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First published online 16 August 2006
doi: 10.1242/dev.02534
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1 Department of Biochemistry, College of Science, Yonsei University, Seoul
120-749, Korea.
2 School of Life Sciences and Biotechnology, Korea University, Seoul 136-701,
Korea.
3 Institute of Molecular Biology, University of Zurich, 8057 Zurich,
Switzerland.
Author for correspondence (e-mail:
kooh{at}yonsei.ac.kr)
Accepted 13 July 2006
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SUMMARY |
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 TOP SUMMARY INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Key words: DICE1 (INTS6), Apoptosis, Mitochondria, Cristae remodeling, C. elegans
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INTRODUCTION |
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TOPSUMMARYINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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).
Although DICE1 mRNA was detected in all human tissues, it was
downregulated or absent in cell lines and tissues of non-small cell lung
carcinoma. Hypermethylation of its promoter region was associated with its
downregulation in several cell lines derived from non-small cell carcinomas
(Wieland et al., 2001) and
prostate cancers (Röpke et al.,
2005). Ectopic expression of DICE1 inhibited colony formation by
cell lines of both non-small cell lung cancer and prostate cancer, and
suppressed anchorage-independent growth of mouse fibroblasts transformed with
the IGF1 receptor gene (Wieland et al.,
2004). Overexpression of a mouse DICE1 homolog (DBI1) also
suppressed induction of cell proliferation by IGF1
(Hoff et al., 1998). DBI1 was
located in the nuclear fraction (Hoff et
al., 1998), in agreement with the nuclear localization of
EGFP-tagged human DICE1 in COS cells
(Wieland et al., 2001).
Although DICE1 homologs in humans and mice are thought to be suppressors of
cell proliferation, the mode of action of the proteins is largely unknown,
because they contain only one functional domain, as defined by current
prediction algorithms: the N terminus of each DICE1 homolog has a conserved
amino acid sequence encoding a von Willebrand factor type A domain (VWF A)
(Fig. 1). This domain is known
to be involved in cell-cell, cell-extracellular matrix and intracellular
protein-protein interactions, as observed in more than 20 human proteins,
including the von Willebrand blood clotting factor, some types of collagens,
integrin subunits and leukocyte adhesion receptor (for reviews, see
Colombatti and Bonaldo, 1991;
Whittaker and Hynes, 2002).
Although the presence of a DEAD-box in human DICE1 suggests a helicase
function, no DEAD box is present in the C. elegans homolog, and a
more extensive helicase motif is not found in any DICE1 homologs. The
likelihood that DICE1 acts as a helicase is thus very low, and the conserved
VWF A domain does not suggest any other potential role.
In this study, we set out to analyze the knockdown and knockout phenotypes of a C. elegans DICE1 homolog in order to elucidate the molecular function of human DICE1. The protein is essential for C. elegans oogenesis, late embryogenesis and larval growth, and its deficiency induces apoptosis. It is localized to the mitochondrial inner membrane, and abnormalities in both overall and inner mitochondrial morphologies result from its absence. The localization of DICE1 and the phenotypes caused by its deficiency suggest that it is involved in remodeling of the mitochondrial cristae in C. elegans. C. elegans DICE1 shows differences in its subcellular location and function from the human homolog, and our results imply either that they have distinct functions due to divergent evolution, or that the proteins have additional cryptic roles.
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MATERIALS AND METHODS |
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TOPSUMMARYINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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). The yk293c3 EST clone of dic-1 (F08B4.1) was
digested with HindIII enzyme, and the resulting 1.6 kb cDNA fragment
was cloned into vector pPD129.36(L4440), which contains convergent T7
polymerase promoters separated by a multicloning site. The resulting construct
was transformed into E. coli HT115(DE3)
(Timmons and Fire, 1998).
Transformants were grown in LB medium containing 100 µg/ml of ampicillin
for 18 hours and seeded onto NGM plates supplemented with 100 µg/ml of
ampicillin and 1 mM isopropyl-thio-ß-D-galactoside (IPTG). To examine
gonads and mitochondria of adult worms, RNAi was performed from the L1 larval
stage. To observe the embryonic phenotypes induced by knockdown of
dic-1, L4 stage worms were grown on NGM plates covered with E.
coli cells producing dsRNA of dic-1 for 24 hours. Adult worms
were transferred to new NGM plates, allowed to lay eggs for 6 hours, and then
removed. Embryo hatching was scored 24 hour later.
Outcross of the dic-1(tm1615) mutant and rescue of larval arrest by wild type dic-1
The dic-1(tm1615) heterozygous strain from the National
Bioresource Project was outcrossed with N2 strains four times to remove
possible unrelated mutations. The mutant harbors a deletion of 588 bp,
corresponding to nucleotides 32905 to 33492 in the cosmid clone F08B4
(Fig. 7). Primers used for PCR
reactions to probe the deletion were as follows: outer sense,
5'-GGTATTTCGTATTCCATCCACTCC; outer antisense,
5'-CCTTGAAATCTCCTGCATAAATAGG; inner sense,
5'-ACGAAGCTTCTCAATTGTATCACC; and inner antisense,
5'-CCTCAATATCAACTTCACTGTTTCC. To observe the phenotypes of
dic-1(tm1615) homozygotes, worms were mounted on slides overlaid with
2% agar and observed with Nomarski optics (DMR HC, Leica). C. elegans
dic-1(tm1615) IV/nT1[qIs51] (IV;V) heterozygotes were generated by
crossing dic-1(tm1615) heterozygotes with nT1[qIs51]/+ males
that were the progeny of a cross between elo-5(ok683)/nT1[qIs51]
hermaphrodites and N2 males
To test for rescue of the homozygotes by the wild-type gene, a DNA fragment
containing wild-type dic-1 gene from 1.9 kb upstream of the start
codon to 1.3 kb downstream of the stop codon was amplified from the F08B4
cosmid genomic DNA clone using the Expand Long Template PCR system (Roche).
The amplified DNA fragment was cloned into pCR2.1-TOPO vector (Invitrogen) and
the recombinant plasmid DNA (2 µg/ml) was microinjected into the gonads of
young adult C. elegans dic-1(tm1615)/nT1[qIs51] heterozygotes,
together with pRF4 plasmid DNA (100 µg/ml)
(Krammer et al., 1990).
Heterozygous transgenic worms in the F1 progeny were selected and maintained
by observing their rolling (rol-6) and gfp-positive
phenotypes. Rescue of homozygotes by the wild-type dic-1 gene was
judged by scoring gfp-negative rollers reaching the adult stage.
To test whether human DICE1 can replace C. elegans DIC-1, a 2.7 kb coding region of human DICE1 gene was amplified from cDNA clone MGC:48751 (Invitrogen) as described above. The amplified cDNA fragment was cloned into pPD49.78 plasmid with an hsp-16.2 promoter, followed by microinjection into the gonads of dic-1(tm1615)/nT1[qIs51] heterozygotes together with pRF4 plasmid DNA. Rolling heterozygotes were allowed to lay embryos and the progeny were treated at the L1 stage by heat shock at 30°C for 1 hour to induce expression of the human DICE1 in their extrachromosomal arrays. The rescue of homozygotes by human DICE1 was judged as above, by scoring gfp-negative rollers reaching the adult stage. As a positive control for the rescue, the coding region of C. elegans DIC-1 was amplified from the yk293c3 cDNA clone and cloned into pPD49.78. A transgenic line was obtained and the rescue of larval arrest in homozygotes was confirmed after heat shock induction of the dic-1 cDNA expression.
GFP-tagged DIC-1 expression in C. elegans
To visualize the subcellular location of C. elegans DIC-1, the
protein was fused with GFP and expressed under the control of either the
hsp-16.2 or myo-3 promoter. The 2.6 kb cDNA fragment of
dic-1 was cloned into pPD118.26 or pPD114.98, and transgenic lines
were generated by microinjecting recombinant plasmid DNA (20 µg/ml)
together with pRF4 plasmid DNA (50 µg/ml) into adult N2 worms. Adult worms
of the transgenic line Phsp-16.2::dic-1::gfp were heat
shocked at 30°C for 1 hour and GFP expression was observed in the progeny
embryos 3 hours later.
Counts of apoptotic cells
Cell corpses in the germ line were counted by mounting worms in a drop of
M9 solution and observing the gonads with Nomarski optics (DMR HC, Leica).
Because corpses are cellularized and more refractive than normal cells, they
can be readily identified under high magnification at the late pachytene
stage. We observed cell corpses in embryos after mounting 380 cell or bean
cell stage embryos on slides coated with 0.1% poly-L-lysine. We also performed
TUNEL staining to detect apoptotic cells in the embryos with an In Situ Cell
Death Detection Kit (Roche) according to the manufacturer's instructions.
Embryos were freeze-cracked, fixed and transferred to the wells of a
polylysine-coated slide. TUNEL reaction mixture (10 µl of enzyme solution
and 90 µl of label solution per well) was added to the fixed embryos and
they were allowed to incubate in the dark for 2 hours at 37°C in a
humidified atmosphere. After washing with PBS, the specimens were stained with
DAPI (4,6-diamidino-2-phenylindole, 1 µg/ml) for 10 minutes and observed
with a fluorescence microscope.
Visualizing mitochondria with mitotracker and using a C. elegans strain expressing GFP in mitochondria
To stain mitochondria in adult worms, L4 stage worms were transferred to
NGM plates containing Mitotracker Red (Molecular Probes, 2 µg/ml) and
incubated in the dark at 20°C for 12 hours. They were then transferred to
new NGM plates to reduce nonspecific staining. After 10 to 20 minutes, they
were mounted on a slide overlaid with 2% agar and observed with a fluorescence
microscope. The Pmyo-3::mitochondrial signal sequence::gfp
transgenic line, in which muscle cell mitochondria are marked by GFP, was used
to observe changes in mitochondrial morphology after dic-1
RNAi.
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).
Embryos were freeze-cracked, fixed, incubated with polyclonal mouse antiserum
against DIC-1 (1:50 dilution), followed by FITC-conjugated goat anti-mouse
immunoglobulin G (1:1000 dilution, Molecular Probes). After staining with DAPI
(1 µg/ml), the specimens were observed with a fluorescence microscope. For
staining, gonads were extruded from decapitated adult worms, fixed in 3%
paraformaldehyde and 0.1 M K2HPO4 (pH 7.2) for 1 hour,
post-fixed with cold methanol, and incubated with primary and secondary
antibodies as described by Jones et al.
(Jones et al., 1996). To
double-stain DIC-1 and cytochrome c oxidase, gonads were incubated
successively with polyclonal mouse antiserum against DIC-1, FITC-conjugated
goat anti mouse immunoglobulin G, and Alex Fluor 594-conjugated mouse
immunoglobulin G against bovine cytochrome c oxidase subunit 1 (Molecular
Probes).
Cryo-electron microscopy
C. elegans worms were plunged into liquid propane at 180°C.
The frozen samples were then transferred to substitution medium (acetone) and
freeze-substituted at 80°C for 72 hours, after which the temperature was
raised to 40°C at a rate of 6°C hr-1. The samples were
washed three times in precooled pure acetone over a period of 1 hour, and then
infiltrated with a spur resin (EMS). Infiltration was performed with
acetone/resin mixtures (v/v) of 3:1, 1:1 and 1:3 for 1 hour each and with the
pure resin for 3 hours. The resin was polymerized at 60°C for 48 hours.
Sections about 60 nm thick were cut horizontal to the plane of the sample
using an ultra-microtome (RMC MTXL) and mounted on nickel grids. They were
stained with uranyl acetate and lead citrate and viewed at 120 kV with a
Technai 12 electron microscope (Philips, Netherlands).
For immunogold labeling, the ultra-thin sections on nickel grids were etched with 10% hydrogen peroxide for 30 minutes, rinsed in deionized water and floated on 0.56 mM sodium metaperiodate. They were then rinsed with deionized water and floated for 1 hour in PBS containing 1% bovine serum albumin (PBS-BSA), followed by overnight incubation at 4°C in antimouse DIC-1 diluted to 1:200 with PBS-BSA. After rinsing several times with PBS-BSA, they were floated for 1 hour in anti-mouse IgG conjugated to 10 nm gold particles (British Biocell International) diluted to 1:50 with PBS-BSA. The gold-labeled sections were rinsed successively with PBS, PBST (containing 1% Triton X-100) and deionized water, air dried, stained and examined as above by electron microscopy.
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RESULTS |
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TOPSUMMARYINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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).
Fig. 2C illustrates
tissue-specific gene expression in wild-type (threefold) and
dic-1(RNAi) embryos at the same time post fertilization,
showing similar overall intensity of GFP fluorescence. Although the
dic-1(RNAi) embryos did not pass the comma stage
morphologically, the extent of GFP expression suggests that tissue-specific
differentiation progressed further; however, the differentiated cells were
disorganized, in agreement with the morphogenic abnormalities seen in
Fig. 2A.
ced-3-dependent apoptosis is induced in dic-1(RNAi) embryos and germ line
Although cell cavity formation in dic-1(RNAi) embryos did
not depend on the major apoptosis pathway
(Fig. 2B), we tested whether
DIC-1 deficiency led to an increased frequency of apoptosis. As
dic-1(RNAi) embryos arrested before the comma stage,
apoptotic cells were scored at the 380 cell and bean cell stages either by
direct observation of cell corpses under Nomarski optics
(Fig. 3A) or after TUNEL
staining (Fig. 3B). By both
measures, ced-3(n717)-dependent apoptosis increased about twofold at
the 380-cell stage and 1.6-fold at the comma stage because of knockdown of
dic-1. Increased apoptosis was also observed in adult gonads upon
RNAi of dic-1 from the L4 stage
(Fig. 4A). This germline
apoptosis was abrogated by the ced-3(n717) mutation, as in embryos,
and also by the ced-4(n1162) mutation, indicating that it was induced
via the major apoptosis pathway. It was not affected in the
cep-1(gk138) background, which rules out DNA damage as the main cause
of the apoptosis.
dic-1(RNAi) blocks oogenesis and generates cell cavities in the germ line
When RNAi of dic-1 was performed from the L1 stage,
various phenotypes appeared, including undersized gonads, inhibition of
oogenesis and cavities in the gonad (Fig.
4B). No apparent defects in germline development were observed
under Nomarski optics from the mitotic to the pachytene region, apart from
some reduction in the number of germ cells. Nevertheless, slightly disturbed
immunolocalization was observed in dic-1(RNAi) gonads for
syntaxin 4 in the cytoplasmic membrane, and for porin in the nuclear membrane
(S.M.H., unpublished). Although meiotic prophase progression was detected by
immunostaining of GLD-1 at the pachytene stage, synapsed chromosomal strands
were not as clear in the dic-1(RNAi) gonad after DAPI
staining as in the wild type (S.M.H., unpublished). As in embryos, gonadal
DIC-1 deficiency had a marked effect on actively differentiating cells such as
late pachytene stage cells and oocytes, but not on rapidly dividing cells. We
observed GFP at the rim of the cavities in a transgenic derivative of
dic-1(RNAi) strain expressing annexin V::GFP, which binds to
phosphatidylserine in the outer layer of the cytoplasmic membrane of apoptotic
and necrotic cells (S. Züllig and T.H.L., unpublished). This suggests
that the cavities resulted from apoptotic or necrotic cell death and were not
merely vacuoles.
DIC-1 protein is localized to the mitochondrial inner membrane
When C. elegans embryos were reacted with a polyclonal mouse
antibody raised against DIC-1, the protein appeared as speckles in the
cytoplasm throughout embryogenesis (Fig.
5A). It was present in germ cells and oocytes, and its particulate
distribution in the cytoplasm was clearly observed
(Fig. 5B,C). At the
post-embryonic stage, DIC-1 was present in all somatic cells, although at a
lower expression level than in germline cells (S.M.H., unpublished). The
speckles of DIC-1 in mitotic germ cells co-localized with anti-cytochrome c
oxidase subunit 1 antibody conjugated to Alexa Fluor 594
(Fig. 5C), supporting a
mitochondrial location of DIC-1. The mitochondrial localization was confirmed
by expressing GFP-tagged DIC-1 under the control of the hsp16.2 or
myo-3 promoter. The fusion protein localized in the cytoplasm of
embryonic cells as foci, as well as in the mitochondria of muscle cells (see
Fig. 6), like the GFP fused to
a mitochondrial localization signal sequence in
Fig. 7B. In agreement with
these results, DIC-1 proved to be located in mitochondria by cryo-electron
microscopy, more precisely on the inner membrane at cristae
(Fig. 5D). The TMpred program
(http://www.ch.embnet.org/software/TMPRED_form.html)
predicts the presence of two transmembrane regions in DIC-1, encompassing
amino acids 129-148 and 377-397, thus supporting the localization in the
membrane.
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L3 growth arrest has been observed in mutants defective in mitochondrial
proteins (Tsang et al., 2001)
and after ethidium bromide treatment, which inhibits mitochondrial DNA
replication (Tsang and Lemire,
2002). This L3 arrest is related to the energy requirement for
entry into the L4 stage, and the fivefold increase in the number of
mitochondrial DNA copies at this time
(Tsang and Lemire, 2002). The
L3 arrest of dic-1(tm1615) homozygotes is yet further evidence that
DIC-1 is crucial for mitochondrial activity. When RNAi of
dic-1 was performed from the L1 stage, a minor fraction of the worms
arrested at the L4 or early adult stage, not at the L3 stage. The
incompleteness of RNAi and the later disappearance of zygotic dic-1
expression may have contributed to the much weaker post-embryonic somatic
phenotype of the knockdown strain compared with the knockout mutant.
Expression of human DICE1 does not rescue the larval arrest of dic-1 homozygotes
In order to determine whether human DICE1 is a functional homolog of C.
elegans DIC-1, we generated a dic-1(tm1615)/nT1[qIs51]
heterozygous strains containing extrachromosomal arrays of human
DICE1 cDNA (or C. elegans dic-1 cDNA as a positive control)
linked to the hsp-16.2 promoter and the rol-6 gene. When
heterozygotes transformed with C. elegans dic-1 cDNA were allowed to
produce progeny, followed by heat shock at the L1 stage, 11% of the roller
progeny (n=682) that reached adulthood were dic-1
homozygotes. This showed that heat shock-induced dic-1 cDNA
expression rescued the mutant phenotype as effectively as the wild-type
dic-1 gene expression described above. By contrast, none of the adult
roller progeny (n=203) was dic-1 homozygotes when the roller
progeny were from heterozygotes transformed with human DICE1 cDNA.
Thus, we were unable to obtain evidence that human DICE1 and C.
elegans DIC-1 are functionally equivalent. Whether this failure reflects
technical limitations with the expression or stability of the human protein in
worms, the inability of human DICE1 to interact with essential C.
elegans partners due to divergence, or a difference in function of the
two proteins is not known.
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DISCUSSION |
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TOPSUMMARYINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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;
Messerschmitt et al., 2003;
Olichon et al., 2003). Fusion
and fission of mitochondria occurs dynamically in the cell, via fusion and
fission of its membranes (Pfanner et al.,
2004; Youle and Karbowski,
2005). In yeast, Mgm1 in the inner membrane, and Ugo1 and Fzo1 in
the outer membrane are involved in the fusion process
(Pfanner et al., 2004;
Wong et al., 2003).
Interestingly, when expression of the mammalian homolog of Mgm1, OPA1, was
reduced in HeLa cells by siRNA, reticular cristae were formed
(Griparic et al., 2004;
Olichon et al., 2003). S.
cerevisiae Mdm33 is involved in the opposite process, fission of the
mitochondrial inner membrane, and its overexpression leads to inner vesicle
formation, as in DIC-1 deficiency, and to partitioning of the matrix by
membrane septa (Messerschmitt et al.,
2003).
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;
Olichon et al., 2003),
implying a functional similarity between DIC-1 and OPA1. Nevertheless,
knockdown of EAT-3, a C. elegans homolog of human OPA1 and of yeast
Mgm1, resulted in mitochondrial fragmentation (T.H.L., unpublished), unlike
the disorganized and interconnected mitochondrial distribution evident in
Fig. 7B. It also resulted in a
reduced number of morphologically normal but longer cristae in cryo-electron
micrographs (J.Y.M., unpublished). Therefore, DIC-1 is not likely to be
involved in fusion of the inner mitochondrial membrane. When FIS1 or DRP1,
both of which are involved in fission of the mitochondrial outer membrane, was
knocked down in HeLa cells, the mitochondria became elongated and apoptosis
was reduced (Lee et al.,
2004). The differences between these phenotypic effects and those
due to DIC-1 deficiency exclude the possibility that DIC-1 participates in
fission of the mitochondrial membrane. Besides fusion and fission proteins,
such proteins as mitofilin and e and g subunits of mitochondrial ATP synthase
control cristae/inner membrane morphology. In mitochondria of HeLa cells
deficient in mitofilin, which normally forms a large multimeric protein
complex in the intermembrane space, concentrically packed inner membranes were
interconnected at numerous sites (John et
al., 2005). This `onion-like' structure was also induced in S.
cerevisiae cells by decreased expression of e or g subunit of the ATP
synthase, both of which are involved not in ATP synthesis but in the
oligomerization of the ATP synthase
(Arselin et al., 2004). Like
mitofilin and the ATP synthase e and g subunits, DIC-1 is thought to be
involved rather in remodeling (as a comprehensive term) of the mitochondrial
cristae/inner membrane, not in its fusion/fission.
|
).
Apoptosis caused by deficiency of dic-1 was observed at late
pachytene and midembryonic stages, the developmental stages in which
physiological apoptosis normally occurs. Cavity formation was observed at the
same or similar stages of germline development and embryogenesis, suggesting a
crucial role for DIC-1 at these stages. During the late pachytene/oogenic and
the mid-embryonic stages, cells undergo active differentiation to form oocytes
and specific tissues, respectively. Efficient mitochondrial activity is
probably more crucial in these differentiating cells than in actively dividing
cells, as has been observed in a dif-1 mutant
(Ahringer, 1995). DIF-1 is a
homolog of mitochondrial carrier proteins involved in solute transport through
the inner mitochondrial membrane and is essential during a 3-hour period of
active embryonic morphogenesis.
Human DICE1 differs in function and location from its C. elegans homolog
In summary, the DICE1 homolog in C. elegans is essential for
normal mitochondrial morphology and function, and its deficiency induces
apoptosis and inhibits development at a number of stages. The major
differences between C. elegans and mammalian DICE1 homologs are in
their subcellular locations and effects on cell proliferation. Although the
C. elegans protein localizes to mitochondria, the human and mouse
homologs localize to nuclei (Hoff et al.,
1998; Wieland et al.,
2001). When their amino acid sequences were analyzed using the
pTARGET program
(http://bioinformatics.albany.edu/~ptarget/),
C. elegans and human DICE1 are predicted to localize in mitochondria
at the confidence levels of 93.9% and 75.1%, respectively. However, nuclear
localization is predicted for both of the proteins by the pSORT program
(http://psort.ims.u-tokyo.ac.jp/)
at the probabilities of 73.9% (in C. elegans) and 82.6% (in humans).
In agreement with this prediction, bipartite nuclear targeting sequences are
identified by the PROSITE program
(http://www.expasy.org./prosite/)
in C. elegans (amino acids 677-693 and 693-709) and human DICE1
(amino acids 643-659). Although an exclusive localization of DIC-1 in
mitochondria has been observed in this study, it is possible that it
translocates to the nucleus under certain conditions. Recently, p53 was found
to translocate to mitochondria, as well as to nuclei upon activation by DNA
damage or oxidative stress, and it was localized in the matrix and outer
membrane of the mitochondria (Chipuk et
al., 2004; Zhao et al.,
2005).
In addition to the difference in subcellular localization, the C.
elegans and mammalian DICE1 homologs have distinct effects on cell
proliferation and differentiation. The C. elegans protein has an
anti-apoptotic role and is more important in differentiating cells than in
actively dividing ones, in contrast to the role of its mammalian homologs as
suppressors of cell proliferation (Hoff et
al., 1998; Wieland et al.,
2004). When dic-1(tm1615) heterozygotes were transformed
with a human DICE1 cDNA under the control of the C. elegans
hsp-16.2 promoter and expression was induced by heat shock at the L1
stage, no rescue of L3 larval arrest was observed in the homozygous
dic-1(tm1615) progeny. This was probably due to differences in the
subcellular location and function of the human homolog and C. elegans
DIC-1. However, it is also possible that a low level expression of the human
DICE1 cDNA or instability of the expressed protein could have caused
the failure. Human DICE1 was recently reported to be a component of the
integrator complex involved in small nuclear RNA processing
(Baillat et al., 2005), in
agreement with its nuclear localization. Nonetheless, the level of DICE1 mRNA
was found to be higher in dominant follicle cells of cow ovaries than in the
subordinate follicle cells that subsequently underwent apoptosis
(Evans et al., 2004). This
result, obtained with cDNA microarrays, suggests an anti-apoptotic role for
DICE1 in cows, as observed for its C. elegans homolog in the present
study. In summary, C. elegans DIC-1 plays a crucial role in the
formation of normal morphology of mitochondrial cristae/inner membrane, unlike
the mammalian homologs that localize in nuclei and are involved in the
suppression of cell proliferation. Our results on the C. elegans
DIC-1 suggest that a re-evaluation and further investigation on mammalian
DICE1 may elucidate its multiple functions and/or very complex intracellular
dynamics.
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ACKNOWLEDGMENTS |
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Footnotes |
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