Institut für Biochemie, Universität Erlangen, Fahrstrasse 17, D-91054 Erlangen, Germany
* Author for correspondence (e-mail: m.wegner{at}biochem.uni-erlangen.de)
Accepted 9 February 2004
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SUMMARY |
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Key words: Sry, High-mobility-group, Sox10, Redundancy, Gene dosage, Oligodendrocyte
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Introduction |
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Sox8 has been recently identified as a member of Sox group E
(Pfeifer et al., 2000;
Schepers et al., 2000
) and
shares extensive sequence homology with Sox9 and Sox10, the other two members
of this group of Sox proteins (Wegner,
1999
). Sox9 and Sox10 have been characterized extensively for
their involvement in chondrogenesis, male sex determination, neural crest
development and gliogenesis, respectively
(Bi et al., 1999
;
Britsch et al., 2001
;
Foster et al., 1994
;
Herbarth et al., 1998b
;
Pingault et al., 1998
;
Southard-Smith et al., 1998
;
Stolt et al., 2003
;
Stolt et al., 2002
;
Wagner et al., 1994
).
Although expression pattern and behaviour in cell culture systems are
indicative of Sox8 functions in the male gonad and in skeletal muscle
(Schepers et al., 2003;
Schmidt et al., 2003
),
analysis of Sox8-deficient mice has so far failed to reveal severe defects in
the development of major organ systems
(Sock et al., 2001
).
Redundancy and functional compensation by related factors is one of the most
plausible explanations for the lack of a severe phenotype in Sox8-deficient
mice. Supporting such an assumption, most Sox8-expressing cells are also
positive for either Sox9 or Sox10 (Sock et
al., 2001
). In case of the male gonad, for example, Sertoli cells
express Sox8 as well as Sox9 (Schepers et
al., 2003
). If functional compensation indeed exists between the
three Sox E proteins, it must be non-reciprocal as both Sox9- and
Sox10-deficient mice exhibit severe developmental defects despite the
continued presence of Sox8.
We have previously noted that Sox8 expression in the developing spinal cord
is indicative of an expression in the oligodendrocyte lineage
(Sock et al., 2001). These
glial cells of the central nervous system arise from neural stem cells in a
limited domain of the ventral ventricular zone
(Pringle and Richardson,
1993
), which earlier gives rise to motoneurons
(Briscoe et al., 1999
). From
this pMN domain, oligodendrocyte progenitors spread throughout the whole
spinal cord as embryogenesis proceeds. At the end of embryogenesis, they
accumulate in the marginal zone and start to differentiate. Terminal
differentiation of oligodendrocytes peaks during the first postnatal weeks,
and is characterized by the production of large amounts of lipids and a
limited set of myelin proteins such as myelin-associated glycoprotein (Mag),
myelin basic protein (Mbp) and proteolipid protein (Plp)
(Lemke, 1988
). The resulting
formation of myelin sheaths around axons allows rapid saltatory conductance in
the central nervous system (CNS).
Recently, several transcription factors have been identified as
cell-intrinsic regulators of oligodendrocyte development. These include the
Olig1 and Olig2 bHLH proteins, neurogenins and Nkx2.2
(Lu et al., 2002;
Nieto et al., 2001
;
Qi et al., 2001
;
Takebayashi et al., 2002
;
Zhou and Anderson, 2002
;
Zhou et al., 2001
). Sox
proteins also feature prominently during development of these cells. Early
specification of oligodendrocyte progenitors requires Sox9
(Stolt et al., 2003
). Once
specified, oligodendrocyte progenitors express both Sox9 and Sox10, and can
cope with loss of either protein (Stolt et
al., 2003
; Stolt et al.,
2002
). A role for Sox10 becomes evident at the onset of terminal
differentiation when Sox9 expression is naturally extinguished in
oligodendrocytes. In Sox10-deficient spinal cords, only few oligodendrocyte
progenitors start to express myelin proteins with delayed kinetics and in
amounts insufficient for myelination
(Stolt et al., 2002
). The
residual myelin gene expression in Sox10-deficient spinal cords is compatible
with the existence of a third activity besides Sox9 and Sox10 with overlapping
expression and function. In view of its possible expression in the
oligodendrocyte lineage (Sock et al.,
2001
), Sox8 is an attractive candidate. Its role during
oligodendrocyte development in the spinal cord is addressed in the present
study.
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Materials and methods |
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Tissue preparation, immunohistochemistry, X-Gal staining, in situ hybridization and quantification of ß-galactosidase
Embryos (from 12.5 dpc to 18.5 dpc) and pups (at postnatal days 1, 3 and 7)
were obtained from staged pregnancies. Spinal cords were additionally
dissected from adult animals. After fixation in paraformaldehyde, genotyped,
age-matched tissues were sectioned on a cryotome. Sections (10 µm) were
used for immunohistochemistry, 20 µm sections for X-Gal staining and in
situ hybridization according to standard protocols as previously described
(Stolt et al., 2003;
Stolt et al., 2002
). For
better comparison, all shown spinal cord sections are from the forelimb level.
For immunohistochemistry, the following primary antibodies were used in
various combinations: anti-Mbp mouse monoclonal (1:100 dilution, Chemicon),
anti-GFAP mouse monoclonal (1:100 dilution, Chemicon), anti-NeuN mouse
monoclonal (1:500 dilution, Chemicon), anti-Mag mouse monoclonal (undiluted,
gift of U. Bartsch, Hamburg University), anti-Olig2 rabbit antiserum (1:2000
dilution, gift of H. Takebayashi, Kyoto University), anti-Sox10 rabbit
antiserum (1:100 dilution) (Stolt et al.,
2003
), anti-ß-galactosidase rabbit antiserum (1:500 dilution,
ICN) or anti-ß-galactosidase goat antiserum (1:500 dilution, Biotrend).
Secondary antibodies conjugated to Cy2 and Cy3 immunofluorescent dyes
(Dianova) were used for detection. In situ hybridization was performed with
DIG-labeled antisense riboprobes for Mbp and Plp
(Stolt et al., 2002
). Samples
were analyzed and documented using either a Leica TCS SL confocal microscope
or a Leica inverted microscope (DMIRB) equipped with a cooled MicroMax CCD
camera (Princeton Instruments, Stanford, CA). Quantification of
ß-galactosidase was performed on extracts prepared by homogenization of
freshly dissected spinal cords using a chemiluminescent ß-galactosidase
detection assay (Roche Biochemicals).
Cell culture, RT-PCR and luciferase assays
N2A neuroblastoma cells were maintained in Dulbecco's Modified Eagle's
Medium containing 5% fetal calf serum, and transfected using Superfect reagent
(Qiagen). Stable N2A cell clones capable of doxycycline-dependent, inducible
Sox8 expression were generated as previously described for Sox10
(Peirano et al., 2000). RNA
from these cells was prepared in both the induced (Sox8 positive) and
uninduced (Sox8 negative) state. After reverse transcription to cDNA,
semi-quantitative PCR was performed to detect products specific for Sox8, Plp,
Mbp and ß-actin. For luciferase assays, N2A cells were transfected
transiently in duplicates in 24-well plates with 200 ng of luciferase reporter
plasmid and 200 ng of effector plasmids per well. Luciferase reporters
containing a long version (positions -656 to +31) and a short version
(positions -256 to +31) of the Mbp promoter were used
(Stolt et al., 2002
). Effector
plasmids corresponded to pCMV5-based expression plasmids for Sox8
(Schmidt et al., 2003
) and
Sox10 (Kuhlbrodt et al.,
1998a
). Cells were harvested 48 hours post-transfection, and
extracts were assayed for luciferase activity
(Stolt et al., 2002
).
Protein extracts and electrophoretic mobility shift assay
Extracts from transfected N2A cells (10 cm dishes) ectopically expressing
full-length or shortened Sox8 and Sox10 proteins, were prepared as described
(Kuhlbrodt et al., 1998b). For
electrophoretic mobility shift assays, protein extracts were incubated with
0.5 ng of 32P-labeled oligonucleotide probes for 20 minutes on ice
in a 20 µl reaction mixture containing 10 mM Hepes (pH 8.0), 5% glycerol,
50 mM NaCl, 5 mM MgCl2, 2 mM DTT, 0.1 mM EDTA, 4 µg of bovine
serum albumin, and 2 µg of poly(dGdC) as unspecific competitor. The
following oligonucleotide probes were used: sites 1-3 from the Mbp
promoter (Stolt et al., 2002
)
and the prototypic dimeric binding site C/C' from the Protein zero
promoter (Peirano et al.,
2000
; Peirano and Wegner,
2000
; Schlierf et al.,
2002
). For supershift experiments, 0.1 µl of antisera were
additionally added after 10 minutes, and incubation was continued for a
further 10 minutes. Samples were loaded onto native 4% polyacrylamide gels and
electrophoresed in 0.5xTBE (45 mM Tris/45 mM boric acid/1 mM EDTA, pH
8.3) at 120 V for 1.5 hours. Gels were dried and exposed for
autoradiography.
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Results |
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At 12.5 days post coitum (dpc), X-gal staining was restricted within the spinal cord to a limited domain in the ventral part of the ventricular zone whose position is identical to or strongly overlapping with the pMN domain (Fig. 1A). Two days later, ß-galactosidase-positive cells were no longer restricted to this domain, but had dispersed throughout the parenchyma of the spinal cord in a pattern typical of glial progenitors (Fig. 1B). The number of these mantle zone cells increased through 16.5 dpc (Fig. 1C). At 18.5 dpc, ß-galactosidase-positive cells had started to accumulate in the marginal zone (Fig. 1D) and continued to do so postnatally (Fig. 1E) as expected if at least some belonged to the oligodendrocyte lineage.
|
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We also generated and analyzed Sox8-deficient mice which additionally lacked one Sox10 allele (Sox8lacZ/lacZ, Sox10+/lacZ). Mice deficient for both Sox8 and Sox10 could not be obtained for the stages analyzed in this study due to early embryonic lethality (C.C.S., P.L. and M.W., unpublished data). Embryonic spinal cords from Sox8lacZ/lacZ, Sox10+/lacZ mice exhibited a normal number of oligodendrocytes at 16.5 dpc and 18.5 dpc as judged from Olig2 and Sox10 expression (Fig. 3D; data not shown). Additionally, the distribution of oligodendrocyte progenitors throughout the spinal cord was indistinguishable from those in control genotypes. Direct quantification of Olig2-positive cells in spinal cord sections of the various genotypes from comparable axial levels confirmed that there is no significant difference between genotypes in the number of Olig2-positive cells at 18.5 dpc (Fig. 4A).
|
Loss of a single Sox10 allele also led to a decrease in both Mbp- and Plp-expressing cells (Fig. 3G,K). According to our quantifications, Sox10+/lacZ spinal cords exhibited a 53-60% reduction at 18.5 dpc. Terminal differentiation of oligodendrocytes is thus slightly stronger affected in Sox10 heterozygous than in Sox8-deficient mice (Fig. 4C).
When both mutations were combined in Sox8lacZ/lacZ, Sox10+/lacZ mice, expression of Mbp and Plp was severely reduced. Almost no terminally differentiating oligodendrocytes were detected at 18.5 dpc by these markers in spinal cords of Sox8lacZ/lacZ, Sox10+/lacZ mice (Fig. 3H,L; Fig. 4C), arguing that both Sox proteins cooperate during this process.
To analyze whether terminal oligodendrocyte differentiation is permanently
or transiently affected, in situ hybridization studies of myelin gene
expression were continued in the postnatal spinal cord. At postnatal day 3, we
still observed a reduction in Mbp- and Plp-expressing cells in both
Sox8-deficient and Sox10 heterozygous spinal cords relative to the wild type
(Fig. 5A-C,E-G). Despite
Olig2-positive cell numbers comparable with the wild type
(Fig. 4B), Mbp- and
Plp-expressing cells were similarly reduced in Sox8-deficient and Sox10
heterozygous mice by 30% (Fig.
4D). Decreased expression of Mbp and Plp was confirmed on the
protein level for both genotypes by immunohistochemistry (see Fig. S1AC,E-G at
http://dev.biologists.org/supplemental/).
Because of the preferential occurrence of both myelin proteins in
oligodendrocyte processes and forming myelin sheaths, it is impossible to tell
from immunohistochemical analyses whether signal reduction is due to reduced
numbers of expressing cells or also due to reduced cellular expression levels.
At postnatal day 3, both Sox8-deficient and Sox10 heterozygous spinal cords
also exhibited weakened immunoreactivity for Mag (see Fig. S1I-K at
http://dev.biologists.org/supplemental/),
which unlike Mbp and Plp is not under direct Sox10 control
(Stolt et al., 2002
).
|
Postnatal mortality among Sox8lacZ/lacZ, Sox10+/lacZ mice is high. Nevertheless, some mice survive for several days so that oligodendrocyte development can be followed in this genotype throughout the first postnatal week. Although spinal cord cells positive for Mbp or Plp transcripts appeared during postnatal development, their number remained strongly reduced at both 3 and 7 days after birth (Fig. 5D,H,L,P). At postnatal day 3, the number of Plp- or Mbp-expressing cells was only 10-15% compared with the wild type (Fig. 4D). Co-immunohistochemistry with antibodies directed against Mbp, Plp or Mag all supported the conclusion that terminal differentiation of oligodendrocytes is severely affected in Sox8lacZ/lacZ, Sox10+/lacZ mice (see Fig. S1D,H,L,P,T,X at http://dev.biologists.org/supplemental). Some Sox8lacZ/lacZ, Sox10+/lacZ exhibited unsteady movements and action tremor, pointing to hypomyelination of the CNS. Although our results point to a severe delay rather than a complete block in oligodendrocyte differentiation, we do not know whether oligodendrocyte differentiation would ever be robust enough for significant myelin formation to occur in the CNS of Sox8lacZ/lacZ, Sox10+/lacZ mice. During this study, Sox8lacZ/lacZ, Sox10+/lacZ mice did not survive past postnatal day 8 (data not shown). In fact, the oligodendrocyte defect and ensuing hypomyelination might be one of the reasons for the high postnatal mortality of these mice.
Sox8 is capable both of binding to myelin gene promoters and interacting with Sox10
One of the mechanisms by which Sox10 is able to influence terminal
differentiation of oligodendrocytes is through direct binding to myelin gene
promoters and transcriptional regulation of myelin gene expression
(Stolt et al., 2002). Sox10
interacts with three binding sites in the proximal part of the Mbp
promoter which together mediate its Sox10-dependent activation
(Stolt et al., 2002
). Two of
these sites (sites 2 and 3) bind a single Sox10 molecule, whereas site 1 is
cooperatively bound by two Sox10 molecules
(Stolt et al., 2002
). When gel
retardation assays were performed with these sites and extracts that contain
either full-length Sox10 or full-length Sox8, we obtained characteristic
protein-DNA complexes on all three sites
(Fig. 6A). Addition of
antibodies directed against Sox10 selectively supershifted the
Sox10-containing complex. In a reciprocal manner, antibodies specific for Sox8
led to a selective supershift of the Sox8-specific complex and left the Sox10
complex unaffected, indicating that Sox8 was indeed contained within the
complex. The mobility obtained for the Sox8-specific complexes was very
similar to that obtained for the corresponding Sox10 complexes in good
agreement with the fact, that both proteins have very similar sizes. It is
also evident that the mobility of those complexes obtained with site 1 was
lower than those obtained with either site 2 or site 3
(Fig. 6A). In case of Sox10,
this lower mobility is due to cooperative binding of two molecules
(Stolt et al., 2002
). We
conclude from the nearly identical mobility of the Sox8-containing complex
that Sox8 also binds to site 1 as a dimer.
|
On dimeric sites, options are different (Fig. 6C). When incubated alone with a dimer site, both a shortened Sox8 and a long Sox10 version yielded one predominant complex with a mobility characteristic of the respective homodimer. When mixed, a new complex with intermediate mobility appeared, showing that heterodimerization between Sox8 and Sox10 had occurred. Thus, dimeric sites in target gene promoters allow Sox8 and Sox10 to functionally interact with each other in DNA-dependent heterodimers.
Sox8 directly activates expression of oligodendroglial myelin genes
Given the fact that Sox8 recognizes bona fide Sox10 target sites by itself
and as heterodimers with Sox10, we analyzed whether Sox8 would be able to
activate the Mbp promoter which has previously been shown to be
responsive in transient transfections to Sox10
(Stolt et al., 2002).
Luciferase expression was stimulated approximately eightfold by Sox10 when
placed under the control of a long version or a short version of the
Mbp promoter (Fig.
7A). Luciferase expression from the Mbp promoter was also
increased by Sox8. Although the approximately five- to sevenfold activation
rates were below those obtained for Sox10, this difference was not
statistically significant. Similar activation rates were obtained in the
presence of both Sox8 and Sox10 (data not shown).
|
Sox8 is expressed at lower levels than Sox10 in terminally differentiating oligodendrocytes
Given the fact that the same lacZ-coding sequence was inserted in
almost identical manner in both the Sox8 and the Sox10 locus
to yield the Sox8lacZ and Sox10lacZ
alleles, it should be possible to compare expression levels of Sox8 and Sox10
in developing oligodendrocytes through measurements of ß-galactosidase
amounts in Sox8+/lacZ and Sox10+/lacZ
spinal cords, provided expression levels do not undergo abrupt changes, that
might not be reproducible by ß-galactosidase with its long half-life.
However, no such drastic changes in expression levels have been observed for
Sox8 or Sox10 in oligodendrocytes (Sock et
al., 2001; Stolt et al.,
2002
) (this study).
At 14.5 dpc, ß-galactosidase was produced at four-fold higher amounts from the Sox10 locus than from the Sox8 locus (Fig. 7C). As development proceeded, this difference became less pronounced. At 16.5 dpc and 18.5 dpc, ß-galactosidase expression from the Sox8 locus reached 50-60% of the levels obtained for Sox10 (Fig. 7C). At the onset of terminal differentiation, Sox10 expression levels were thus approximately twice as high as Sox8 levels. This difference vanished further in adult animals, where 85% of Sox10 levels were achieved for Sox8 (Fig. 7C). From 18.5 dpc onwards, ß-galactosidase levels in spinal cords from mice with various combinations of lacZ alleles corresponded to the additive value obtained for the single Sox8lacZ and Sox10lacZ alleles (data not shown), arguing that at these times Sox8 and Sox10 expression in oligodendrocytes are largely independent of each other.
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Discussion |
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We show that in the late embryonic, early postnatal and adult spinal cord,
Sox8 is selectively expressed in the oligodendrocyte lineage. At these times,
we failed to detect expression in either astrocytes or neurons. In agreement
with previous observations about the general expression pattern of Sox8
(Sock et al., 2001),
oligodendrocytic expression was already present before terminal
differentiation, and maintained throughout and thereafter. This expression
pattern is strongly reminiscent of the related Sox10
(Stolt et al., 2002
) and
additionally shows a strong overlap with Sox9
(Stolt et al., 2003
).
Oligodendrocytes thus express all three group E Sox proteins before terminal
differentiation, and a combination of Sox8 and Sox10 from thereon. Despite
this significant overlap in Sox gene expression, there are defined
defects in oligodendrocyte development in Sox9- and Sox10-deficient spinal
cords, with Sox9 playing a role in oligodendrocyte specification
(Stolt et al., 2003
) and Sox10
being required for terminal differentiation
(Stolt et al., 2002
).
Sox8-deficient mice are viable and reach a normal lifespan without
exhibiting overt neurological symptoms
(Sock et al., 2001).
Therefore, severe oligodendrocyte defects were not expected. There was,
however, a transient delay in terminal differentiation and myelination of
spinal cord oligodendrocytes. Delayed terminal differentiation has previously
been observed in Nkx2.2-deficient mice and in Olig1-deficient mice for
oligodendrocytes (Lu et al.,
2002
; Qi et al.,
2001
), and in Oct6-deficient mice for Schwann cells, the
oligodendrocyte counterparts of the peripheral nervous system
(Bermingham et al., 1996
;
Jaegle et al., 1996
).
Interestingly, we also found a differentiation delay in
Sox10+/lacZ oligodendrocytes arguing that this process is
as sensitive to Sox10 gene dosage as the development of melanocytes
and the enteric nervous system (Britsch et
al., 2001
; Herbarth et al.,
1998a
; Pingault et al.,
1998
; Southard-Smith et al.,
1998
).
The delay of terminal oligodendrocyte differentiation in
Sox10+/lacZ spinal cords was drastically increased by the
absence of Sox8, and still prominent in Sox8lacZ/lacZ,
Sox10+/lacZ mice at the end of the first postnatal week.
Owing to lethality, we were unable to determine whether terminal
differentiation of oligodendrocytes would ever be sufficient for sustained CNS
myelination in this genotype. Retinal transplantation of neural stem cells
indicated that this is not the case for Sox10-deficient oligodendrocytes
(Stolt et al., 2002). Thus, it
will be interesting to see in future experiments whether the terminal
differentiation defect in Sox8lacZ/lacZ,
Sox10+/lacZ oligodendrocytes is as severe as in
Sox10lacZ/lacZ oligodendrocytes.
Our data argue that Sox8 and Sox10 perform redundant functions during
terminal differentiation of oligodendrocytes. We have shown here that Sox8 is
able to bind to a number of bona fide Sox10 response elements in a manner
indistinguishable from Sox10. Additionally, Sox8 and Sox10 bind cooperatively
to some response elements as heterodimers in a manner similar to the
respective homodimers. Thus, there is no evidence for differential DNA binding
or promoter recognition between both Sox proteins. In good agreement, Sox8
activated expression from the Mbp promoter in transient transfections
and induced expression from the endogenous Mbp and Plp genes
in N2A cells in a manner similar to Sox10
(Bondurand et al., 2001;
Peirano et al., 2000
;
Stolt et al., 2002
).
Interestingly, we have previously observed in vivo a weak residual Mbp
expression in some Sox10-deficient oligodendrocytes
(Stolt et al., 2002
). As Sox8
was the only remaining Sox E protein in the cells, the residual Mbp expression
might be attributed to the activity of Sox8.
Redundancy of structurally related transcription factors has been frequently observed. Usually, the redundant proteins can fully compensate each other's loss so that their function is unmasked only after combined deletion. This is not the case for Sox8 and Sox10, as Sox10 compensates loss of Sox8 much more effectively than vice versa.
One possible explanation for this non-reciprocal compensation is that Sox8 can only substitute for Sox10 in specific functions, so that Sox8 turns on only a subset of Sox10 target genes. The failure to activate the full complement of Sox10 target genes would then be causative for the severe terminal differentiation defect in Sox10-deficient oligodendrocytes. Although it is impossible to discount such a model in the absence of the full list of oligodendrocytic target genes for both proteins, there is little evidence so far that would favor this model as the main cause for the observed effects.
If Sox8 activates more or less the same transcriptional program in
oligodendrocytes as Sox10, it might do so less efficiently. One reason for a
reduced efficiency could be a reduced intrinsic transactivation capacity of
the Sox8 protein relative to Sox10. Sequence conservation between Sox8 and
Sox10 proteins are significant within the respective transactivation domains
of both proteins but not as high as in the DNA-binding domain
(Kuhlbrodt et al., 1998a;
Schepers et al., 2000
). Thus,
it is at least conceivable that interaction with the transcription machinery,
chromatin remodeling activities or transcriptional co-factors is qualitatively
or quantitatively different between both Sox proteins, as suggested for Sox8
and Sox9 (Schepers et al.,
2003
). Our transient transfection data would not exclude such a
model as maximal induction rates obtained for saturating amounts of Sox8 were
lower than those obtained for Sox10. However, induction rates did not differ
enough to identify the intrinsic transactivation capacities of the proteins as
the main difference between Sox8 and Sox10 in oligodendrocytes.
Another explanation focuses on the lower expression levels of Sox8 relative to Sox10 in oligodendrocytes as determined by comparison of ß-galactosidase activities in Sox8+/lacZ and Sox10+/lacZ mice. The difference is most pronounced during early oligodendrocyte development. However, even at the time of terminal oligodendrocyte differentiation, we still detected a twofold higher expression level of the lacZSox10 allele. Although certainly oversimplified, Sox gene function in the oligodendrocyte lineage might be explained at first approximation by a model in which the different expression levels of the respective Sox proteins are taken into account and functions are regarded as approximately equal. Thus during terminal differentiation, one Sox10 allele contributes roughly one-third to the overall Sox gene activity, whereas one Sox8 allele accounts for one-sixth. If the total amount of Sox protein drops to approximately two-thirds (as is the case for the Sox10 heterozygote and the Sox8-deficient mouse), minor disturbances become evident in the form of a transient delay of terminal differentiation. Severity of this defect increases with decreasing amounts of residual Sox protein as is the case after combined losses of Sox8 and Sox10 alleles. This leads to the dramatically extended terminal differentiation delay in Sox8lacZ/lacZ, Sox10+/lacZ mice, in which the total amount of remaining Sox protein has dropped to approximately one-third.
Although differences in expression levels are thus capable of explaining the different role of Sox8 and Sox10 during terminal differentiation of oligodendrocytes, it is almost certain that there will be other modulating factors such as differences in protein stability or, as already discussed, differences in transactivation capacities between both Sox proteins. Their contribution will be most easily revealed in mouse models in which Sox8 is inserted into the Sox10 locus and therefore expressed at levels characteristic of Sox10.
Sox8 and Sox10 are not the only structurally related transcription factors
that are co-expressed during development of myelinating glia. The bHLH
transcription factors Olig1 and Olig2 are both found in cells of the
oligodendrocyte lineage and behave similarly in gain-of-function analyses.
Nevertheless, their deletion leads to different phenotypes in the mouse with
loss of Olig2 leading to an early specification defect and loss of Olig1
affecting maturation of oligodendrocytes
(Lu et al., 2002). This argues
that loss of Olig1 is compensated unidirectionally by Olig2 during early
phases of oligodendrocyte development.
Myelinating Schwann cells express Oct6 and the closely related POU protein
Brn2 with a similar developmental profile. Analysis of mice deficient for both
Oct6 and Brn2 and the phenotypic rescue of Oct6-deficiency by ectopic Brn2
expression both support the notion that the two POU proteins perform redundant
functions (Jaegle et al.,
2003). Nevertheless, the delay in Schwann cell differentiation is
more pronounced in Oct6-deficient mice than in mice that lack Brn2, indicating
that both proteins have a different importance during Schwann cell
development. There is also some indication that Brn2 is present in lower
amounts than Oct6. The relationship between Oct6 and Brn2 in Schwann cells is
thus strongly reminiscent of the one between Sox10 and Sox8 in
oligodendrocytes. It remains to be seen whether these unidirectional
redundancies are a more general phenomenon for important developmental
regulators of myelinating glia. Because of their unidirectionality, it is
tempting to speculate that they do not primarily present a fail-safe mechanism
to ensure proper myelination but rather are an expression of the relatively
young evolutionary history of vertebrate glia.
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ACKNOWLEDGMENTS |
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Footnotes |
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