig-32 acts in the same genetic pathway as the mes genes to
position Ray 1.
|
; Ginzburg et al.,
2002), acting through the unc-73 guanine nucleotide
exchange factor and the ced-10/Rac mig-2/Rho GTPases
(Dalpe et al., 2004). We
constructed double mutants between mig-32 and each of these genes and
examined the position of Ray 1 in the male tails of the animals. In each case
we identified significantly enhanced defects in the position of Ray 1
(Fig. 4), suggesting that
mig-32 acts parallel to these pathways to position Ray 1.
mig-32 and spat-3a are required for HSN neuronal migration and axon extension
We observed that mig-32 mutants were variably egg-laying
defective, with some animals in a population carrying more eggs than wild-type
animals. In staged adults the wild-type strain N2 carried an average of
17.5±5 eggs, compared with 28.8±11 eggs in mig-32
mutants (n=55 animals for each genotype, P<0.0001,
unpaired two-tailed t-test). Egg-laying requires a vulva through
which the eggs are laid, muscles to expel the eggs, and neurons to control the
vulval muscles (Trent et al.,
1983). We found that the two HSN neurons, which are essential for
egg-laying (Trent et al.,
1983), are often abnormal in mig-32 mutants (Figs
5 and
6). During embryogenesis the
HSN neurons migrate from the tail to the midbody
(Sulston et al., 1983). Using
the Pnlp-1gfp reporter to identify the HSN neurons
(Li et al., 1999), we observed
that the HSNs of mig-32(n4275) mutants failed to reach the midbody in
41% of mutants; by comparison, all HSNs migrated to their normal position in
otherwise wild-type animals carrying the Pnlp–1gfp
reporter (Fig. 6). The axons
extended by the HSN neurons were also abnormal. In wild-type animals, each HSN
extends an axon from the vulval region ventrally to the ventral nerve cord;
the axons then turn anterior and extend to the head. Of the HSN axons of
mig-32 mutants, 56% failed to reach the head; by comparison, all HSN
axons extended to the head of otherwise wild-type animals carrying the
Pnlp-1gfp reporter (Figs
5 and
6). All HSNs of mig-32
mutants expressed the Pnlp-1gfp reporter, suggesting that
the HSN neurons correctly establish their identity and that the defects in HSN
migration result from a requirement for mig-32 in other processes
important for migration and axon extension. The defects in HSN migration and
axon extension are likely to account for the variable defects in egg-laying we
observed in mig-32 mutants; such variability has been associated with
other mutants with defects in HSN migration
(Desai et al., 1988).
|
|
mig-32 mutants have additional defects in laterality of commissures, process extension and midline crossing by axons
Using gfp reporters, we surveyed mig-32 mutants for
defects in neuronal migration and processes extended by other neurons. The
Punc-47gfp reporter is expressed in the VD and DD
motoneurons of the ventral nerve cord
(McIntire et al., 1997). In
otherwise wild-type animals carrying the Punc-47gfp
reporter, the VD and DD neurons extend commissures from the ventral nerve cord
laterally along the body wall to the dorsal nerve cord. In wild-type animals,
all but one pair of commissures track along the right side of the animal; only
4% of otherwise wild-type animals carrying the Punc-47gfp
reporter had more than one pair of commissures on the wrong side
(Fig. 5;
Table 2). By contrast, 63% of
mig-32 mutants had more than two left-sided commissures, with some
animals having as many as five commissures on the wrong side
(Fig. 5;
Table 2). The number and
positions of VD and DD neurons, the total number of commissures and the
expression of Punc-47gfp were normal in mig-32
mutants (data not shown).
|
). Using
the Psra-6gfp reporter, which is expressed in the PVQ
neurons (Troemel et al.,
1995), we observed that the PVQ axons of mig-32 mutants
did not respect the midline boundary and crossed inappropriately
(Fig. 5;
Table 2). Of the PVQ axons of
mig-32 mutants, 26% crossed the midline inappropriately, compared
with 8% of otherwise wild-type animals carrying the
Psra-6gfp reporter. All PVQ neurons of mig-32
mutants expressed Psra-6gfp at levels similar to wild-type
animals.
|
). The Q
cells migrate anteriorly and divide, with the QR cell (right-sided Q cell)
generating the AVM mechanosensory neuron and QL (left-sided Q cell) the PVM
neuron. The ALM neurons migrate posteriorly from the head. Using the
Pmec-7gfp reporter to label the mechanosensory neurons and
their axons, we found that the ALM neurons of mig-32 mutants were
slightly defective in posterior migration. Of 150 ALM neurons observed in
mig-32 mutants, 15 ALMs did not complete their migrations
(Fig. 6). The processes of the
ALM neurons were similar in mig-32 and wild-type animals (data not
shown). The PLM neurons did not migrate, but their posterior processes were
often foreshortened in mig-32 mutants. Of 100 PLM neurons of
mig-32 mutants, the posterior process terminated prematurely in 55%
of animals, extending less than half the normal distance. By contrast, the
posterior processes of the PLM neurons terminated prematurely in only 6% of
100 wild-type animals. The locations of the AVM and PVM neurons and the
morphology of their processes were similar, comparing 75 mig-32
mutants with 50 wild-type animals (data not shown). Migration of the distal tip cells, which are somatic cells that lead the anterior and posterior arms of the proliferating germline along the body wall, appeared normal, comparing 23 mig-32 mutants with 18 wild-type animals (data not shown).
In summary, mig-32 is required for normal migration of the HSN neurons and for extension of some neuronal processes. It participates in ensuring that VD neuronal commissures extend along the correct side of the animal, and that the PVQ axons do not cross the midline inappropriately. The defects we observed in mig-32 mutants are unlikely to be a result of markedly altered neuronal differentiation, because expression of all the gfp reporters used for these experiments were expressed in the expected patterns in mig-32 mutants (a list of reporters used is found in Materials and methods).
mig-32 acts parallel to most known pathways that act in HSN migration
Several genetic pathways have been identified that ensure correct migration
of the HSN neurons from the tail to the vulva. To determine whether
mig-32 acts within one of these pathways, we constructed double
mutants between mig-32 and other genes that regulate HSN migration,
including mig-1, a Wnt receptor and Frizzled homolog
(Pan et al., 2006),
mig-10, a cytoplasmic protein that mediates attractive and repulsive
guidance signals by unc-6/Netrin and slt-1
(Chang et al., 2006;
Quinn et al., 2006),
respectively, and slt-1 (Hao et
al., 2001). In each case, the double mutants with mig-32
had significantly enhanced defects in HSN migration assayed with the
Pnlp-1gfp reporter
(Fig. 6). These data suggest
that mig-32 acts genetically parallel to these pathways to promote
HSN migration.
mig-32 and spat-3 repress ectopic vulval development
The lin-15 locus is an operon that includes two genes,
lin-15A and lin-15B, both of which are redundant repressors
of a vulval fate for hypodermal descendants of the P cells in the vulval
equivalence group (Clark et al.,
1994). Mutants carrying the temperature-sensitive allele
n765 are morphologically normal when raised at 15°C but show a
multiple vulva (Muv) phenotype when raised at 20°C; the
lin-15(n765ts) allele has a single mutation that impairs function of
both lin-15A and lin-15B
(Cui et al., 2008). During
strain constructions, we observed that mig-32; lin-15(n765ts) double
mutants are 100% Muv at 15°C (data not shown). We used this observation to
test whether spat-3 and mig-32 might act similarly to
repress vulval fates. We subjected lin-15(n765ts) mutants to
mig-32(RNAi) or spat-3a(RNAi) and raised the animals at
15°C; the spat-3a(RNAi) construct specifically targeted the
spat-3a transcript that encodes the RING domain. Reducing either
mig-32 or spat-3a activity resulted in a highly penetrant
Muv phenotype. Seventy-three percent of lin-15(n765ts); mig-32(RNAi)
mutants (n=320) and 42% of lin-15(n765ts); spat-3(RNAi)
mutants (n=200) were Muv, compared with 3% of lin-15(n765ts);
unc-22(RNAi) mutants (n=100). These data suggest that
mig-32 and spat-3a act similarly to repress vulval fates in
hypodermal cells that do not normally contribute to vulval development.
|
DISCUSSION |
|---|
|
TOP
SUMMARY
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
|---|
;
Li et al., 2006;
Wang et al., 2004a;
Wang et al., 2004b).
Consistent with MIG-32 and SPAT-3A acting together in a PRC1-like complex,
mig-32 and spat-3 mutants are markedly defective in H2A
ubiquitylation and have very similar defects in the anatomy of their nervous
systems and as repressors of ectopic vulval fates in hypodermal cells. We
therefore propose that MIG-32 and SPAT-3A are core subunits of a PRC1-like
complex in C. elegans, although we have not yet demonstrated complex
formation by these proteins.
The relationship of MIG-32 and SPAT-3A with PcG complexes of C. elegans
There are at least two complexes in C. elegans that are
functionally related to the Drosophila and mammalian Polycomb
repression complexes. Strong genetic and biochemical evidence indicates that
the MES-2–MES-3–MES-6 complex is functionally analogous to the
PRC2 complex (Xu et al.,
2001a), which places the histone H3K27me3 mark characteristic of
Polycomb repression (Bender et al.,
2004; Fong et al.,
2002; Holdeman et al.,
1998; Xu et al.,
2001a). mes mutants are sterile, probably as a
consequence of inappropriate expression of genes normally silenced in the
developing germ cells. If PRC1 were required for regulation of gene expression
by PRC2, mig-32 and spat-3 mutants might be expected to
share the Mes phenotype of sterility. However, both mutants are healthy and
fertile. mig-32 and spat-3 could be redundant with other
proteins or complexes responsible for H2A ubiquitylation in the germline, or
H2A ubiquitylation could be dispensable for the silencing function that is
thought to be the essential role of the mes genes in ensuring
germline integrity. We favor the latter possibility, as western blots of
L1-stage wild-type animals with rudimentary germlines have relatively high
steady state levels of ubiquitylated H2A and H2B, but we detected little
ubiquitylated histone in L4 or young adult animals with proliferative
germlines (our unpublished observations).
More recently the sop-2, sor-1 and sor-3 genes have been
proposed as components of a distinct Polycomb complex-like repressive
mechanism in C. elegans. Mutations affecting these genes result in
expanded domains of Hox gene expression
(Wang et al., 2004a;
Yang et al., 2007;
Zhang et al., 2003;
Zhang et al., 2004). All are
essential genes, and mutants carrying partial loss-of-function alleles have
severe defects not observed in mes null mutants, suggesting that the
sop and sor genes have many important functions in somatic
cells and do not simply maintain the pattern of gene expression established by
the mes genes, which have subtle functions in somatic cells
(Ross and Zarkower, 2003). The
pattern of nuclear fluorescence we observed for a rescuing MIG-32:GFP fusion
protein suggests that MIG-32 does not co-localize with the SOP-2 and SOR-1
proteins (Saurin et al., 1998;
Yang et al., 2007;
Zhang et al., 2006). Given the
numerous and severe defects of the sop and sor mutants and
the comparatively limited defects of mig-32 and spat-3
mutants, MIG-32 and SPAT-3A are unlikely to be essential components of a
putative SOP/SOR Polycomb-like complex.
The roles of Polycomb complexes in nervous system development
The mig-32 homolog Bmi-1 has been intensively studied
following its isolation as a target gene upregulated by proviral integration
in Eµ-myc-driven lymphomas in mice
(Haupt et al., 1991;
van Lohuizen et al., 1991). In
the mammalian nervous system Bmi-1 is required for the self-renewal
of neural stem cells (Molofsky et al.,
2005; Molofsky et al.,
2003), and epigenetic regulation of the cell cycle is a crucial
function of Bmi-1 in the hematopoietic and nervous systems
(Jacobs et al., 1999;
Molofsky et al., 2005;
Molofsky et al., 2003). We
have not observed abnormalities in cell numbers in mig-32 mutants
that would suggest an essential role in the regulation of the cell cycle, but
we have not directly examined this possibility.
Expression of additional MIG-32 homologs in the mammalian nervous system
has been reported (Gunster et al.,
1997; Jacobs et al.,
1999; Kim et al.,
2005; Leung et al.,
2004; Molofsky et al.,
2003; Nunes et al.,
2001; Schoorlemmer et al.,
1997; Shakhova et al.,
2005; van der Lugt et al.,
1994), but with the exception of Bmi-1 little is known
about what these genes contribute to nervous system function. Our data suggest
that epigenetic regulation of gene expression by PRC1 and related complexes
will participate in neuronal migration and process extension, but the precise
basis for the defects we observe in mig-32 and spat-3
mutants is not yet clear. Specifically, does PRC1 regulate transcription of
individual gene targets that are crucial regulators of individual cell
migrations or process extensions, or does loss of PRC1 result in a `noisy'
pattern of gene expression to which some cells are more sensitive? Recent
genomic screens in Drosophila and vertebrates have identified targets
of Polycomb repression complexes (Boyer et
al., 2006; Bracken et al.,
2006; Lee et al.,
2006; Negre et al.,
2006; Schwartz et al.,
2006; Tolhuis et al.,
2006). Many of these biochemically defined targets are involved in
nervous system patterning, and our data raise the possibility that
Polycomb-group complexes regulate these targets in a functionally important
way in the developing nervous system. Our observation that most cells appear
to adopt fates similar to those of wild-type animals, as suggested by normal
expression of the cell-type-specific gfp reporters used in this
study, suggests that mig-32 is not crucial for establishing cell
fates, but instead acts in a subtle manner to refine cellular phenotypes.
Defects in Hox gene expression are unlikely to be central for the mig-32 or spat-3a mutant phenotypes
Mutations affecting Hox genes result in abnormal neuronal migrations in
C. elegans (Baum et al.,
1999; Chalfie and Sulston,
1981; Chisholm,
1991; Clark et al.,
1993; Harris et al.,
1996; Kenyon,
1986; Salser and Kenyon,
1992; Wang et al.,
1993), and given the classical role of Polycomb family members as
repressors of Hox gene activity we asked whether abnormal Hox gene activity
might underlie the defects in the nervous system we observed. However, our
data suggest that regulation of Hox gene expression by mig-32 and
spat-3 may be subtle. In general, the migration and neuronal process
extension defects of mig-32 and spat-3 mutants have little
in common with those observed in gain- or loss-of-function Hox mutants. In
addition, the expression domains of the Pegl-5gfp Hox and
Ppkd-2gfp reporters do not appear to be expanded in
mig-32 mutants, and mig-32 mutations do not suppress
pal-1 mutations, which reduce Hox activity (our unpublished
observations). These data suggest that non-Hox targets of mig-32 and
spat-3 are more likely to be responsible for the defects we describe,
and are consistent with the Polycomb-group targets of Drosophila and
mammalian cells, the vast majority of which are not Hox genes. However, our
data do not rule out a role for MIG-32 or SPAT-3A as regulators of Hox gene
activity, and further work to define the genetic and biochemical properties of
mig-32 and spat-3 in germline and somatic cells will allow
comparison to the Polycomb group complexes of other species.
|
Footnotes |
|---|
|
REFERENCES |
|---|
|
TOP
SUMMARY
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
|---|
Alkema, M. J., Bronk, M., Verhoeven, E., Otte, A., van't Veer,
L. J., Berns, A. and van Lohuizen, M. (1997). Identification
of Bmi1-interacting proteins as constituents of a multimeric mammalian
polycomb complex. Genes Dev.
11,226
-240.
Bateman, A., Birney, E., Cerruti, L., Durbin, R., Etwiller, L.,
Eddy, S. R., Griffiths-Jones, S., Howe, K. L., Marshall, M. and Sonnhammer, E.
L. (2002). The Pfam protein families database.
Nucleic Acids Res. 30,276
-280.
Baum, P. D., Guenther, C., Frank, C. A., Pham, B. V. and
Garriga, G. (1999). The Caenorhabditis elegans gene
ham-2 links Hox patterning to migration of the HSN motor neuron.
Genes Dev. 13,472
-483.
Belote, J. M., Hoffmann, F. M., McKeown, M., Chorsky, R. L. and
Baker, B. S. (1990). Cytogenetic analysis of chromosome
region 73AD of Drosophila melanogaster. Genetics
125,783
-793.[Abstract]
Ben-Saadon, R., Zaaroor, D., Ziv, T. and Ciechanover, A.
(2006). The polycomb protein Ring1B generates self atypical mixed
ubiquitin chains required for its in vitro histone H2A ligase activity.
Mol. Cell 24,701
-711.[CrossRef][Medline]
Bender, L. B., Cao, R., Zhang, Y. and Strome, S.
(2004). The MES-2/MES-3/MES-6 complex and regulation of histone
H3 methylation in C. elegans. Curr. Biol.
14,1639
-1643.[CrossRef][Medline]
Boyer, L. A., Plath, K., Zeitlinger, J., Brambrink, T.,
Medeiros, L. A., Lee, T. I., Levine, S. S., Wernig, M., Tajonar, A., Ray, M.
K. et al. (2006). Polycomb complexes repress developmental
regulators in murine embryonic stem cells. Nature
441,349
-353.[CrossRef][Medline]
Bracken, A. P., Dietrich, N., Pasini, D., Hansen, K. H. and
Helin, K. (2006). Genome-wide mapping of Polycomb target
genes unravels their roles in cell fate transitions. Genes
Dev. 20,1123
-1136.
Brenner, S. (1974). The genetics of
Caenorhabditis elegans. Genetics
77, 71-94.
Cameron, S., Clark, S. G., McDermott, J. B., Aamodt, E. and
Horvitz, H. R. (2002). PAG-3, a Zn-finger transcription
factor, determines neuroblast fate in C. elegans.Development 129,1763
-1774.
Cao, R., Wang, L., Wang, H., Xia, L., Erdjument-Bromage, H.,
Tempst, P., Jones, R. S. and Zhang, Y. (2002). Role of
histone H3 lysine 27 methylation in Polycomb-group silencing.
Science 298,1039
-1043.
Cao, R., Tsukada, Y. and Zhang, Y. (2005). Role
of Bmi-1 and Ring1A in H2A ubiquitylation and Hox gene silencing.
Mol. Cell 20,845
-854.[CrossRef][Medline]
Capowski, E. E., Martin, P., Garvin, C. and Strome, S.
(1991). Identification of grandchildless loci whose products are
required for normal germ-line development in the nematode Caenorhabditis
elegans. Genetics 129,1061
-1072.[Abstract]
Chalfie, M. and Sulston, J. (1981).
Developmental genetics of the mechanosensory neurons of Caenorhabditis
elegans. Dev. Biol. 82,358
-370.[CrossRef][Medline]
Chang, C., Adler, C. E., Krause, M., Clark, S. G., Gertler, F.
B., Tessier-Lavigne, M. and Bargmann, C. I. (2006).
MIG-10/lamellipodin and AGE-1/PI3K promote axon guidance and outgrowth in
response to slit and netrin. Curr. Biol.
16,854
-862.[CrossRef][Medline]
Chisholm, A. (1991). Control of cell fate in
the tail region of C. elegans by the gene egl-5.Development 111,921
-932.
Clark, S. G., Chisholm, A. D. and Horvitz, H. R.
(1993). Control of cell fates in the central body region of
C. elegans by the homeobox gene lin-39. Cell
74, 43-55.[CrossRef][Medline]
Clark, S. G., Lu, X. and Horvitz, H. R. (1994).
The Caenorhabditis elegans locus lin-15, a negative
regulator of a tyrosine kinase signaling pathway, encodes two different
proteins. Genetics 137,987
-997.[Abstract]
Cui, M., Allen, M. A., Larsen, A., Macmorris, M., Han, M. and
Blumenthal, T. (2008). Genes involved in pre-mRNA
3'-end formation and transcription termination revealed by a
lin-15 operon Muv suppressor screen. Proc. Natl. Acad.
Sci. USA 105,16665
-16670.
Czermin, B., Melfi, R., McCabe, D., Seitz, V., Imhof, A. and
Pirrotta, V. (2002). Drosophila enhancer of Zeste/ESC
complexes have a histone H3 methyltransferase activity that marks chromosomal
Polycomb sites. Cell
111,185
-196.[CrossRef][Medline]
Dalpe, G., Zhang, L. W., Zheng, H. and Culotti, J. G.
(2004). Conversion of cell movement responses to Semaphorin-1 and
Plexin-1 from attraction to repulsion by lowered levels of specific RAC
GTPases in C. elegans. Development
131,2073
-2088.
Desai, C., Garriga, G., McIntire, S. L. and Horvitz, H. R.
(1988). A genetic pathway for the development of the
Caenorhabditis elegans HSN motor neurons.
Nature 336,638
-646.[CrossRef][Medline]
Emmons, S. W. (2005). Male development. In
WormBook (ed. The C. elegans Research
Community).
www.wormbook.org.
Fire, A., Xu, S., Montgomery, M. K., Kostas, S. A., Driver, S.
E. and Mello, C. C. (1998). Potent and specific genetic
interference by double-stranded RNA in Caenorhabditis elegans.Nature 391,806
-811.[CrossRef][Medline]
Fong, Y., Bender, L., Wang, W. and Strome, S.
(2002). Regulation of the different chromatin states of autosomes
and X chromosomes in the germ line of C. elegans.Science 296,2235
-2238.
Francis, N. J., Kingston, R. E. and Woodcock, C. L.
(2004). Chromatin compaction by a polycomb group protein complex.
Science 306,1574
-1577.
Fujii, T., Nakao, F., Shibata, Y., Shioi, G., Kodama, E.,
Fujisawa, H. and Takagi, S. (2002). Caenorhabditis
elegans PlexinA, PLX-1, interacts with transmembrane semaphorins and
regulates epidermal morphogenesis. Development
129,2053
-2063.
Ginzburg, V. E., Roy, P. J. and Culotti, J. G.
(2002). Semaphorin 1a and semaphorin 1b are required for correct
epidermal cell positioning and adhesion during morphogenesis in C.
elegans. Development 129,2065
-2078.
Gunster, M. J., Satijn, D. P., Hamer, K. M., den Blaauwen, J.
L., de Bruijn, D., Alkema, M. J., van Lohuizen, M., van Driel, R. and Otte, A.
P. (1997). Identification and characterization of
interactions between the vertebrate polycomb-group protein BMI1 and human
homologs of polyhomeotic. Mol. Cell. Biol.
17,2326
-2335.[Abstract]
Hamelin, M., Scott, I. M., Way, J. C. and Culotti, J. G.
(1992). The mec-7 beta-tubulin gene of
Caenorhabditis elegans is expressed primarily in the touch receptor
neurons. EMBO J. 11,2885
-2893.[Medline]
Hao, J. C., Yu, T. W., Fujisawa, K., Culotti, J. G.,
Gengyo-Ando, K., Mitani, S., Moulder, G., Barstead, R., Tessier-Lavigne, M.
and Bargmann, C. I. (2001). C. elegans slit acts in
midline, dorsal-ventral, and anterior-posterior guidance via the SAX-3/Robo
receptor. Neuron 32,25
-38.[CrossRef][Medline]
Harris, J., Honigberg, L., Robinson, N. and Kenyon, C.
(1996). Neuronal cell migration in C. elegans:
regulation of Hox gene expression and cell position.
Development 122,3117
-3131.[Abstract]
Haupt, Y., Alexander, W. S., Barri, G., Klinken, S. P. and
Adams, J. M. (1991). Novel zinc finger gene implicated as myc
collaborator by retrovirally accelerated lymphomagenesis in E mu-myc
transgenic mice. Cell
65,753
-763.[CrossRef][Medline]
Holdeman, R., Nehrt, S. and Strome, S. (1998).
MES-2, a maternal protein essential for viability of the germline in
Caenorhabditis elegans, is homologous to a Drosophila Polycomb group
protein. Development
125,2457
-2467.[Abstract]
Hunter, C. P., Harris, J. M., Maloof, J. N. and Kenyon, C.
(1999). Hox gene expression in a single Caenorhabditis
elegans cell is regulated by a caudal homolog and intercellular signals
that inhibit wnt signaling. Development
126,805
-814.[Abstract]
Irminger-Finger, I. and Nothiger, R. (1995).
The Drosophila melanogaster gene lethal(3)73Ah encodes a
ring finger protein homologous to the oncoproteins MEL-18 and BMI-1.
Gene 163,203
-208.[CrossRef][Medline]
Jackson, J. P., Johnson, L., Jasencakova, Z., Zhang, X.,
PerezBurgos, L., Singh, P. B., Cheng, X., Schubert, I., Jenuwein, T. and
Jacobsen, S. E. (2004). Dimethylation of histone H3 lysine 9
is a critical mark for DNA methylation and gene silencing in Arabidopsis
thaliana. Chromosoma
112,308
-315.[CrossRef][Medline]
Jacobs, J. J., Kieboom, K., Marino, S., DePinho, R. A. and van
Lohuizen, M. (1999). The oncogene and Polycomb-group gene
bmi-1 regulates cell proliferation and senescence through the
ink4a locus. Nature
397,164
-168.[CrossRef][Medline]
Kennison, J. A. and Tamkun, J. W. (1988).
Dosage-dependent modifiers of polycomb and antennapedia mutations in
Drosophila. Proc. Natl. Acad. Sci. USA
85,8136
-8140.
Kenyon, C. (1986). A gene involved in the
development of the posterior body region of C. elegans.Cell 46,477
-487.[CrossRef][Medline]
Kim, M. H., Gunnersen, J. M. and Tan, S. S.
(2005). Mph2 expression in germinal zones of the mouse
brain. Dev. Dyn. 232,209
-215.[CrossRef][Medline]
King, I. F., Francis, N. J. and Kingston, R. E.
(2002). Native and recombinant polycomb group complexes establish
a selective block to template accessibility to repress transcription in
vitro. Mol. Cell. Biol. 22,7919
-7928.
Kuzmichev, A., Nishioka, K., Erdjument-Bromage, H., Tempst, P.
and Reinberg, D. (2002). Histone methyltransferase activity
associated with a human multiprotein complex containing the Enhancer of Zeste
protein. Genes Dev. 16,2893
-2905.
Labbe, J. C., Pacquelet, A., Marty, T. and Gotta, M.
(2006). A genomewide screen for suppressors of par-2
uncovers potential regulators of PAR protein-dependent cell polarity in
Caenorhabditis elegans. Genetics
174,285
-295.
Lee, T. I., Jenner, R. G., Boyer, L. A., Guenther, M. G.,
Levine, S. S., Kumar, R. M., Chevalier, B., Johnstone, S. E., Cole, M. F.,
Isono, K. et al. (2006). Control of developmental regulators
by Polycomb in human embryonic stem cells. Cell
125,301
-313.[CrossRef][Medline]
Leung, C., Lingbeek, M., Shakhova, O., Liu, J., Tanger, E.,
Saremaslani, P., Van Lohuizen, M. and Marino, S. (2004).
Bmi1 is essential for cerebellar development and is overexpressed in
human medulloblastomas. Nature
428,337
-341.[CrossRef][Medline]
Levine, S. S., Weiss, A., Erdjument-Bromage, H., Shao, Z.,
Tempst, P. and Kingston, R. E. (2002). The core of the
polycomb repressive complex is compositionally and functionally conserved in
flies and humans. Mol. Cell. Biol.
22,6070
-6078.
Lewis, E. B. (1978). A gene complex controlling
segmentation in Drosophila. Nature
276,565
-570.[CrossRef][Medline]
Li, C., Nelson, L. S., Kim, K., Nathoo, A. and Hart, A. C.
(1999). Neuropeptide gene families in the nematode
Caenorhabditis elegans. Ann. NY Acad. Sci.
897,239
-252.[CrossRef][Medline]
Li, Z., Cao, R., Wang, M., Myers, M. P., Zhang, Y. and Xu, R.
M. (2006). Structure of a Bmi-1-Ring1B polycomb group
ubiquitin ligase complex. J. Biol. Chem.
281,20643
-20649.
McIntire, S. L., Reimer, R. J., Schuske, K., Edwards, R. H. and
Jorgensen, E. M. (1997). Identification and characterization
of the vesicular GABA transporter. Nature
389,870
-876.[CrossRef][Medline]
Molofsky, A. V., He, S., Bydon, M., Morrison, S. J. and Pardal,
R. (2005). Bmi-1 promotes neural stem cell self-renewal and
neural development but not mouse growth and survival by repressing the
p16Ink4a and p19Arf senescence pathways. Genes Dev.
19,1432
-1437.
Molofsky, A. V., Pardal, R., Iwashita, T., Park, I. K., Clarke,
M. F. and Morrison, S. J. (2003). Bmi-1 dependence
distinguishes neural stem cell self-renewal from progenitor proliferation.
Nature 425,962
-967.[CrossRef][Medline]
Muller, J., Hart, C. M., Francis, N. J., Vargas, M. L.,
Sengupta, A., Wild, B., Miller, E. L., O'Connor, M. B., Kingston, R. E. and
Simon, J. A. (2002). Histone methyltransferase activity of a
Drosophila Polycomb group repressor complex. Cell
111,197
-208.[CrossRef][Medline]
Negre, N., Hennetin, J., Sun, L. V., Lavrov, S., Bellis, M.,
White, K. P. and Cavalli, G. (2006). Chromosomal distribution
of PcG proteins during Drosophila development. PLoS
Biol. 4,e170
.[CrossRef][Medline]
Nunes, M., Blanc, I., Maes, J., Fellous, M., Robert, B. and
McElreavey, K. (2001). NSPc1, a novel mammalian
Polycomb gene, is expressed in neural crest-derived structures of the
peripheral nervous system. Mech. Dev.
102,219
-222.[CrossRef][Medline]
Nusslein-Volhard, C., Kluding, H. and Jurgens, G.
(1985). Genes affecting the segmental subdivision of the
Drosophila embryo. Cold Spring Harb. Symp. Quant.
Biol. 50,145
-154.
Osley, M. A. (2006). Regulation of histone H2A
and H2B ubiquitylation. Brief. Funct. Genomic.
Proteomic. 5,179
-189.
Pan, C. L., Howell, J. E., Clark, S. G., Hilliard, M., Cordes,
S., Bargmann, C. I. and Garriga, G. (2006). Multiple Wnts and
frizzled receptors regulate anteriorly directed cell and growth cone
migrations in Caenorhabditis elegans. Dev. Cell
10,367
-377.[CrossRef][Medline]
Pujol, N., Torregrossa, P., Ewbank, J. J. and Brunet, J. F.
(2000). The homeodomain protein CePHOX2/CEH-17 controls
antero-posterior axonal growth in C. elegans.Development 127,3361
-3371.[Abstract]
Quinn, C. C., Pfeil, D. S., Chen, E., Stovall, E. L., Harden, M.
V., Gavin, M. K., Forrester, W. C., Ryder, E. F., Soto, M. C. and Wadsworth,
W. G. (2006). UNC-6/netrin and SLT-1/slit guidance cues
orient axon outgrowth mediated by MIG-10/RIAM/lamellipodin. Curr.
Biol. 16,845
-853.[CrossRef][Medline]
Ross, J. M. and Zarkower, D. (2003). Polycomb
group regulation of Hox gene expression in C. elegans. Dev.
Cell 4,891
-901.[CrossRef][Medline]
Salser, S. J. and Kenyon, C. (1992). Activation
of a C. elegans Antennapedia homologue in migrating cells controls
their direction of migration. Nature
355,255
-258.[CrossRef][Medline]
Sanchez, C., Sanchez, I., Demmers, J. A., Rodriguez, P.,
Strouboulis, J. and Vidal, M. (2007). Proteomics analysis of
Ring1B/Rnf2 interactors identifies a novel complex with the Fbxl10/Jhdm1B
histone demethylase and the Bcl6 interacting corepressor. Mol. Cell
Proteomics 6,820
-834.
Saurin, A. J., Shiels, C., Williamson, J., Satijn, D. P., Otte,
A. P., Sheer, D. and Freemont, P. S. (1998). The human
polycomb group complex associates with pericentromeric heterochromatin to form
a novel nuclear domain. J. Cell Biol.
142,887
-898.
Saurin, A. J., Shao, Z., Erdjument-Bromage, H., Tempst, P. and
Kingston, R. E. (2001). A Drosophila Polycomb group complex
includes Zeste and dTAFII proteins. Nature
412,655
-660.[CrossRef][Medline]
Schoorlemmer, J., Marcos-Gutierrez, C., Were, F., Martinez, R.,
Garcia, E., Satijn, D. P., Otte, A. P. and Vidal, M. (1997).
Ring1A is a transcriptional repressor that interacts with the Polycomb-M33
protein and is expressed at rhombomere boundaries in the mouse hindbrain.
EMBO J. 16,5930
-5942.[CrossRef][Medline]
Schwartz, Y. B., Kahn, T. G., Nix, D. A., Li, X. Y., Bourgon,
R., Biggin, M. and Pirrotta, V. (2006). Genome-wide analysis
of Polycomb targets in Drosophila melanogaster. Nat.
Genet. 38,700
-705.[CrossRef][Medline]
Shakhova, O., Leung, C. and Marino, S. (2005).
Bmi1 in development and tumorigenesis of the central nervous system.
J. Mol. Med. 83,596
-600.[CrossRef][Medline]
Shao, Z., Raible, F., Mollaaghababa, R., Guyon, J. R., Wu, C.
T., Bender, W. and Kingston, R. E. (1999). Stabilization of
chromatin structure by PRC1, a Polycomb complex. Cell
98, 37-46.[CrossRef][Medline]
Simon, J., Chiang, A. and Bender, W. (1992).
Ten different Polycomb group genes are required for spatial control of the
abdA and AbdB homeotic products. Development
114,493
-505.[Abstract]
Spieth, J., Brooke, G., Kuersten, S., Lea, K. and Blumenthal,
T. (1993). Operons in C. elegans: polycistronic mRNA
precursors are processed by trans-splicing of SL2 to downstream
coding regions. Cell 73,521
-532.[CrossRef][Medline]
Sulston, J. E., Schierenberg, E., White, J. G. and Thomson, J.
N. (1983). The embryonic cell lineage of the nematode
Caenorhabditis elegans. Dev. Biol.
100,64
-119.[CrossRef][Medline]
Tolhuis, B., de Wit, E., Muijrers, I., Teunissen, H., Talhout,
W., van Steensel, B. and van Lohuizen, M. (2006). Genome-wide
profiling of PRC1 and PRC2 Polycomb chromatin binding in Drosophila
melanogaster. Nat. Genet.
38,694
-699.[CrossRef][Medline]
Trent, C., Tsung, N. and Horvitz, H. R. (1983).
Egg-laying defective mutants of the nematode Caenorhabditis elegans.Genetics 104,619
-647.
Trimarchi, J. M., Fairchild, B., Wen, J. and Lees, J. A.
(2001). The E2F6 transcription factor is a component of the
mammalian Bmi1-containing polycomb complex. Proc. Natl. Acad. Sci.
USA 98,1519
-1524.
Troemel, E. R., Chou, J. H., Dwyer, N. D., Colbert, H. A. and
Bargmann, C. I. (1995). Divergent seven transmembrane
receptors are candidate chemosensory receptors in C. elegans.Cell 83,207
-218.[CrossRef][Medline]
van der Lugt, N. M., Domen, J., Linders, K., van Roon, M.,
Robanus-Maandag, E., te Riele, H., van der Valk, M., Deschamps, J., Sofroniew,
M., van Lohuizen, M. et al. (1994). Posterior transformation,
neurological abnormalities, and severe hematopoietic defects in mice with a
targeted deletion of the bmi-1 proto-oncogene. Genes
Dev. 8,757
-769.
van Lohuizen, M., Verbeek, S., Scheijen, B., Wientjens, E., van
der Gulden, H. and Berns, A. (1991). Identification of
cooperating oncogenes in E mu-myc transgenic mice by provirus
tagging. Cell 65,737
-752.[CrossRef][Medline]
Wang, B. B., Muller-Immergluck, M. M., Austin, J., Robinson, N.
T., Chisholm, A. and Kenyon, C. (1993). A homeotic gene
cluster patterns the anteroposterior body axis of C. elegans.Cell 74,29
-42.[CrossRef][Medline]
Wang, H., Wang, L., Erdjument-Bromage, H., Vidal, M., Tempst,
P., Jones, R. S. and Zhang, Y. (2004a). Role of histone H2A
ubiquitination in Polycomb silencing. Nature
431,873
-878.[CrossRef][Medline]
Wang, L., Brown, J. L., Cao, R., Zhang, Y., Kassis, J. A. and
Jones, R. S. (2004b). Hierarchical recruitment of polycomb
group silencing complexes. Mol. Cell
14,637
-646.[CrossRef][Medline]
Wei, J., Zhai, L., Xu, J. and Wang, H. (2006).
Role of Bmi1 in H2A ubiquitylation and Hox gene silencing. J. Biol.
Chem. 281,22537
-22544.
White, J. G., Southgate, E., Thomson, J. N. and Brenner, S.
(1986). The structure of the nervous system of the nematode
Caenorhabditis elegans. Philos. Trans. R. Soc. Lond. B Biol.
Sci. 311,1
-340.
Xu, L., Fong, Y. and Strome, S. (2001a). The
Caenorhabditis elegans maternal-effect sterile proteins, MES-2,
MES-3, and MES-6, are associated in a complex in embryos. Proc.
Natl. Acad. Sci. USA 98,5061
-5066.
Xu, L., Paulsen, J., Yoo, Y., Goodwin, E. B. and Strome, S.
(2001b). Caenorhabditis elegans MES-3 is a target of
GLD-1 and functions epigenetically in germline development.
Genetics 159,1007
-1017.
Yang, Y., Sun, Y., Luo, X., Zhang, Y., Chen, Y., Tian, E.,
Lints, R. and Zhang, H. (2007). Polycomb-like genes are
necessary for specification of dopaminergic and serotonergic neurons in
Caenorhabditis elegans. Proc. Natl. Acad. Sci. USA
104,852
-857.
Yu, H., Pretot, R. F., Burglin, T. R. and Sternberg, P. W.
(2003). Distinct roles of transcription factors EGL-46 and DAF-19
in specifying the functionality of a polycystin-expressing sensory neuron
necessary for C. elegans male vulva location behavior.
Development 130,5217
-5227.
Zallen, J. A., Yi, B. A. and Bargmann, C. I.
(1998). The conserved immunoglobulin superfamily member
SAX-3/Robo directs multiple aspects of axon guidance in C. elegans.Cell 92,217
-227.[CrossRef][Medline]
Zhang, H. and Emmons, S. W. (2001). The novel
C. elegans gene sop-3 modulates Wnt signaling to regulate
Hox gene expression. Development
128,767
-777.[Abstract]
Zhang, H., Azevedo, R. B., Lints, R., Doyle, C., Teng, Y.,
Haber, D. and Emmons, S. W. (2003). Global regulation of Hox
gene expression in C. elegans by a SAM domain protein.
Dev. Cell 4,903
-915.[CrossRef][Medline]
Zhang, H., Smolen, G. A., Palmer, R., Christoforou, A., van den
Heuvel, S. and Haber, D. A. (2004). SUMO modification is
required for in vivo Hox gene regulation by the Caenorhabditis
elegans Polycomb group protein SOP-2. Nat. Genet.
36,507
-511.[CrossRef][Medline]
Zhang, T., Sun, Y., Tian, E., Deng, H., Zhang, Y., Luo, X., Cai,
Q., Wang, H., Chai, J. and Zhang, H. (2006). RNA-binding
proteins SOP-2 and SOR-1 form a novel PcG-like complex in C. elegans.Development 133,1023
-1033.
Zhu, P., Zhou, W., Wang, J., Puc, J., Ohgi, K. A.,
Erdjument-Bromage, H., Tempst, P., Glass, C. K. and Rosenfeld, M. G.
(2007). A histone H2A deubiquitinase complex coordinating histone
acetylation and H1 dissociation in transcriptional regulation. Mol.
Cell 27,609
-621.[CrossRef][Medline]
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati 
Twitter What's this?
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
