 |
 Previous Article | Next Article 
The Journal of Neuroscience, May 15, 1998, 18(10):3749-3756
A Neuronal Form of the Cell Adhesion Molecule L1 Contains a
Tyrosine-Based Signal Required for Sorting to the Axonal Growth
Cone
Hiroyuki
Kamiguchi and
Vance
Lemmon
Department of Neurosciences, Case Western Reserve University,
Cleveland, Ohio 44106
 |
ABSTRACT |
The neural cell adhesion molecule L1, which is present on axons and
growth cones, plays a crucial role in the formation of major axonal
tracts such as the corticospinal tract and corpus callosum. L1 is
preferentially transported to axons and inserted in the growth cone
membrane. However, how L1 is sorted to axons remains unclear.
Tyr1176 in the L1 cytoplasmic domain is adjacent to
a neuron-specific alternatively spliced sequence, RSLE
(Arg-Ser-Leu-Glu). The resulting sequence of
YRSLE conforms to a tyrosine-based consensus
motif (YxxL) for sorting of integral membrane proteins into specific cellular compartments. To study a possible role of the YRSLE sequence in L1 sorting, chick DRG neurons were transfected with human L1 cDNA
that codes for full-length L1 (L1FL), a non-neuronal
form of L1 that lacks the RSLE sequence (L1 RSLE), mutant
L1 with a Y1176A substitution (L1Y1176A), or L1
truncated immediately after the RSLE sequence (L1C77).
L1FL and L1C77, both of which possess the
YRSLE sequence, were expressed in the axonal growth cone and to a
lesser degree in the cell body. In contrast, expression of both
L1RSLE and L1Y1176A was restricted to the
cell body and proximal axonal shaft. We also found that
L1RSLE and L1Y1176A were integrated into the
plasma membrane in the cell body after missorting. These data
demonstrate that the neuronal form of L1 carries the tyrosine-based
sorting signal YRSLE, which is critical for sorting L1 to the axonal
growth cone.
Key words:
neural cell adhesion molecule; L1; axon; growth cone; protein sorting; tyrosine-based sorting signal
| |
INTRODUCTION |
The neural cell adhesion molecule L1
is a transmembrane protein belonging to the immunoglobulin (Ig)
superfamily (Moos et al., 1988 ). The amino acid sequence of the
cytoplasmic domain of L1 is identical among mammalian species (Hlavin
and Lemmon, 1991). The L1 gene is composed of 28 exons, two of which
(exons 2 and 27) are alternatively spliced (Reid and Hemperly, 1992; Takeda et al., 1996). The exon 27 encodes the four amino acids Arg-Ser-Leu-Glu (RSLE) near the middle of the cytoplasmic domain (Miura
et al., 1991), which is expressed in neurons but not in the other
L1-expressing cells, such as Schwann cells, melanocytes, and
lymphocytes (Takeda et al., 1996).
L1 is involved in various important processes during nervous system
development, including neuronal migration (Lindner et al., 1983; Barami
et al., 1994), neurite growth (Lagenaur and Lemmon, 1987), and neurite
fasciculation (Stallcup and Beasley, 1985). L1 is expressed on the
surface of axonal shafts and growth cones of developing neurons.
Homophilic binding of L1 molecules is probably its most common mode of
action in promoting axonal extension along a bundle of preexisting
axons, forming fascicles (Grumet and Edelman, 1988; Lemmon et al.,
1989). In humans, mutations of the L1 gene cause X-linked
hydrocephalus, in which defects of the corticospinal tract and corpus
callosum are also found (Wong et al., 1995b). Similar malformations
have been reported in the L1 knock-out mouse (Cohen et al., 1997; Dahme
et al., 1997). The dramatic alterations in the formation of major
axonal tracts after mutations in the L1 gene clearly demonstrate that
L1 plays a crucial role in nervous system development.
During the period when axons elongate toward their proper targets,
their tips form a specialized sensory structure called the growth cone,
which interacts with environmental cues to produce directed axonal
growth. Growth cones undergo rapid changes in shape with concomitant
reorganization of cytoskeletal elements when they encounter L1 borders
in a substrate (Burden-Gulley et al., 1995; Burden-Gulley and Lemmon,
1996). Consequently, the growth cone is well positioned as the major
site of action of L1, where in response to extracellular ligand
binding, L1 generates intracellular signals and interacts with the
cytoskeleton to regulate axonal growth (Kamiguchi and Lemmon,
1997).
Immunohistochemical analyses of developing mice demonstrated the
polarized expression of L1 in pyramidal cells, granule cells, and
interneurons in the hippocampus. L1 is expressed on axons but not on
dendrites or cell bodies of these cells (Persohn and Schachner, 1990).
Furthermore, Vogt et al. (1996) have shown that NgCAM, a chick homolog
of L1, is transported directly to the axonal growth cone of dorsal root
ganglia (DRG) neurons and inserted exclusively in the growth cone
membrane. Subsequently, NgCAM spreads from the growth cone into the
axonal shaft as the axon elongates by preferentially adding new
membrane components into the growth cone (Craig et al., 1995). As a
result, L1 is preferentially localized in the axonal growth cone and
axonal shaft. This raises the question as to how L1 is sorted into a
specific population of transport vesicles destined for the axonal
growth cone.
It is thought that many proteins carry sorting information encrypted
within their structure (Rothman and Wieland, 1996). In some cases, the
sorting information consists of structural motifs known as sorting
signals, which are recognized by intracellular receptor-like molecules.
Interaction of a sorting signal with its receptor results in sorting of
the signal-bearing protein into a specific population of vesicles
destined for a specific cellular compartment. A well known example is a
tyrosine-based sorting signal that conforms to the motif Yxx , where
x is any amino acid and is an amino acid with a bulky hydrophobic
side chain (Trowbridge et al., 1993). The tyrosine and residues are critical, whereas the importance of the x residues is less well defined
(Boll et al., 1996; Ohno et al., 1996). Various protein-sorting events
have been attributed to the tyrosine-based sorting signals, which
include clathrin-mediated endocytosis, targeting to endosomes en route
to the basolateral plasma membrane or to lysosomes, and localization to
the trans-Golgi network (Mellman, 1996; Marks et al.,
1997).
The neuron-specific alternatively spliced RSLE sequence in the L1
cytoplasmic domain is immediately preceded by a tyrosine residue. When
the RSLE sequence is spliced in, the resulting
YRSLE sequence conforms to the tyrosine-based
sorting motif. In this paper, we demonstrate that the YRSLE sequence is
critical for sorting L1 to the axonal growth cone, revealing a novel
function of the RSLE exon in neurons.
| |
MATERIALS AND METHODS |
Generation of wild-type and mutant L1 expression
vectors. The pBluescript II KS+, which contains a cDNA encoding
the full-length human L1 (L1FL) in its
EcoRI-HindIII site (Wong et al., 1995a), was
mutagenized to create mutant L1 cDNAs. A cDNA encoding for either an
RSLE-minus form of L1 (L1RSLE) or mutant L1 with a
Y1176A substitution (L1Y1176A) was generated by
oligonucleotide-directed mutagenesis according to the manufacturer's
protocol (Clontech Laboratories, Palo Alto, CA). For both mutations, a
selection primer (5'-CTCCACCGCGGTGGATGCCGCTCTAGAAC-3') was used to
mutate a unique NotI restriction site of pBluescript, which
is located upstream from the L1 insert. A mutagenic primer
(5'-GACCTTCGGCGAGTACAGTGACAACGAGGAG-3' or
5'-GACCTTCGGCGAGGCCAGGTCCCTGGAGAG-3') was used for
L1RSLE or L1Y1176A, respectively.
Both mutations were confirmed by dideoxy sequencing using Sequenase
v2.0 (United States Biochemicals, Cleveland, OH). A cDNA encoding for a
truncation mutant of L1 (L1C77) that lacks the
C-terminal 77 amino acids located immediately after the RSLE sequence
was created by PCR. Primers used for the PCR amplification are as
follows: a sense primer corresponding to nucleotides 2901-2918 of the
L1 cDNA; an antisense primer
(5'-CCAAGCTTACTTACCTACTCCAGGGACCTGTA-3') that contains a fragment
corresponding to 3427-3441 (Y1176RSLE) followed
immediately by a stop codon. The latter primer also has a
HindIII restriction site added to its 5'-end. The PCR product was digested with BsiWI (located at 3015) and
HindIII, and ligated into a
BsiWI/HindIII-digested pBluescript containing the
L1FL cDNA. The entire PCR-amplified region was confirmed by sequencing.
Subsequently, the L1 cDNAs were excised from the pBluescript with
EcoRI and XhoI, and subcloned into the
EcoRI/XhoI restriction site of pcDNA3
(Invitrogen, San Diego, CA). This placed the L1 cDNA under
transcriptional control of a cytomegalo virus enhancer-promoter. The
plasmid DNAs were purified using a Qiagen miniprep kit (Qiagen, Chatsworth, CA).
DNA transfection into NIH-3T3 cells and immunocytochemistry.
NIH-3T3 cells (American Type Culture Collection, Rockville, MD) were
plated on a two-chamber plastic slide (Lab-Tek, Naperville, IL) coated
with fibronectin (5 µg/cm2; Boehringer Mannheim,
Indianapolis, IN) at a density of ~0.5 × 105
cells/cm2 in minimum essential medium (MEM) (Life
Technologies, Gaithersburg, MD) supplemented with 10% fetal bovine
serum (FBS). After 16 hr, the cells were transfected with L1 expression
plasmids by DOTAP liposomal transfection according to the
manufacturer's protocol (Boehringer Mannheim). After an additional 24 hr incubation, the cells were processed for immunocytochemistry. For
living cell staining, the cells were incubated with monoclonal
anti-human L1 antibody (Wolff et al., 1988) for 1 hr at 4°C followed
by fixation with Bouin's fluid (75% saturated aqueous picric acid,
20% formalin, 5% glacial acetic acid) for 20 min. For fixed cell
staining, the cells were fixed with Bouin's fluid, washed with PBS,
blocked with 20% horse serum in PBS (HS-PBS), and incubated with
monoclonal anti-human L1 antibody for 16 hr at 4°C. In both staining
procedures, the cells were then incubated with Texas Red-conjugated
anti-mouse IgG (Molecular Probes, Eugene, OR) in HS-PBS (1:200) for 1 hr at 20°C, rinsed with PBS, mounted with SlowFade (Molecular
Probes), and observed using a Zeiss LSM 410 confocal laser scan
microscope (Carl Zeiss, Göttingen, Germany).
Primary cultures of DRG neurons and DNA transfection. DRGs
were dissected from embryonic day 10 chicks and dissociated
sequentially with 2.4 U/ml dispase II (Boehringer Mannheim) and 0.1 mg/ml DNase (Boehringer Mannheim) in
Ca2+/Mg2+-free PBS. The
dissociated cells were washed twice with MEM followed by incubation in
MEM supplemented with 5 µg/ml DOTAP lipofection reagent for 10 min.
Subsequently, 2.5 µg/ml of plasmid DNA was added to the medium, and
the cells were incubated for an additional 40 min on a culture dish.
Then 10% FBS and 20 ng/ml of nerve growth factor (NGF) were added, and
the cells that were detached by shaking the dish were replated on a
two-chamber plastic slide that was precoated sequentially with
poly-L-lysine and 10 µg/ml laminin (Life Technologies) at
an initial density of ~0.5 × 105
cells/cm2. The lipofection procedure described above
is based on the improved method for DOTAP-mediated gene transfer into
primary cultures of hippocampal neurons reported by Kaech et al.
(1996).
For DRG neuronal culture without DNA transfection, the dissociated
cells were resuspended in MEM supplemented with 10% FBS and were
preplated for 1 hr. Then 20 ng/ml of NGF was added to the medium, and
the neuron-enriched culture was prepared by replating the detached
cells on a poly-L-lysine and laminin-coated two-chamber slide at a density of ~0.5 × 104
cells/cm2.
The cultures were maintained in a humid atmosphere of 95% air, 5%
CO2 at 37°C.
Immunocytochemistry of DRG neurons. The DRG neurons were
fixed with Bouin's fluid at 14 or 24 hr after plating, washed with PBS, blocked with HS-PBS, and incubated with primary antibodies for 16 hr at 4°C. The primary antibodies used were monoclonal anti-human L1
(Wolff et al., 1988), polyclonal anti-chick NCAM (kind gift of Dr. Urs
Rutishauser, Case Western Reserve University, Cleveland OH), monoclonal
anti-microtubule-associated protein-2 (MAP-2) (5 µg/ml; Boehringer
Mannheim), and polyclonal anti-NgCAM antibodies (Lemmon and McLoon,
1986). After washes with PBS, the cells were incubated with Oregon
Green-conjugated anti-rabbit IgG (Molecular Probes) and Texas
Red-conjugated anti-mouse IgG in HS-PBS (1:200) for 1 hr at 20°C,
rinsed with PBS, and mounted with SlowFade.
In some experiments, cell-surface molecules were labeled by incubating
living cells with monoclonal anti-human L1 and polyclonal anti-chick
NCAM antibodies for 1 hr at 37°C. The cells were then fixed with 4%
formaldehyde in PBS, followed by incubation with secondary antibodies.
Human L1-transfected neurons that were double-stained for chick NCAM
and human L1 were observed using a Zeiss LSM 410 confocal laser scan
microscope with an oil immersion 100× objective. The whole area of a
Lab-Tek slide chamber was screened, and all the neurons expressing
human L1 were included in this study. The cells were scanned, and
fluorescence images were obtained in 0.5 µm steps in the
z-axis. A projected image was then generated from a series
of sectional images that encompassed the entire cell (10-20 µm in
the z-axis).
| |
RESULTS |
Generation of L1 mutants and expression in NIH-3T3 cells
The L1 cytoplasmic domain consists of 114 amino acids (1144-1257)
(Hlavin and Lemmon, 1991), and a part of the cytoplasmic domain
(1176-1185) is shown in the single-letter amino acid code (Fig.
1). In the wild-type full-length L1
(L1FL), a tyrosine residue at position 1176 is
adjacent to the RSLE sequence (shaded region), and the
resulting sequence of YRSLE conforms to a tyrosine-based sorting motif,
YxxL (underlined region). To characterize the structural significance of the YRSLE sequence for proper localization of L1 in
neurons, we have generated one non-neuronal isoform and two mutant
forms of L1. In the non-neuronal form of L1 lacking the RSLE sequence
(L1RSLE), a hydrophobic amino acid leucine at position
Y+3, which is a key residue in the motif, is replaced by a polar amino
acid Asn. L1Y1176A has a single amino acid substitution (Y1176A) that mutates a critical tyrosine residue in the motif. L1C77 is a truncation mutant that lacks the C-terminal
77 amino acids located immediately after the RSLE sequence.
Consequently, L1FL and L1C77 carry an amino
acid sequence that conforms to the tyrosine-based sorting motif;
however, this sequence is disrupted in L1RSLE and
L1Y1176A. We have generated pcDNA3-based expression
plasmids containing human L1 cDNA that codes for
L1FL, L1RSLE,
L1Y1176A, or L1C77. Both the
extracellular and transmembrane domains are identical among all the
forms of L1.
View larger version (17K):
[in this window]
[in a new window]
|
Figure 1.
Schematic illustration of the cytoplasmic domain
of wild-type and mutant forms of L1. A part of the L1 cytoplasmic
domain is shown in the single-letter amino acid code. In the
neuronal form of L1 (L1FL),
Tyr1176 is adjacent to the RSLE sequence
(shaded region). The resulting sequence of YRSLE
conforms to a tyrosine-based sorting motif, Yxx
(underlined region), where x is any amino acid and
is an amino acid with a bulky hydrophobic side chain. In the
non-neuronal form of L1 (L1RSLE) that lacks the RSLE
sequence, a hydrophobic amino acid leucine at position Y+3 is replaced
by a polar amino acid, Asn. L1Y1176A has a single amino
acid substitution (Y1176A) that mutates a critical tyrosine residue in
the motif. L1C77, which lacks the C-terminal 77 amino
acids located immediately after the RSLE exon, still carries the YRSLE
sequence.
|
|
To show that the L1 constructs are processed correctly and that the
gene products are integrated properly into the plasma membrane in
nonpolarized cells, NIH-3T3 cells were transfected with the L1
expression plasmids. Both living cells and fixed cells were
immunostained with monoclonal anti-human L1 antibody. Figure 2 shows single-section confocal images
(0.71-µm-thick) of the cells stained alive. L1FL,
L1RSLE, L1Y1176A and L1C77 are expressed on the cell surface with a similar pattern in both living
and fixed cells (data not shown), indicating that all the forms of L1
are integrated properly into the membrane in NIH-3T3 cells.
View larger version (99K):
[in this window]
[in a new window]
|
Figure 2.
Cell surface expression of wild-type and mutant
forms of L1 in NIH-3T3 cells. Immunocytochemical localization of
L1FL (A), L1RSLE
(B), L1Y1176A
(C), and L1C77
(D) in NIH-3T3 cells is shown. Live transfected
cells were stained with anti-human L1 antibody, and single-section
confocal images (0.71-µm-thick) were taken. All the forms of L1 are
expressed on the cell surface with a similar pattern in both living and
fixed cells (data not shown), indicating that they are correctly
processed and integrated into the plasma membrane. Scale bar, 25 µm.
|
|
The YRSLE sequence is required for sorting L1 to the axonal growth
cone in DRG neurons
To study axonal sorting of L1, we first characterized neurites
from cultured DRG neurons (axons vs dendrites). Peripheral sensory
neurons including DRG neurons are pseudounipolar in situ (Tandrup, 1995), and their somata lack dendrites (Pannese, 1981). An
axon of DRG neurons bifurcates shortly after it emerges from the cell
body: one branch extends peripherally, and the other branch extends
centrally. Immunocytochemical studies showed that neurites from
cultured DRG neurons express an axonal marker, L1, but not a dendritic
marker, MAP-2 (Letourneau and Shattuck, 1989). Koninck et al. (1993)
have shown that NGF induces sensory neurons from the nodose ganglia to
extend dendrites if the neurons are cultured in the absence of
satellite cells for a long term (1-3 weeks). To characterize neurites
from DRG neurons under the culture conditions in the present study, the
cells were double-labeled with polyclonal anti-NgCAM (a chick homolog
of L1) and monoclonal anti-MAP-2 antibodies (Fig.
3). All the neurites expressed NgCAM but
not MAP-2 in 322 DRG neurons examined after 14 hr in culture and in 346 neurons after 24 hr in culture. Similar results have been reported by
Honig and Kueter (1995) showing that all the growth cones of DRG
explants expressed NgCAM after 1.5 d in culture. These findings
indicate that DRG neurons used in the following experiments bear only
axons and lack dendrites.
View larger version (34K):
[in this window]
[in a new window]
|
Figure 3.
Immunocytochemical characterization of neurites
from cultured DRG neurons. Immunofluorescence micrographs of a chick
DRG neuron double-labeled with anti-NgCAM (A) and
anti-MAP-2 antibody (B) are shown. All the
neurites express an axonal marker, NgCAM, but the expression of a
dendritic marker, MAP-2, is restricted to the cell body. Scale bar, 50 µm.
|
|
To study the role of the YRSLE sequence in the localization of L1 in
neurons, primary cultures of chick DRG neurons were transfected with
pcDNA3 containing human L1 cDNA that codes for L1FL,
L1RSLE, L1Y1176A, or
L1C77. After 14 or 24 hr, expression of the transgene
was visualized by immunocytochemistry using a monoclonal anti-human L1
antibody that does not cross-react with NgCAM. The transfected cells
were double-labeled with polyclonal anti-chick NCAM antibody to
visualize the entire outline of neurons. An L1FL-expressing DRG neuron shown in Figure
4A,B
is representative of 11 cells from five independent experiments.
L1FL was highly expressed in the axonal growth cone and to
a lesser degree in the cell body, consistent with the observation that
L1 is preferentially transported to the axonal growth cone and inserted
exclusively into the growth cone membrane in DRG neurons (Vogt et al.,
1996). Similar results were obtained in all of the 11 cells, which
expressed L1FL (eight cells after 14 hr, three cells after
24 hr). These data also indicate that L1 can be synthesized from the
transgene and transported to the growth cone within 14 hr after
transfection. L1RSLE-expressing DRG neurons shown in
Figure 5 are representative of 23 cells
from four independent experiments (13 cells after 14 hr, 10 cells after 24 hr). Expression of L1RSLE was restricted to the cell
body, or in some neurons L1RSLE extended into the
proximal axonal shaft. None of the 23 neurons expressed
L1RSLE either in the distal half of the axonal shaft or
in the growth cone. The middle and bottom panels in Figure 5 show the
most extreme examples of neurons grown for 14 hr and 24 hr after
transfection, respectively, where L1RSLE extended
farthest into the axonal shaft. These data indicate that the RSLE
sequence has to be spliced in for L1 to reach the growth cone in DRG
neurons.
View larger version (74K):
[in this window]
[in a new window]
|
Figure 4.
Subcellular localization of L1FL
(A, B) and L1C77
(C, D) in DRG neurons. Projected confocal
images of human L1-expressing DRG neurons at 14 hr after transfection
are shown. Expression of L1FL or L1C77 was
visualized by monoclonal anti-human L1 antibody (B,
D). The cells were double-labeled with polyclonal
anti-chick NCAM antibody to visualize the entire outline of neurons
(A, C). Both L1FL and
L1C77 are highly expressed in the axonal growth cone and
to a lesser degree in the cell body. Scale bar, 25 µm.
|
|
View larger version (62K):
[in this window]
[in a new window]
|
Figure 5.
Subcellular localization of L1RSLE
in DRG neurons. Projected confocal images of
L1RSLE-expressing DRG neurons at 14 hr
(A-D) or 24 hr (E,
F) after transfection are shown. Expression of
L1RSLE was visualized by monoclonal anti-human L1
antibody (B, D, F).
The cells were double-labeled with polyclonal anti-chick NCAM antibody
to visualize the entire outline of neurons (A,
C, E). Expression of
L1RSLE is restricted to the cell body
(B), or L1RSLE extends into the
proximal axonal shaft but does not reach the distal half of the axon
and the growth cone (D, F). Scale
bar, 25 µm.
|
|
Because the RSLE sequence constitutes a tyrosine-based sorting motif in
the L1 cytoplasmic domain, we then analyzed whether a mutation of the
critical tyrosine residue in the motif influences the cellular
localization of L1. L1Y1176A-expressing DRG neurons shown
in Figure 6 are representative of 23 cells from four independent experiments (5 cells after 14 hr, 18 cells
after 24 hr). Similar to the localization pattern of
L1RSLE, expression of L1Y1176A was
restricted to the cell body and proximal axonal shaft, and never
extended into the distal axonal shaft or growth cone. This observation
indicates that Tyr1176 as well as the RSLE sequence
is critical for targeting L1 to the axonal growth cone.
L1C77, which carries the YRSLE sequence, consistently
reached the axonal growth cone in 19 of 20 DRG neurons from three
independent experiments (9 cells after 14 hr, 11 cells after 24 hr). A
representative L1C77-expressing cell is shown in Figure
4C,D. The localization pattern of
L1C77 was similar to that of L1FL,
with one exception in which L1C77 stayed in the cell
body and proximal axonal shaft.
View larger version (71K):
[in this window]
[in a new window]
|
Figure 6.
Subcellular localization of L1Y1176A
in DRG neurons. Projected confocal images of
L1Y1176A-expressing DRG neurons at 14 hr (A,
B) or 24 hr (C, D) after
transfection are shown. Expression of L1Y1176A was
visualized by monoclonal anti-human L1 antibody (B,
D). The cells were double-labeled with polyclonal
anti-chick NCAM antibody to visualize the entire outline of neurons
(A, C). Expression of
L1Y1176A is restricted to the cell body
(B), or L1Y1176A extends into the
proximal axonal shaft but does not reach the distal half of the axon
and the growth cone (D). Scale bar, 25 µm.
|
|
These observations are summarized in Table
1, where the subcellular localization of
transfected L1 in DRG neurons was categorized into four groups on the
basis of the extent of L1 expression along the axonal shaft. These
results demonstrate that the YRSLE sequence is required for targeting
L1 to the axonal growth cone, and that the tyrosine residue is critical
for the function of this signal.
The failure of L1RSLE and L1Y1176A to follow
the proper transport pathway raises a question as to whether these
forms of L1 are on the cell surface or trapped within intracellular vesicles. Living DRG neurons that express either L1RSLE
or L1Y1176A were incubated with the anti-human L1 antibody
before fixation, and confocal sections (0.83-µm-thick) through the
middle of the cell body were obtained (Fig.
7). Both forms of L1 mostly overlapped
with chick NCAM on the cell surface, as evidenced by a yellow color in
the superimposed images where chick NCAM is colored green and
transfected L1 is red. These observations indicate that these forms of
L1 are integrated into the plasma membrane in the somata after
missorting.
View larger version (100K):
[in this window]
[in a new window]
|
Figure 7.
Cell surface localization of L1RSLE
and L1Y1176A in the DRG somata. Confocal sections
(0.83-µm-thick) through the middle of the cell body of DRG neurons
that express L1RSLE
(A-C) or L1Y1176A
(D-F) are shown. The living cells
were double-labeled with polyclonal anti-chick NCAM (A,
D) and monoclonal anti-human L1 antibody
(B, E). Superimposition of the images
with chick NCAM colored in green and transfected L1 in
red (C, F) exhibits
clear overlap (yellow) on the cell surface. Scale
bar, 5 µm.
|
|
| |
DISCUSSION |
The interaction between cytoplasmic coat proteins and specific
signals in the cytoplasmic domains of integral membrane proteins is a
general mechanism controlling protein sorting. The best-characterized sorting signal is the tyrosine-based sorting motif that is involved in
various sorting events by interaction with adaptor proteins (Trowbridge
et al., 1993; Mellman, 1996; Marks et al., 1997). The YRSLE sequence
present in the cytoplasmic domain of the neuronal form of L1 conforms
to the tyrosine-based sorting motif. We have found that the AP-2
adaptor specifically recognizes and interacts with the YRSLE sequence,
resulting in clathrin-mediated endocytosis of L1 (H. Kamiguchi, K. E. Long, M. Pendergast, A. W. Schaefer, I. Rapoport, T. Kirchhausen, and V. Lemmon, unpublished observations). These
observations indicate that the YRSLE sequence of L1 actually functions
as a tyrosine-based sorting signal. The L1 cytoplasmic domain contains
three other tyrosine residues (Hlavin and Lemmon, 1991), although none
is situated three amino acids N-terminal to a hydrophobic residue and
therefore do not constitute a recognized sorting sequence. Homologs of
mammalian L1 in chick (NgCAM/8D9/G4) and zebrafish (L1.1 and L1.2)
contain the YRSLE sequence, but it is absent in Drosophila
(neuroglian) (Tongiorgi et al., 1995; Hortsch, 1996). In addition, it
is conserved in two other members of the L1 subfamily, neurofascin and
NrCAM (Kayyem et al., 1992; Volkmer et al., 1992), implying an
important function for this sequence.
The tyrosine-based sorting motif in the L1 cytoplasmic domain is
immediately followed by a cluster of acidic amino acids containing Ser1181 that can be phosphorylated by casein kinase
II (CKII) in vivo and in vitro (Fig. 1) (Wong et
al., 1996). Interestingly, an acidic stretch with CKII phosphorylation
sites is also located C-terminal to a tyrosine-based signal in several
proteins that are targeted to post-Golgi compartments, such as furin,
mannose 6-phosphate receptors, and varicella-zoster virus glycoprotein
I. Intracellular trafficking of these proteins is regulated by the CKII
phosphorylation sites as well as by the tyrosine-based sorting signal
(Trowbridge et al., 1993; Jones et al., 1995; Takahashi et al., 1995;
Alconada et al., 1996; Breuer et al., 1997), implying that the
phosphorylation of L1 by CKII might play a role in L1 trafficking.
Maintenance of spatial distribution of integral membrane proteins is an
essential function of polarized cells. This is evident in developing
neurons in which the production of specific cell-surface domains is
necessary to explore guidance cues and coordinate axonal growth, to
establish appropriate connections with their targets, and to transmit
or receive both chemical and electrical information. Because L1
expressed in the growth cone plays a crucial role in regulating axonal
growth, it is important for developing neurons to maintain the
polarized expression of L1 by preferentially targeting both newly
synthesized and recycled L1 molecules to the axonal growth cone. The
present study demonstrates that the YRSLE sequence of the L1
cytoplasmic domain functions as a tyrosine-based sorting signal that is
required for sorting L1 to the axonal growth cone. This result strongly
suggests that neurons express an intracellular adaptor-like molecule
that specifically recognizes and interacts with the YRSLE sequence,
resulting in sorting L1 into a specific population of vesicles destined
for the axonal growth cone. Functional interactions between
tyrosine-based sorting signals and adaptor proteins depend on multiple
factors, such as the exact position of the signal in the cytoplasmic
tail (Rohrer et al., 1996), amino acids surrounding the critical
tyrosine (Ohno et al., 1996), and possible secondary signals that
operate in concert (Rohrer et al., 1995; Schweizer et al., 1996).
Therefore, although the YRSLE sequence is essential for sorting to
axons, it is likely that additional features of the L1 cytoplasmic
domain N-terminal to this sequence are also important for targeting to
the axon.
The mechanism of protein sorting is well characterized in polarized
epithelial cells (Matter and Mellman, 1994; Mellman, 1996). The
cytoplasmic domain of some integral membrane proteins contains tyrosine-based sorting signals that mediate sorting to the basolateral plasma membrane. The AP-1 adaptor specifically localizes to coated buds
and vesicles of the trans-Golgi network, the primary sorting site of newly synthesized proteins, and mediates vesicular transport to
the basolateral plasma membrane. Accumulating evidence has suggested
that epithelial apical and neuronal axonal membranes, as well as
epithelial basolateral and neuronal somatodendritic membranes, may be
equivalent domains in terms of polarized protein localization (Craig
and Banker, 1994; Scannevin et al., 1996). This has led to the proposal
that epithelial cells and neurons may share common molecular mechanisms
for protein sorting. However, several membrane proteins that contradict
this hypothesis have also been reported (Craig and Banker, 1994;
Higgins et al., 1997). Given the existence of nervous tissue-specific
isoforms of adaptor complex subunits (Pevsner et al., 1994; Ball et
al., 1995), it is most likely that neurons and epithelial cells use
similar but distinct molecules to govern protein sorting. To further
clarify the mechanism of protein sorting in neurons, it is essential to identify molecules that interact with tyrosine-based sorting signals and mediate axonal sorting.
Interestingly, expression of the YRSLE sequence depends on the
neuron-specific alternatively spliced exon 27 that codes for RSLE. In
the RSLE-minus form of L1 that is expressed exclusively in non-neuronal
cells, the critical leucine residue at position Y+3 is replaced by a
polar amino acid, Asn, resulting in disruption of the sorting signal.
Functional differences between the RSLE-positive and RSLE-negative
forms of L1 have been studied using L1-transfected L cells (Takeda et
al., 1996). The cells expressing either form of L1 showed homophilic
adhesivity and promoted neurite growth and neuronal migration to a
similar extent, but the RSLE-positive L1-expressing cells migrated
faster than the other cell line on an L1 substrate. However, the
question regarding the function of the RSLE exon in neurons remained
unanswered. Our findings showed that the RSLE exon regulates sorting
into the axon. Non-neuronal cells that express the RSLE-minus form of
L1, such as Schwann cells, melanocytes, and lymphocytes, do not require
the YRSLE sequence as a sorting signal, probably because these cells
are not highly polarized.
Another possible function of the YRSLE sequence in neurons is that it
may interact with Src homology 2 (SH2)-containing signaling proteins
(Songyang and Cantley, 1995). Indeed, it has been demonstrated that the
cytoplasmic domain of L1 is phosphorylated on tyrosine, possibly on
Try1176 (Heiland et al., 1996). Because the tyrosine
in the sorting motif has to be in a nonphosphorylated state for the
signal to be active (Boll et al., 1996; Ohno et al., 1996), the YRSLE
sequence might have dual roles (sorting signal or SH2 binding)
depending on the phosphorylation state of the tyrosine.
In humans, mutations of the L1 gene cause hydrocephalus as well as
disruptions of the formation of major axonal tracts such as the
corticospinal tract and corpus callosum. Patients with L1 mutations
almost always have spastic paraparesis and adducted thumbs, which are
caused by impaired formation of the corticospinal tract and loss of
innervation to the extensor pollicis muscle, respectively (Wong et al.,
1995b; Kamiguchi et al., 1998). Interestingly, mutations that affect
only the L1 cytoplasmic domain are much less likely to cause severe
hydrocephalus than mutations affecting the extracellular domain
(Yamasaki et al., 1997). However, mutations of the cytoplasmic domain
are just as likely to produce abnormal development of axonal pathways
as mutations of the extracellular domain. Because deletion of the
cytoplasmic domain from L1 does not alter its homophilic adhesive
property (Wong et al., 1995a), the cytoplasmic domain must serve other
important roles that are critical in axonal tract development. This is
likely to include activation of second messenger systems and
interactions with the cytoskeleton (Kamiguchi and Lemmon, 1997).
Mutations of the cytoplasmic domain must impair one or more of these L1
functions, resulting in abnormal development of major axon tracts.
However, it is possible that mutations affecting the YRSLE sorting
signal may also prevent L1 from being expressed in axons. So far, two
different mutations that disrupt the YRSLE sequence in human L1 have
been published: a frameshift mutation in exon 26 (Jouet et al., 1994)
and a point mutation in the acceptor site in intron 26 that would
truncate the protein immediately before the RSLE sequence (Jouet et
al., 1995). Both mutations cause spastic paraparesis and adducted
thumbs, but neither causes progressive hydrocephalus or death at an
early age. In the affected individuals, loss of L1 expression in axons probably accounts for the malformation of axonal tracts.
| |
FOOTNOTES |
Received Dec. 29, 1997; revised March 3, 1998; accepted March 5, 1998.
This work was supported by National Institutes of Health Grants EY-5285
and NS-34252 to V.L. We acknowledge the excellent technical assistance
of Guanghui Cheng, Maryanne Pendergast, and Zhenhua Miao. The
anti-human L1 antibody (5G3) was the kind gift of Dr. Ralph Reisfeld.
We also thank Dr. Martin Snider and Dr. Susan Burden-Gulley for helpful
comments on this manuscript.
Correspondence should be addressed to Vance Lemmon, Department of
Neurosciences, Case Western Reserve University, 2109 Adelbert Road,
Cleveland, OH 44106-4975.
| |
REFERENCES |
-
Alconada A,
Bauer U,
Hoflack B
(1996)
A tyrosine-based motif and a casein kinase II phosphorylation site regulate the intracellular trafficking of the varicella-zoster virus glycoprotein I, a protein localized in the trans-Golgi network.
EMBO J
15:6096-6110[Web of Science][Medline].
-
Ball CL,
Hunt SP,
Robinson MS
(1995)
Expression and localization of
 -adaptin isoforms.
J Cell Sci
108:2865-2875[Abstract]. -
Barami K,
Kirschenbaum B,
Lemmon V,
Goldman SA
(1994)
N-cadherin and Ng-CAM/8D9 are involved serially in the migration of newly generated neurons into the adult songbird brain.
Neuron
13:567-582[Web of Science][Medline].
-
Boll W,
Ohno H,
Songyang Z,
Rapoport I,
Cantley LC,
Bonifacino JS,
Kirchhausen T
(1996)
Sequence requirements for the recognition of tyrosine-based endocytic signals by clathrin AP-2 complexes.
EMBO J
15:5789-5795[Web of Science][Medline].
-
Breuer P,
Körner C,
Böker C,
Herzog A,
Pohlmann R,
Braulke T
(1997)
Serine phosphorylation site of the 46 kDa mannose 6-phosphate receptor is required for transport to the plasma membrane in Madin-Darby canine kidney and mouse fibroblast cells.
Mol Biol Cell
8:567-576[Abstract].
-
Burden-Gulley SM,
Lemmon V
(1996)
L1/8D9, N-cadherin and laminin induce distinct distribution patterns of cytoskeletal elements in growth cones.
Cell Motil Cytoskel
35:1-23[Web of Science][Medline].
-
Burden-Gulley SM,
Payne HR,
Lemmon V
(1995)
Growth cones are actively influenced by substrate-bound adhesion molecules.
J Neurosci
15:4370-4381[Abstract].
-
Cohen NR,
Taylor JSH,
Scott LB,
Guillery RW,
Soriano P,
Furley AJW
(1997)
Errors in corticospinal axon guidance in mice lacking the neural cell adhesion molecule L1.
Curr Biol
8:26-33.
-
Craig AM,
Banker G
(1994)
Neuronal polarity.
Annu Rev Neurosci
17:267-310[Web of Science][Medline].
-
Craig AM,
Wyborski RJ,
Banker G
(1995)
Preferential addition of newly synthesized membrane protein at axonal growth cones.
Nature
375:592-594[Medline].
-
Dahme M,
Bartsch U,
Martini R,
Anliker B,
Schachner M,
Mantei N
(1997)
Disruption of the mouse L1 gene leads to malformations of the nervous system.
Nat Genet
17:346-349[Web of Science][Medline].
-
Grumet M,
Edelman GM
(1988)
Neuron-glia cell adhesion molecule interacts with neurons and astroglia via different binding mechanisms.
J Cell Biol
106:487-503[Abstract/Free Full Text].
-
Heiland PC,
Hertlein B,
Traub O,
Griffith LS,
Schmitz B
(1996)
The neural adhesion molecule L1 is phosphorylated on tyrosine and serine residues.
NeuroReport
7:2675-2678[Web of Science][Medline].
-
Higgins D,
Burack M,
Lein P,
Banker G
(1997)
Mechanisms of neuronal polarity.
Curr Opin Neurobiol
7:599-604[Web of Science][Medline].
-
Hlavin ML,
Lemmon V
(1991)
Molecular structure and functional testing of human L1CAM: an interspecies comparison.
Genomics
11:416-423[Web of Science][Medline].
-
Honig MG,
Kueter J
(1995)
The expression of cell adhesion molecules on the growth cones of chick cutaneous and muscle sensory neurons.
Dev Biol
167:563-583[Web of Science][Medline].
-
Hortsch M
(1996)
The L1 family of neural cell adhesion molecules: old proteins performing new tricks.
Neuron
17:587-593[Web of Science][Medline].
-
Jones BG,
Thomas L,
Molloy SS,
Thulin CD,
Fry MD,
Walsh KA,
Thomas G
(1995)
Intracellular trafficking of furin is modulated by the phosphorylation state of a casein kinase II site in its cytoplasmic tail.
EMBO J
14:5869-5883[Web of Science][Medline].
-
Jouet M,
Rosenthal A,
Armstrong G,
MacFarlane J,
Stevenson R,
Paterson J,
Metzenberg A,
Ionasescu V,
Temple K,
Kenwrick S
(1994)
X-linked spastic paraplegia (SPG1), MASA syndrome and X-linked hydrocephalus result from mutations in the L1 gene.
Nat Genet
7:402-407[Web of Science][Medline].
-
Jouet M,
Moncla A,
Paterson J,
Mckeown C,
Fryer A,
Carpenter N,
Holmberg E,
Wadelius C,
Kenwrick S
(1995)
New domains of neural cell-adhesion molecule L1 implicated in X-linked hydrocephalus and MASA syndrome.
Am J Hum Genet
56:1304-1314[Web of Science][Medline].
-
Kaech S,
Kim JB,
Cariola M,
Ralston E
(1996)
Improved lipid-mediated gene transfer into primary cultures of hippocampal neurons.
Mol Brain Res
35:344-348[Medline].
-
Kamiguchi H,
Lemmon V
(1997)
Neural cell adhesion molecule L1: signaling pathways and growth cone motility.
J Neurosci Res
49:1-8[Web of Science][Medline].
-
Kamiguchi H,
Hlavin ML,
Yamasaki M,
Lemmon V
(1998)
Adhesion molecules and inherited diseases of the human nervous system.
Annu Rev Neurosci
21:97-125[Web of Science][Medline].
-
Kayyem JF,
Roman JM,
de la Rosa EJ,
Schwarz U,
Dreyer WJ
(1992)
Bravo/Nr-CAM is closely related to the cell adhesion molecules L1 and Ng-CAM and has a similar heterodimer structure.
J Cell Biol
118:1259-1270[Abstract/Free Full Text].
-
Koninck PD,
Carbonetto S,
Cooper E
(1993)
NGF induces neonatal rat sensory neurons to extend dendrites in culture after removal of satellite cells.
J Neurosci
13:577-585[Abstract].
-
Lagenaur C,
Lemmon V
(1987)
An L1-like molecule, the 8D9 antigen, is a potent substrate for neurite extension.
Proc Natl Acad Sci USA
84:7753-7757[Abstract/Free Full Text].
-
Lemmon V,
McLoon S
(1986)
The appearance of an L1-like molecule in the chick primary visual pathway.
J Neurosci
6:2987-2994[Abstract].
-
Lemmon V,
Farr K,
Lagenaur C
(1989)
L1-mediated axon outgrowth occurs via a homophilic binding mechanism.
Neuron
2:1597-1603[Web of Science][Medline].
-
Letourneau PC,
Shattuck TA
(1989)
Distribution and possible interactions of actin-associated proteins and adhesion molecules of nerve growth cones.
Development
105:505-519[Abstract/Free Full Text].
-
Lindner J,
Rathjen FG,
Schachner M
(1983)
L1 mono- and polyclonal antibodies modify cell migration in early postnatal mouse cerebellum.
Nature
305:427-430[Medline].
-
Marks MS,
Ohno H,
Kirchhausen T,
Bonifacino JS
(1997)
Protein sorting by tyrosine-based signals: adapting to the Ys and wherefores.
Trends Cell Biol
7:124-128.[Medline]
-
Matter K,
Mellman I
(1994)
Mechanisms of cell polarity: sorting and transport in epithelial cells.
Curr Opin Cell Biol
6:545-554[Web of Science][Medline].
-
Mellman I
(1996)
Endocytosis and molecular sorting.
Annu Rev Cell Dev Biol
12:575-625.[Web of Science][Medline]
-
Miura M,
Kobayashi M,
Asou H,
Uyemura K
(1991)
Molecular cloning of cDNA encoding the rat neural cell adhesion molecule L1
 two L1 isoforms in the cytoplasmic region are produced by differential splicing.
FEBS Lett
289:91-95[Web of Science][Medline]. -
Moos M,
Tacke R,
Scherer H,
Teplow D,
Fruh K,
Schachner M
(1988)
Neural adhesion molecule L1 as a member of the immunoglobulin superfamily with binding domains similar to fibronectin.
Nature
334:701-703[Medline].
-
Ohno H,
Fournier MC,
Poy G,
Bonifacino JS
(1996)
Structural determinants of interaction of tyrosine-based sorting signals with the adaptor medium chains.
J Biol Chem
271:29009-29015[Abstract/Free Full Text].
-
Pannese E
(1981)
The satellite cells of the sensory ganglion.
Adv Anat Embryol Cell Biol
65:1-111[Medline].
-
Persohn E,
Schachner M
(1990)
Immunohistological localization of the neural adhesion molecules L1 and N-CAM in the developing hippocampus of the mouse.
J Neurocytol
19:807-819[Web of Science][Medline].
-
Pevsner J,
Volknandt W,
Wong BR,
Scheller RH
(1994)
Two rat homologs of clathrin-associated adaptor proteins.
Gene
146:279-283[Web of Science][Medline].
-
Reid RA,
Hemperly JJ
(1992)
Variants of human L1 cell adhesion molecule arise through alternate splicing of RNA.
J Mol Neurosci
3:127-135[Web of Science][Medline].
-
Rohrer J,
Schweizer A,
Johnson KF,
Kornfeld S
(1995)
A determinant in the cytoplasmic tail of the cation-dependent mannose 6-phosphate receptor prevents trafficking to lysosomes.
J Cell Biol
130:1297-1306[Abstract/Free Full Text].
-
Rohrer J,
Schweizer A,
Russell D,
Kornfeld S
(1996)
The targeting of Lamp1 to lysosomes is dependent on the spacing of its cytoplasmic tail tyrosine sorting motif relative to the membrane.
J Cell Biol
132:565-576[Abstract/Free Full Text].
-
Rothman JE,
Wieland FT
(1996)
Protein sorting by transport vesicles.
Science
272:227-234[Abstract].
-
Scannevin RH,
Murakoshi H,
Rhodes KJ,
Trimmer JS
(1996)
Identification of a cytoplasmic domain important in the polarized expression and clustering of the Kv2.1 K+ channel.
J Cell Biol
135:1619-1632[Abstract/Free Full Text].
-
Schweizer A,
Kornfeld S,
Rohrer J
(1996)
Cysteine34 of the cytoplasmic tail of the cation-dependent mannose 6-phosphate receptor is reversibly palmitoylated and required for normal trafficking and lysosomal enzyme sorting.
J Cell Biol
132:577-584[Abstract/Free Full Text].
-
Songyang Z,
Cantley LC
(1995)
Recognition and specificity in protein tyrosine kinase-mediated signaling.
Trends Biol Sci
20:470-475.
-
Stallcup WB,
Beasley L
(1985)
Involvement of the nerve growth factor-inducible large external glycoprotein (NILE) in neurite fasciculation in primary cultures of rat brain.
Proc Natl Acad Sci USA
82:1276-1280[Abstract/Free Full Text].
-
Takahashi S,
Nakagawa T,
Bonno T,
Watanabe T,
Murakami K,
Nakayama K
(1995)
Localization of furin to the trans-Golgi network and recycling from the cell surface involves Ser and Tyr residues within the cytoplasmic domain.
J Biol Chem
270:28397-28401[Abstract/Free Full Text].
-
Takeda Y,
Asou H,
Murakami Y,
Miura M,
Kobayashi M,
Uyemura K
(1996)
A nonneuronal isoform of cell adhesion molecule L1: tissue-specific expression and functional analysis.
J Neurochem
66:2338-2349[Web of Science][Medline].
-
Tandrup T
(1995)
Are the neurons in the dorsal root ganglion pseudounipolar? A comparison of the number of neurons and number of myelinated and unmyelinated fibres in the dorsal root.
J Comp Neurol
357:341-347[Web of Science][Medline].
-
Tongiorgi E,
Bernhardt RR,
Schachner M
(1995)
Zebrafish neurons express two L1-related molecules during early axonogenesis.
J Neurosci Res
42:547-561[Web of Science][Medline].
-
Trowbridge IS,
Collawn JF,
Hopkins CR
(1993)
Signal-dependent membrane protein trafficking in the endocytic pathway.
Annu Rev Cell Biol
9:129-161[Web of Science].
-
Vogt L,
Giger RJ,
Ziegler U,
Kunz B,
Buchstaller A,
Hermens WTJMC,
Kaplitt MG,
Rosenfeld MR,
Plaff DW,
Verhaagen J,
Sonderegger P
(1996)
Continuous renewal of the axonal pathway sensor apparatus by insertion of new sensor molecules into the growth cone membrane.
Curr Biol
6:1153-1158[Web of Science][Medline].
-
Volkmer H,
Hassel B,
Wolff JM,
Frank R,
Rathjen FG
(1992)
Structure of the axonal surface recognition molecule neurofascin and its relationship to a neural subgroup of the immunoglobulin superfamily.
J Cell Biol
118:149-161[Abstract/Free Full Text].
-
Wolff JM,
Frank R,
Mujoo K,
Spiro RC,
Reisfeld RA,
Rathjen FG
(1988)
A human brain glycoprotein related to the mouse cell adhesion molecule L1.
J Biol Chem
263:11943-11947[Abstract/Free Full Text].
-
Wong EV,
Cheng G,
Payne HR,
Lemmon V
(1995a)
The cytoplasmic domain of the cell adhesion molecule L1 is not required for homophilic adhesion.
Neurosci Lett
200:155-158[Web of Science][Medline].
-
Wong EV,
Kenwrick S,
Willems P,
Lemmon V
(1995b)
Mutations in the cell adhesion molecule L1 cause mental retardation.
Trends Neurosci
18:168-172[Web of Science][Medline].
-
Wong EV,
Schaefer AW,
Landreth G,
Lemmon V
(1996)
Casein kinase II phosphorylates the neural cell adhesion molecule L1.
J Neurochem
66:779-786[Web of Science][Medline].
-
Yamasaki M,
Thompson P,
Lemmon V
(1997)
CRASH syndrome: mutations in L1CAM correlate with severity of the disease.
Neuropediatrics
28:175-178[Web of Science][Medline].
Copyright © 1998 Society for Neuroscience 0270-6474/98/18103749-08$05.00/0
This article has been cited by other articles:
|
|
|
|
 
C. Bel, K. Oguievetskaia, C. Pitaval, L. Goutebroze, and C. Faivre-Sarrailh
Axonal targeting of Caspr2 in hippocampal neurons via selective somatodendritic endocytosis
J. Cell Sci.,
September 15, 2009;
122(18):
3403 - 3413.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. Thelen, T. Georg, S. Bertuch, P. Zelina, and G. E. Pollerberg
Ubiquitination and Endocytosis of Cell Adhesion Molecule DM-GRASP Regulate Its Cell Surface Presence and Affect Its Role for Axon Navigation
J. Biol. Chem.,
November 21, 2008;
283(47):
32792 - 32801.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. C. Yap, R. L. Nokes, D. Wisco, E. Anderson, H. Folsch, and B. Winckler
Pathway selection to the axon depends on multiple targeting signals in NgCAM
J. Cell Sci.,
May 1, 2008;
121(9):
1514 - 1525.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. C. Yap, D. Wisco, P. Kujala, Z. M. Lasiecka, J. T. Cannon, M. C. Chang, H. Hirling, J. Klumperman, and B. Winckler
The somatodendritic endosomal regulator NEEP21 facilitates axonal targeting of L1/NgCAM
J. Cell Biol.,
February 25, 2008;
180(4):
827 - 842.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. Diestel, D. Schaefer, H. Cremer, and B. Schmitz
NCAM is ubiquitylated, endocytosed and recycled in neurons
J. Cell Sci.,
November 15, 2007;
120(22):
4035 - 4049.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
Y. Nasu-Nishimura, T. Hayashi, T. Ohishi, T. Okabe, S. Ohwada, Y. Hasegawa, T. Senda, C. Toyoshima, T. Nakamura, and T. Akiyama
Role of the Rho GTPase-activating protein RICS in neurite outgrowth
Genes Cells,
June 1, 2006;
11(6):
607 - 614.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
P. Gutwein, A. Stoeck, S. Riedle, D. Gast, S. Runz, T. P. Condon, A. Marme, M.-C. Phong, O. Linderkamp, A. Skorokhod, et al.
Cleavage of L1 in Exosomes and Apoptotic Membrane Vesicles Released from Ovarian Carcinoma Cells
Clin. Cancer Res.,
April 1, 2005;
11(7):
2492 - 2501.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
L. Cheng, K. Itoh, and V. Lemmon
L1-Mediated Branching Is Regulated by Two Ezrin-Radixin-Moesin (ERM)-Binding Sites, the RSLE Region and a Novel Juxtamembrane ERM-Binding Region
J. Neurosci.,
January 12, 2005;
25(2):
395 - 403.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. D. Malhotra, V. Thyagarajan, C. Chen, and L. L. Isom
Tyrosine-phosphorylated and Nonphosphorylated Sodium Channel {beta}1 Subunits Are Differentially Localized in Cardiac Myocytes
J. Biol. Chem.,
September 24, 2004;
279(39):
40748 - 40754.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. Nishimura, F. Yoshihara, T. Tojima, N. Ooashi, W. Yoon, K. Mikoshiba, V. Bennett, and H. Kamiguchi
L1-dependent neuritogenesis involves ankyrinB that mediates L1-CAM coupling with retrograde actin flow
J. Cell Biol.,
December 8, 2003;
163(5):
1077 - 1088.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
D. Wisco, E. D. Anderson, M. C. Chang, C. Norden, T. Boiko, H. Folsch, and B. Winckler
Uncovering multiple axonal targeting pathways in hippocampal neurons
J. Cell Biol.,
September 29, 2003;
162(7):
1317 - 1328.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. F. Wilkemeyer, C. E. Menkari, and M. E. Charness
Novel Antagonists of Alcohol Inhibition of L1-Mediated Cell Adhesion: Multiple Mechanisms of Action
Mol. Pharmacol.,
November 1, 2002;
62(5):
1053 - 1060.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
T. C. Dickson, C. D. Mintz, D. L. Benson, and S. R.J. Salton
Functional binding interaction identified between the axonal CAM L1 and members of the ERM family
J. Cell Biol.,
June 24, 2002;
157(7):
1105 - 1112.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. Thelen, V. Kedar, A. K. Panicker, R.-S. Schmid, B. R. Midkiff, and P. F. Maness
The Neural Cell Adhesion Molecule L1 Potentiates Integrin-Dependent Cell Migration to Extracellular Matrix Proteins
J. Neurosci.,
June 15, 2002;
22(12):
4918 - 4931.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
G. A. Smith, S. P. Gross, and L. W. Enquist
Herpesviruses use bidirectional fast-axonal transport to spread in sensory neurons
PNAS,
March 13, 2001;
98(6):
3466 - 3470.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
L. K. Needham, K. Thelen, and P. F. Maness
Cytoplasmic Domain Mutations of the L1 Cell Adhesion Molecule Reduce L1-Ankyrin Interactions
J. Neurosci.,
March 1, 2001;
21(5):
1490 - 1500.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S Alais, N Allioli, C Pujades, J. Duband, O Vainio, B. Imhof, and D Dunon
HEMCAM/CD146 downregulates cell surface expression of (&bgr;)1 integrins
J. Cell Sci.,
January 5, 2001;
114(10):
1847 - 1859.
[Abstract]
[PDF]
|
|
|
|
|
|
|
J. M. Vila, J. Calvo, L. Places, O. Padilla, M. Arman, I. Gimferrer, C. Aussel, J. Vives, and F. Lozano
Role of Two Conserved Cytoplasmic Threonine Residues (T410 and T412) in CD5 Signaling
J. Immunol.,
January 1, 2001;
166(1):
396 - 402.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R.-S. Schmid, W. M. Pruitt, and P. F. Maness
A MAP Kinase-Signaling Pathway Mediates Neurite Outgrowth on L1 and Requires Src-Dependent Endocytosis
J. Neurosci.,
June 1, 2000;
20(11):
4177 - 4188.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
H. Kamiguchi and V. Lemmon
Recycling of the Cell Adhesion Molecule L1 in Axonal Growth Cones
J. Neurosci.,
May 15, 2000;
20(10):
3676 - 3686.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
D. Peretti, L. Peris, S. Rosso, S. Quiroga, and A. Caceres
Evidence for the Involvement of Kif4 in the Anterograde Transport of L1-Containing Vesicles
J. Cell Biol.,
April 3, 2000;
149(1):
141 - 152.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. Kenwrick, A. Watkins, and E. D. Angelis
Neural cell recognition molecule L1: relating biological complexity to human disease mutations
Hum. Mol. Genet.,
April 1, 2000;
9(6):
879 - 886.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. W. Schaefer, H. Kamiguchi, E. V. Wong, C. M. Beach, G. Landreth, and V. Lemmon
Activation of the MAPK Signal Cascade by the Neural Cell Adhesion Molecule L1 Requires L1 Internalization
J. Biol. Chem.,
December 31, 1999;
274(53):
37965 - 37973.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C.A. Haney, Z. Sahenk, C. Li, V.P. Lemmon, J. Roder, and B.D. Trapp
Heterophilic Binding of L1 on Unmyelinated Sensory Axons Mediates Schwann Cell Adhesion and Is Required for Axonal Survival
J. Cell Biol.,
September 6, 1999;
146(5):
1173 - 1184.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. F. Bearer, A. R. Swick, M. A. O'Riordan, and G. Cheng
Ethanol inhibits L1-mediated neurite outgrowth in postnatal rat cerebellar granule cells.
J. Biol. Chem.,
July 9, 1999;
274(28):
20046 - 20046.
[Full Text]
[PDF]
|
|
|
|
|
|
|
H. Kamiguchi, K. E. Long, M. Pendergast, A. W. Schaefer, I. Rapoport, T. Kirchhausen, and V. Lemmon
The Neural Cell Adhesion Molecule L1 Interacts with the AP-2 Adaptor and Is Endocytosed via the Clathrin-Mediated Pathway
J. Neurosci.,
July 15, 1998;
18(14):
5311 - 5321.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
K. E. Long, H. Asou, M. D. Snider, and V. Lemmon
The Role of Endocytosis in Regulating L1-mediated Adhesion
J. Biol. Chem.,
January 5, 2001;
276(2):
1285 - 1290.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. Koroll, F. G. Rathjen, and H. Volkmer
The Neural Cell Recognition Molecule Neurofascin Interacts with Syntenin-1 but Not with Syntenin-2, Both of Which Reveal Self-associating Activity
J. Biol. Chem.,
March 30, 2001;
276(14):
10646 - 10654.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
E. De Angelis, T. Brummendorf, L. Cheng, V. Lemmon, and S. Kenwrick
Alternative Use of a Mini Exon of the L1 Gene Affects L1 Binding to Neural Ligands
J. Biol. Chem.,
August 24, 2001;
276(35):
32738 - 32742.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
Top
Abstract
Introduction
