1 Department of Molecular and Cellular Biology, Baylor College of Medicine, One
Baylor Plaza, TX 77030, USA
2 Program in Developmental Biology, Baylor College of Medicine, One Baylor
Plaza, TX 77030, USA
3 Department of Ophthalmology, Baylor College of Medicine, One Baylor Plaza, TX
77030, USA
* Author for correspondence (e-mail: kchoi{at}bcm.tmc.edu)
Accepted 4 June 2003
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SUMMARY |
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Key words: Drosophila, Crumbs, Discs-lost, Par-6, Photoreceptor, Rhabdomere
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INTRODUCTION |
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The crb and sdt genes were identified genetically as
essential components for organizing apicobasal polarity and AJs in embryonic
epithelia (Bachmann et al.,
2001; Bhat et al.,
1999
; Hong et al.,
2001
; Tepass et al.,
1990
). Genetic interaction studies suggested that sdt
acts downstream of crb in the same pathway
(Grawe et al., 1996
;
Tepass and Knust, 1993
).
Molecular analysis of Crb and Sdt has shown that they are directly associated
in the apical plasma membranes of epithelial cells
(Bachmann et al., 2001
;
Hong et al., 2001
). Crb is a
transmembrane protein with a long extracellular domain and a short C-terminal
cytoplasmic tail. The Crb-Sdt interaction is mediated by the single PDZ domain
of a MAGUK (a membrane-associated guanylate kinase) family protein Sdt and the
C-terminal PDZ domain binding motif (PBM) of the cytoplasmic domain of Crb
(Bachmann et al., 2001
;
Hong et al., 2001
). Crb complex
proteins are evolutionarily conserved. Studies in mammalian cell culture
systems indicate that Pals1 (mammalian Sdt homolog) links PATJ (Dlt homolog)
and CRB1 (Crb homolog) to form a Crb complex
(Roh et al., 2002
). Pals1 (or
Sdt) binds to the Crb-PBM motif (CrbPBM) via its single PDZ domain.
The L27 (Lin-2 and Lin-7 homology) domain of Pals1 interacts with a novel
protein-binding motif located at the N terminus of PATJ (or Dlt) termed MAGUK
recruitment domain (MRE) (Roh et al.,
2002
). However, in vivo function of vertebrate Sdt homologs is
largely unknown except that a zebrafish homolog Nagie oko localizes to the
apical cell junctions of the retinal neuroepithelium and may function as a
putative scaffolding factor (Wei and
Malicki, 2002
).
In addition to the Crb complex, apicobasal cell polarity is also regulated
by the Par-6 complex. Par-6 (partition-defective-6), initially found in C.
elegans, is essential for the establishment of asymmetry of early worm
embryo and for asymmetric localization of Par-3 (the homolog of
Drosophila Baz). Par-6 interacts with aPKC and Baz to form a
conserved ternary protein complex and affect aPKC activity. In contrast to Crb
and Sdt, Par-6 complex proteins are not only crucial for epithelial cell
polarity but also important for asymmetric division of various cell types in
different organisms (Rose and Kemphues,
1988; Suzuki et al.,
2001
; Wodarz et al.,
2000
). In Drosophila, homologs of these proteins (Par-6,
aPKC and Baz) colocalize at the apical side of epithelial cells and
neuroblasts in the embryo, and are essential for the establishment of
apicobasal asymmetry in both of these cell types
(Kuchinke et al., 1998
;
Petronczki and Knoblich, 2001
;
Wodarz et al., 2000
).
Mammalian homologs (PAR-6, aPKC and Par-3 or ASIP) form a similar ternary
complex that localizes to the tight junctions
(Ohno, 2001
). Studies in
cultured epithelial cells have suggested that this complex is important for
the formation of tight junctions and the specification of apical and
basolateral membrane domains (Izumi et
al., 1998
; Joberty et al.,
2000
; Lin et al.,
2000
; Suzuki et al.,
2001
). Thus, the Par-6 complex appears to act as a conserved
functional cassette for the generation of apicobasal asymmetries in diverse
cell types (Ohno, 2001
).
Recent studies on embryonic epithelia in Drosophila have provided
genetic evidence for interaction of Crb and Par-6 complex proteins
(Bilder et al., 2003
;
Tanentzapf and Tepass, 2003
).
Although a physical interaction between Sdt and Par-6 has been found in
mammalian cell cultures (Hurd et al.,
2003
), its role in epithelial tissues in developing animals has
not been demonstrated.
In this study, we focus on the analysis of mechanisms how the apical
domains of photoreceptor cells are organized. The apicobasal polarity is
prominent in the photoreceptors owing to the photosensitive organ, rhabdomere,
formed on the apical surface of the cell. During pupal eye development, the
apical domain of differentiating photoreceptors undergoes dynamic
reorganization of the cell shape and size, resulting in the formation of
rhabdomeres and new AJs (Kumar and Ready,
1995; Longley and Ready,
1995
). Recent studies have shown that Crb plays important roles in
morphogenesis of the photoreceptor rhabdomere, providing evidence that at
least some proteins involved in the apicobasal polarity of embryonic epithelia
are essential for the organization of photoreceptors
(Izaddoost et al., 2002
;
Pellikka et al., 2002
). Crb is
specifically localized to the rhabdomere stalk, a membrane domain that is
juxtaposed apically to the emerging rhabdomere and basally to the AJ. Crb is
required for positioning and growth of rhabdomere and AJ during the crucial
period of photoreceptor extension along the proximodistal axis of the retina.
Further analysis of Crb function has shown that the intracellular domain is
necessary for the recruitment of AJ as well as localization of Dlt
(Izaddoost et al., 2002
).
Importantly, the mammalian homolog of Crb localizes to the region
corresponding to the rhabdomere stalk membrane, that is, the inner segment
between the outer segment (analogous to the rhabdomere) and the AJ of rod
photoreceptors (Pellikka et al.,
2002
). Furthermore, mutations in CRB1, one of Crb
homologs in human, cause severe retinal dystrophies such as retinitis
pigmentosa type 12 (RP12) and Leber congenital amaurosis (LCA)
(den Hollander et al., 1999
;
den Hollander et al., 2001a
).
These studies suggest that Crb and other molecular components involved in the
specification of apical membrane of photoreceptors might be evolutionarily
conserved.
To understand further the molecular events required for apical morphogenesis of photoreceptors, we examined the localization, function and interaction of components of Crb and Par-6 complexes. We show that all proteins except Baz in the Crb and Par-6 ternary complexes colocalize to the rhabdomere stalk and are mutually required for their localizations. Baz localizes to the AJ but is essential for apical targeting of Crb and Par-6 complex proteins. Furthermore, these two protein complexes are linked by direct binding of Dlt and Par-6, providing the molecular basis for the interdependence of these two protein complexes.
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MATERIALS AND METHODS |
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Immunohistochemistry
For immunohistochemistry, pupal eyes at 40-50% pupal development (pd) were
fixed and stained with combinations of antibodies as previously described
(Izaddoost et al., 2002).
Primary antibodies used were rabbit anti-Dlt (1:500), mouse anti-Dlt (1:500),
rat anti-Crb (1:400) (Bhat et al.,
1999
), rabbit anti-Sdt (1:100)
(Bachmann et al., 2001
), mouse
anti-Arm (1:200; Hybridoma Bank), rat anti-DE-cad (1:50)
(Takeichi, 1988
), rabbit
anti-Baz (1:500) (Wodarz et al.,
1999
), rabbit anti-Par-6 (1:500)
(Petronczki and Knoblich,
2001
) and rabbit anti-PKC
C20 (1:500; Santa Cruz
Biotechnology). TRITC-conjugated phalloidin was from Sigma, whereas
fluorescent secondary antibodies were from Jackson Immunochemicals. Images
were scanned using a Zeiss LSM laser-scanning confocal microscope.
DNA constructs
GST-Dlt and its deletion constructs were described previously
(Bhat et al., 1999).
GST-Dlt-PDZ3 was constructed by deletion from the internal BamHI site
in the PDZ4 to the C terminus of Dlt. GST-Dlt-PDZ4 was made by deletion from
the N terminus of GST-PDZ34 to the internal ScaI site in the PDZ3.
GST- or MBP-fusion plasmids of Par-6 and aPKC were obtained by inserting
fragments amplified from an embryonic cDNA library (provided from K. Zinn)
into pGEX (Pharmacia) or pMal (NEB) in-frame. GST-Baz plasmid was obtained by
inserting NruI-SalI fragment containing three PDZ domains
(amino acids 110-947) from LD13977 (BDGP; the ORF is identical to that
available under GenBank Accession Number AJ130871) into pGEX.
MBP-Par-6
C was constructed using an internal BamHI site to
delete the C-terminal 107 amino acids. GST-Par-6
PDZ was generated by
PCR-based deletion of an internal domain (amino acids 171-247).
GST-Par-6
N was made by deletion of N-terminal 136 amino acids using an
internal EcoRV site.
In vitro pull-down binding assay using purified proteins
GST-, or MBP-tagged proteins were expressed in E. coli strain
BL21-CodonPlus (Stratagene), and purified by glutathione-agarose
(Sigma) or amylose resin (NEB), respectively. Equal amount of purified GST- or
MBP- fusion proteins immobilized on glutathioneagarose or amylose in PBS
containing 0.1% ß-mercaptoethanol, 0.1% BSA, and 0.05% Tween-20 were
mixed and incubated with equal amount of purified GST- or MBP-fusion proteins
for overnight at 4°C. After extensive washing with PBS containing 0.1%
ß-mercaptoethanol and 0.05% Tween-20 or 0.1% NP-40, the bound proteins
were eluted with 10 mM glutathione or 10 mM maltose and subjected to SDS-PAGE.
Eluted GST- or MBP-fusion proteins were detected by immunoblotting using
anti-GST (Sigma) or anti-MBP (NEB) antibody.
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RESULTS |
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The intracellular domain of Crb consists of the juxtamembrane domain (JM)
and the PDZ domain-binding motif (PBM)
(Klebes and Knust, 2000). It
has been shown that the JM and PBM are important for AJ and Dlt localization,
respectively (Izaddoost et al.,
2002
). To examine the Crb-dependent localization of Sdt further,
CrbPBM was overexpressed in the eye using GMR-GAL4
(Izaddoost et al., 2002
).
CrbPBM is the Crb intracellular domain in which the PBM is intact
but the JM is mutated (Klebes and Knust,
2000
). Overexpression of CrbPBM resulted in
mislocalization of Dlt throughout the cell membrane
(Fig. 2D). In this case, Sdt
was also mislocalized together with Dlt
(Fig. 2E,F). The
comislocalization of Sdt and Dlt was also observed when the
Crbintra or CrbMyc-intra was overexpressed using
GMR-GAL4 (data not shown). These results indicate that the
localization of Sdt in apical membrane is strictly dependent on Crb and
CrbPBM is sufficient to recruit Sdt.
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Sdt is required for positioning and extension of rhabdomere
In the absence of Crb, rhabdomeres of photoreceptors fail to expand along
the growing axis of the cells (Izaddoost
et al., 2002; Pellikka et al.,
2002
). To determine whether Sdt is also required for growth and/or
maintenance of rhabdomeres, we examined sdt- mutant
photoreceptors using phalloidin (Fig.
3I) as an F-actin marker. Phalloidin staining showed fragmented or
mispositioned rhabdomeres (Fig.
3I), indicating that rhabdomeres fail to grow normally and/or
maintain the structure along the length of photoreceptor cells. In a normal
ommatidium, the apical surface of photoreceptors points toward the center of
each cluster. In sdt- mutant clones, the apical domain of
photoreceptors failed to orient to the center, resulting in the formation of
rhabdomeres in displaced positions (Fig.
3I, arrows). These defects in AJs and rhabdomeres in
sdt- mutant clones mimic the phenotypes of
crb- photoreceptors
(Izaddoost et al., 2002
;
Pellikka et al., 2002
). As the
JM domain of Crb is important for correct positioning of AJ
(Klebes and Knust, 2000
;
Izaddoost et al., 2002
) and
the PBM domain of Crb is important for binding the Sdt
(Bachmann et al., 2001
;
Hong et al., 2001
),
sdt- phenotypes are likely to be due to the loss of Crb in
sdt mutant clones.
AJs in sdt- photoreceptors
(Fig. 3G) were mispositioned
basolaterally. A side view of wild-type ommatidium
(Fig. 3J) shows smooth and
well-defined AJs spanning the length of the retina from the surface to the
floor. But, the sdt- cells
(Fig. 3J) show discontinuous
and mispositioned Arm staining. Furthermore, a longitudinal section of the
sdt- clones in adult eye shows defects of rhabdomere
elongation (Fig. 3L). An
oblique section of the same eye (labeled with bracket) shows that the
rhabdomeres are missing in proximal region, but are relatively well organized
in distal region (Fig. 3K).
This phenotype of discontinuous rhabdomeres is similar to that of the
crb- mutant (Izaddoost
et al., 2002; Pellikka et al.,
2002
). Transmission electron microscopy was used to examine the
ultrastructure of sdt- rhabdomeres and AJs in adult eyes.
Rhabdomeres and AJs of wild-type ommatidia adjacent to
sdt- mutant clones in the same mosaic eye are uniform in
size and properly positioned (Fig.
3M). But, the sdt- photoreceptors show bulky
and fused rhabdomeres (Fig.
3N). The similar adult phenotype of sdt- and
crb- clones suggests that Crb and Sdt are involved in the
same cellular function.
Par-6 and aPKC colocalize with Crb complex in the rhabdomere
stalk
It has been shown that baz, encoding one of the components of
Par-6 complex, genetically interact with sdt
(Müller and Wieschaus,
1996). This led us to postulate that Crb and Par-6 complexes might
function together for the formation and/or maintenance of rhabdomere and AJ in
the photoreceptors. To test this, we examined whether the components of
the
Par-6 complex colocalize with Crb complex proteins in photoreceptor cells during pupal stages. As shown in Fig. 4A and B, both Par-6 and aPKC colocalized with Dlt at the rhabdomere stalks, suggesting that the Par-6 complex may play important roles in photoreceptor morphogenesis similar to Crb. By contrast, Baz colocalized with Arm at the AJ of photoreceptors basal to Par-6 and aPKC at the rhabdomere stalk (Fig. 4C,D). We examined third instar eye discs to see whether Baz and Par-6 might colocalize to the apical membrane in earlier stages of eye development. However, even in the third instar eye disc Par-6 and Baz localized separately to the apical Dlt domain and the AJ of photoreceptors, respectively, indicating that Baz is targeted to AJs from the early stage of photoreceptor differentiation (Fig. 4E-G). Interestingly, aPKC not only colocalizes with Dlt in the apical membrane but also overlaps with Arm (Fig. 4B; data not shown). Therefore, in contrast to Crb complex proteins that specifically localized to the apical membrane, Par-6 complex proteins appear to be sorted differently for localizations to the apical Crb domain and/or the AJ. Based on the separate localization of Baz from Par-6 and aPKC, `Par-6 complex' in photoreceptor membranes will be referred hereafter as a complex of Par-6 and aPKC.
|
To identify the region of Dlt that binds Par-6, we tested mutant forms of
Dlt proteins deleted in different PDZ domains for binding to Par-6. This assay
showed that the region of PDZ domain 3 (PDZ3) directly interact with Par-6
(Fig. 4I-J). As Par-6 contains
a conserved C-terminal PDZ-domain binding motif (-VLHL) of the class II
(Hung and Sheng, 2002), we
tested whether Dlt binds to Par-6 via the PBM. Interestingly, a mutated Par-6
protein lacking the C-terminal PBM residues was able to bind Dlt similar to
full-length of Par-6 (Fig. 4K).
As PDZ domains can interact with other PDZ domains
(Hung and Sheng, 2002
), we
tested whether the Dlt interacts with the PDZ domain of Par-6. Par-6
PDZ
mutant protein deleted in the PDZ domain still showed binding to Dlt. However,
Par-6 protein deleted in the N-terminal domain (Par-6
N) failed to bind
Dlt (Fig. 4L). This result
suggests that the N-terminal region of Par-6 is necessary for binding to the
PDZ3 region of Dlt (Fig.
4N).
Crb, Sdt and Dlt are required for localization of Par-6 complex
We have shown that Par-6 and Crb complexes not only colocalize but also
directly interact. To evaluate the physiological significance of this
interaction, we examined the effects of crb mutation on the
localization of Par-6/aPKC. In crb- photoreceptors, both
Par-6 and aPKC (Fig. 5A-F) were
displaced or partially lost from the apical region of the rhabdomere stalks.
Therefore, Crb is important for proper localization and maintenance of normal
level of Par-6 and aPKC although it may not be required for the formation of
Par-6 complex. It is important to note that the pattern of mislocalized Par-6
in crb- clones was identical to that of aPKC and Dlt, that
is, either mislocalized or lost altogether
(Fig. 5A-F). This suggests that
Dlt may associate with Par-6/aPKC complex in the absence of Crb.
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As shown earlier, the localization of Crb and Sdt is interdependent.
Therefore, we also examined whether the localizations of Par-6-aPKC-Baz are
similarly altered in sdt- mutant cells
(Fig. 6). Dlt was almost
entirely lost in the absence of Sdt, but Par-6 and aPKC were mislocalized and
significantly reduced (Fig.
6A-F). This phenotype appeared to be very similar to the
mislocalization of Par-6 and aPKC in crb- clones. The
pattern of abnormal Baz localization in sdt- and
crb- clones was also comparable (Fig.
5G-L and
6G-I). Therefore, loss of
either component of the Crb complex results in similar effects on the
localization of Par-6 complex proteins. As no dlt null mutation is
available, we used dltMY10, a 7.5kb deletion that
uncovers dlt and other flanking genes,
-spectrin,
cdc37 and JTBR (Jumping Translocation Breakpoint).
dltMY10 clones showed nearly complete loss of Sdt from the
apical membrane (Fig. 6J-L).
Moreover, sdt (Fig.
3D-F; Fig. 6A-I)
and dltMY10 (data not shown) clones showed essentially
indistinguishable pattern of mislocalization of Crb and Par-6 proteins,
consistent with the interdependent requirement of Dlt and Sdt for apical
organization of photoreceptors.
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In par-6- photoreceptors, Dlt and Arm were strongly reduced and/or mislocalized (Fig. 8A-B). Dlt and Arm were also mislocalized in aPKC- mutant photoreceptors (Fig. 8C-D). Interestingly, Dlt failed to localize to the membrane, resulting in a diffused distribution from the apical membrane (Fig. 8C). In baz- mutants, Arm was mislocalized (Fig. 8F) consistent with the localization of Baz to the AJ. The mislocalized Arm was detectable in proximal region, but completely lost in distal region (data not shown). Surprisingly, the baz- mutation caused almost complete loss of Dlt which is localized apical to the AJ in normal photoreceptors (Fig. 8E).
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DISCUSSION |
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Interdependency of Crb-Sdt-Dlt for their apical membrane
localization
In the absence of Crb, Sdt is mislocalized together with Dlt from the
rhabdomere stalk. By contrast, both Crb and Dlt are almost absent in
sdt mutants. Our results provide in vivo evidence for the
inter-dependent function of Crb complex proteins in the developing retina. The
strong dependence of Crb localization on Sdt and Dlt suggests that Crb may be
destabilized or may not be targeted to the membrane in the absence of Sdt or
Dlt. It is intriguing that Sdt and Dlt are lost only partially in the absence
of Crb. Our findings of a direct interaction between Dlt and Par-6 suggest
that Sdt-Dlt can still be targeted to the membrane in the absence of Crb
through the binding of Dlt to the Par-6 complex. However, it is important to
note that Dlt is essentially lost in sdt mutant clones and vice
versa. This raises an intriguing possibility that Dlt or Sdt are
dependent on each other in vivo to be targeted to the apical membrane via
binding to either Crb or Par-6. This mutual dependency between Dlt and Sdt may
explain why Dlt and Sdt are lost in the absence of the other, rather than
being associated with the Par-6 complex.
Direct interaction of Crb and Par-6 complexes
The interaction between the Crb and Par-6 complexes is mediated by the PDZ3
region of Dlt and the N-terminal domain of Par-6
(Fig. 4N). The N-terminal
domain of Par-6 (Fig. 4L) is
also used for binding aPKC (Joberty et
al., 2000; Lin et al.,
2000
; Qiu et al.,
2000
). Therefore, a potential function of Dlt is to bind Par-6 in
competition with aPKC or to facilitate the interaction of Par-6 with aPKC or
other Par-6 binding proteins. Our mutant analysis indicates that loss of Dlt
and Sdt in sdt- clones causes mislocalization of both Crb
and Par-6 complex proteins (Fig.
3D-F; Fig. 6A-I).
This suggests that Sdt-Dlt interaction provides a scaffold to recruit Crb
complex to the Par-6 complex and enhance the stability of these two complexes
rather than functioning as a competitor for aPKC.
Proteins in Crb and Par-6 complexes consist of multiple functional domains
which may be involved in diverse protein-protein interactions. A recent study
has shown that in mammalian cell culture systems the PDZ domain of Par-6 binds
not only Par-3 but also the N terminus of Pals1
(Hurd et al., 2003). These
results suggest that the crosstalk between the Crb and Par-6 complexes is
mediated by multiple domain-specific interactions. Evidence from our genetic
analysis using mutants suggests that the crosstalk between the two complexes
is mutually required for normal organization of apical membranes and AJs in
vivo, and also provides a basis for partial redundancy of these complexes in
the organization of photoreceptor cell polarity. Interestingly, when either
Crb or Sdt is lost, mislocalization or elimination of other associated
components including Par-6 complex proteins becomes more severe in the
age-dependent manner. This suggests that the Crb complex may be required for
the maintenance rather than the formation of the Par-6 complex. The
age-dependent degenerative phenotype may be related to the requirement of
extensive apical membrane growth to make rhabdomeres and AJs along the growing
axis of photoreceptors during pupal stage. Loss of any one component of the
Crb complex is likely to be increasingly more detrimental as the process of
membrane reorganization proceeds. In crb- or
sdt- mutants, significant fractions of Par-6 complex
proteins remain in the membrane despite the age-dependent and progressive
mislocalization of apical markers. By contrast, loss of Par-6 or aPKC resulted
in mislocalization of Dlt from the apical membrane
(Fig. 8). This suggests that
the Par-6 complex plays essential functions for membrane localization of Crb
complex proteins. Furthermore, both Par-6 and aPKC seem to be important for
survival and/or proliferation of retinal cells as mutant clones were very
small compared with adjacent twin spots and often completely disrupted
probably due to cell death. This is consistent with the findings of frequent
apoptosis in aPKC- or par-6- embryos
(Petronczki and Knoblich,
2001
; Wodarz et al.,
2000
).
Localization of Baz at AJs of photoreceptors
An important distinction of Par-6 complex in the photoreceptors from other
epithelia is the localization of Baz. Baz localizes with Crb complex in the
subapical membrane (Kuchinke et al.,
1998; Wodarz et al.,
2000
) or both the subapical region and AJ
(Bilder et al., 2003
) in the
Drosophila embryonic epithelia. Vertebrate Par-3 also localizes to
the apical tight junction in vertebrate epithelial cells
(Izumi et al., 1998
;
Suzuki et al., 2001
). By
contrast, Baz in the photoreceptors are specifically positioned in the AJs
basal to the all other proteins in the Crb/Par-6 complexes. As shown in
Fig. 7E, Baz and Arm are
recruited together to ectopic membrane sites by misexpression of
CrbJM, suggesting that Baz is an integral component of AJ. However,
Baz is not recruited by CrbPBM, whereas Par-6 and aPKC can be
ectopically recruited by CrbPBM rather than CrbJM.
Therefore, Baz appears to be recruited to AJ independently of Par-6/aPKC.
Intriguingly, despite its specific localization to AJs, loss of Baz
resulted in most severe disruption of AJ as well as the more apical Dlt
domain. It has been proposed that the Par-6/aPKC cassette is recruited to the
site of cell-cell contact and then moves along the most apical zone of the
developing cell-cell contact. In this process, an important step for cell
polarity formation is to tether the cytoplasmic Par-6/aPKC complex to the site
of cell-cell contact at the membrane which is mediated by the interaction of
Par-3 and a membrane protein JAM (Ohno,
2001). Therefore, our results that baz mutation causes
loss of Dlt and AJs support the crucial role of Baz in the initial step of
cell polarization. However, the distinct localization of Baz from Par-6 and
aPKC in the photoreceptors suggests that the mode of Baz localization varies
in different systems. In photoreceptors, Baz may be targeted to the membrane
with Par-6 but be sorted out from Par-6 in subsequent steps of polarization to
remain in the AJs, whereas Par-6-aPKC-Baz cassette remains together in the
complex in other epithelia. In contrast to Baz, aPKC localizes to both
rhabdomere stalk and AJ (Fig.
4B), suggesting that Baz and Par-6 are completely separated during
polarization while aPKC is not sorted from both Par-6 and Baz. The critical
function of Baz in the localization of Crb complex in the rhabdomere stalk is
consistent with the requirement of Baz for Crb localization in embryonic
epithelia (Bilder et al.,
2003
). However, the requirement of Baz in the embryo appears to be
dependent on the stage of development as Crb distribution in the absence of
Baz becomes normal in late embryos
(Tanentzapf and Tepass, 2003
).
On the contrary, such stage-dependent recovery of Crb complex localization has
not been observed in baz- photoreceptor cells.
Recent studies have shown that mutations in human CRB1 cause RP12
and LCA, severe recessive retinal diseases, emphasizing the importance of Crb
family proteins in the eyes of mammals including humans. The
Drosophila Crb and human CRB1 are localized in analogous subcellular
membrane domains of photoreceptors, the rhabdomere stalk and the inner segment
in Drosophila and human photoreceptors, respectively. Besides similar
subcellular localization, Crb and human CRB1 are functionally conserved
(den Hollander et al., 2001b;
Izaddoost et al., 2002
;
Roh et al., 2002
).
Age-dependent photoreceptor defects in crb mutant also provide
analogy to age-dependent retinal degeneration in RP12/LCA patients. Our
studies here imply that hCRB1 may function as a protein complex with homologs
of Sdt and Dlt and such complex may interact with a homologous Par-6 complex.
Whether such homologous human genes are the targets of inherited retinal
diseases such as RP remains to be studied.
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ACKNOWLEDGMENTS |
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REFERENCES |
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