The Pancreas Center, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts 02215
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
We have shown that endocytosis at the apical plasma membrane of pancreatic acinar cells is regulated by the pH of the acinar lumen and is associated with cleavage of GP2, a glycosyl phosphatidylinositol-anchored protein. The aim of this study was to determine the transduction pathway by which endocytosis is activated. Apical endocytosis was studied in rat pancreatic acini by prestimulation with cholecystokinin followed by measurement of horseradish peroxidase (HRP) uptake. Lanthanum, staurosporine, and forskolin had no effect on HRP uptake. Cytochalasin D significantly inhibited endocytosis, indicating a dependence on actin filament integrity. Genistein and the specific tyrphostin inhibitor B42 also inhibited HRP uptake, implicating tyrosine kinases in the regulation of HRP uptake. With the use of an Src kinase-specific substrate, Src kinase activity was temporally related to activation of endocytosis. The tyrosine-dependent phosphorylation of an 85-kDa substrate in both rat and mouse pancreatic acini correlated with Src kinase activation and pH-dependent regulation of HRP uptake. These results indicate that apical endocytosis in acinar cells is associated with tyrosine kinase activation and is dependent on the actin cytoskeleton.
acini; pancreas
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
IN THE PANCREATIC ACINAR CELL, hormonal stimulation results in the insertion of large amounts of secretory granule membrane into the apical plasma membrane (APM). This membrane is postulated to remain as discrete lipid patches (6), which are then recycled back to the Golgi for reuse in the regulated secretory pathway. We have recently demonstrated that this apical endocytic process is regulated by the pH of the acinar lumen (10, 11). Specifically, endocytosis of horseradish peroxidase (HRP) was inhibited at pH values of <6.2, whereas maximal uptake occurred at pH values of 7.4-8.3. The activation of HRP uptake was associated with the cleavage of GP2, a glycosyl phosphatidylinositol (GPI)-anchored protein from the APM (12).
Endocytosis can occur at the basolateral or apical domain, potentially through different mechanisms. In renal proximal tubule cells, apical endocytosis of the ANG II receptor was dependent on microfilament and microtubule integrity, as evidenced by inhibition with cytochalasin D or colchicine, respectively (20). Similarly, actin microfilaments were found to be critical for endocytosis at the apical but not the basolateral surface of Madin-Darby canine kidney (MDCK) cells (13) as well as in Caco-2 cells (23).
Studies utilizing phorbol myristate have indicated that calcium signaling via protein kinase C is important in the regulation of apical endocytosis in both Caco-2 and MDCK cells (16, 23). In addition, activation of heterotrimeric GTP-binding proteins with mastoparan or increases in intracellular cAMP levels promoted the internalization of ricin at the apical surface of MDCK cells through a clathrin-independent pathway (8). In lymphocytes, modulation of protein kinase C or protein kinase A affected internalization of CD 59, a GPI-anchored protein (7). Taken together, these studies demonstrate that, in addition to the cytoskeletal matrix, signaling through cAMP and/or calcium pathways may play an important role in apical endocytic pathways.
Tyrosine kinases have also been implicated in endocytosis. Caveolin, an integral membrane component of caveolae, undergoes tyrosine-dependent phosphorylation both in vitro and in vivo in v-Src-transformed NIH/3T3 cells (17). Caveolae are important in potocytosis and may activate transmembrane signaling through their association with GPI-anchored proteins (1). Endocytosis of the asialoglycoprotein receptor in hepatocytes is dependent on tyrosine-induced phosphorylation (9, 15). Inhibiting phosphorylation of the receptor with genistein or tyrphostin blocks endocytosis at an early step. However, there are no data on whether nonreceptor tyrosine-dependent kinases are involved in apical endocytic events.
Given that endocytosis can be selectively activated by cleavage of GP2 from the APM, we sought to determine the signal transduction mechanism by which this process is activated in rat pancreas. The data indicate that apical endocytosis is regulated through a tyrosine kinase-dependent pathway.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
HRP, cytochalasin D, lanthanum, forskolin, and sodium orthovanadate were obtained from Sigma Chemical (St. Louis, MO); cholecystokinin (CCK-8) was from Penninsula Laboratories (Belmont, CA); genistein was from Research Biochemicals International (Natick, MA); tyrphostin A1 and B42 and staurosporine were from Calbiochem (La Jolla, CA); protein tyrosine kinase (Src family) assay kit, phosphotyrosine monoclonal antibody, and alkaline phosphatase-protein G were from Upstate Biotechnology (Lake Placid, NY); [32P]ATP was from New England Nuclear (Boston, MA); Immobilon-P was from Millipore (Bedford, MA); and phosphatidylinositol-specific phospholipase C (PI-PLC; 9.42 U/ml) was from ICN Biochemicals (Irvine, CA).
Analysis of HRP uptake in pancreatic acini. Rat (male Wistar, 75 g) or mouse (C57, adult males) pancreatic acini were prepared by collagenase and mechanical dissociation (10). Endocytosis at the APM was examined using our previously described two-step protocol (10), in which, in step one, acini are incubated for 1 h at 37°C with 0.5 nM CCK-8 in Krebs-Henseleit buffer (KHB) to maximize exocytosis and the insertion of zymogen granule membrane into the APM. Cells are then washed in buffer alone, and then, in step two, endocytosis is assessed by incubation of acini with HRP (5 mg/ml final concentration) in KHB at pH 7.4. For studies examining the effects of various agents on the inhibition of HRP uptake, acini prestimulated with CCK were incubated with the various agents at 4°C for 10 min in KHB-containing HRP followed by incubation for 30 min at 37°C. HRP uptake was measured using a colorimetric assay (3). Statistical analyses were performed using Student's t-test, and results were expressed as means ± SD.
Src kinase assay. Using the protocol supplied with the Src assay kit, 5 µl of the substrate peptide were added to 5 µl of reaction buffer containing (in mM) 100 Tris (pH 7.2), 125 MgCl2, 25 MnCl2, 2 EGTA, 0.25 sodium orthovanadate, and 2 dithiothreitol. Sample (10 µl) was added at 4°C, and the reaction initiated by addition of 10 µl of a solution containing 75 mM MnCl2 and 0.5 mM ATP plus 10 µCi of [32P]ATP (3,000 Ci/mmol). After incubation for various times at 30°C, 20 µl of 40% TCA were added to precipitate the peptide. After incubating at room temperature for 5 min, 25 µl were spotted on 2 × 2 cm P81 filter paper. The filters were washed three times with 0.75% phosphoric acid and once with acetone and then were transferred to scintillation vials. Radioactivity was measured in a Wallack model 1409 liquid scintillation counter.
Western blot analysis. With the use of the same protocol for the analysis of HRP uptake, these same samples were subjected to SDS-PAGE using 10% polyacrylamide gels. This was followed by transfer to Immobilon-P membranes. The blot was then incubated for 2 h at room temperature with a 1:2,000 dilution of a mouse monoclonal phosphotyrosine antibody. The blot was then washed and incubated for 1 h in protein G conjugated to alkaline phosphatase. Bound antibody was detected by reacting with alkaline phosphatase. Phosphorylated polypeptides on Western blots were scanned with a Hewlett-Packard 4C scanner, and the intensity of the bands was quantitated using NIH Image software.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Determination of the signal transduction process in
apical endocytosis. To determine the signal
transduction process by which endocytosis is regulated, pancreatic
acini prestimulated with CCK were incubated with 20 µM cytochalasin D
(microfilament inhibitor), 100 µM lanthanum (calcium uptake
inhibitor), 100 nM staurosporine (protein kinase C inhibitor), 10 µM
forskolin (protein kinase A activator), 50 µM genistein (tyrosine
kinase inhibitor), or 200 µM orthovanadate (tryosine phosphatase
inhibitor). As shown in Fig.
1, there was ~90%
inhibition of HRP uptake by cytochalasin D, confirming previous studies
showing that actin filaments are required for endocytosis at the apical
surface of polarized epithelial cells (13). Lanthanum was without
significant effect as well as staurosporine, consistent with our prior
data on calcium signaling, demonstrating that HRP uptake could not be
activated with a phorbol ester or diacylglycerol (12). Similarly,
activation of protein kinase A with forskolin had no significant effect
compared with the control condition. In contrast, genistein inhibited
HRP uptake by ~50%. If HRP uptake is regulated by tyrosine kinases,
inhibition of tyrosine phosphatase would be expected to enhance HRP
uptake. The 62% increase in HRP uptake observed with orthovanadate
compared with the control condition is consistent with
tyrosine-dependent regulation of HRP uptake.
|
In Fig. 2, the effect of different
concentrations of tyrphostin A1, an inactive analog of the tyrphostins,
is shown. No significant inhibition of HRP uptake was observed. In
contrast, the active analog tyrphostin B42 resulted in ~70%
inhibition of HRP uptake at concentrations of 50-100 µM. This is
similar to the degree of inhibition observed by these agents as well as
genistein in the tyrosine-dependent epidermal growth
factor regulation of rabbit intestinal smooth muscle (14).
These results demonstrate that HRP uptake is regulated through tyrosine
kinase activation.
|
Src kinase activity correlates with activation of HRP
uptake. To determine whether the tyrosine-dependent
regulation of HRP uptake may be mediated through Src kinases, acini
prestimulated with CCK were incubated for various times at 37°C in
KHB buffered at either pH 6.0 or 7.4. Src kinase activity was measured
by incubating cell lysates with an Src-specific peptide substrate in
vitro and measuring labeled phosphate incorporation in the presence of
[-32P]ATP.
As shown in Fig. 3, Src kinase activity in
acini incubated in KHB at pH 7.4 peaked at 15 min and then decreased
over the subsequent 30 min examined. Src kinase activity did not
increase in acini incubated in KHB at pH 6.0. No significant activity
was detected using an Src nonspecific peptide substrate or a
calcium-dependent phosphoserine-specific substrate under these
conditions.
|
The tyrosine-dependent phosphorylation of an 85-kDa
polypeptide correlates with HRP uptake. To examine the
substrates phosphorylated by tyrosine-dependent kinases, acini
incubated under the same conditions used to examine HRP uptake were
subjected to Western blot analysis using a monoclonal antibody that
binds to tyrosine phosphoproteins. We have previously shown that HRP
uptake is abolished when acini are incubated in KHB buffered at pH 6.0, whereas maximal stimulation occurs at pH 7.4 (10). This block in
endocytosis of HRP at pH 6.0 could be completely overcome by addition
of PI-PLC. The effect of PI-PLC on restoring HRP uptake at pH 6.0 was
due to the cleavage of GPI-anchored proteins, specifically GP2 (12). To
determine whether phosphorylation of specific proteins correlates with
HRP uptake, acini preincubated for 1 h with CCK were subsequently incubated in KHB buffered at pH 6.0 or 7.4 for 30 min at 37°C. Western blot analysis of these fractions is shown in Fig.
4A. With
the use of rat pancreatic acini, the phosphorylation of an 85-kDa
protein was specifically increased when cells were incubated at pH 7.4 compared with 6.0. The addition of PI-PLC in KHB buffered at pH 6.0 also increased the phosphorylation of this protein. No other
phosphorylated substrates reproducibly showed this correlation with
endocytosis.
|
A similar phosphoprotein should be demonstrable in other species, given the universal nature of apical endocytosis. Therefore, mouse pancreatic acini were examined using the same protocol utilized for rat. As shown in Fig. 4B, an 80- to 85-kDa phosphoprotein also demonstrated an increase in phosphorylation as the pH of the medium was increased from 6.0 to 7.4. Similarly, PI-PLC increased the phosphorylation of this protein in cells incubated in KHB buffered at pH 6.0. This phosphoprotein localizes to a 100,000-g particulate fraction (Fig. 4C). This indicates that, in two species, phosphorylation of a membrane-associated 85-kDa polypeptide shows a tight correlation with endocytosis as assessed by HRP uptake.
To examine the relevance to endocytosis, the effects of genistein and
orthovanadate on the tyrosine-dependent phosphorylation of the 85-kDa
protein was examined. As shown in Fig. 5,
50 µM genistein inhibited phosphorylation of this substrate in intact acini by 56% compared with the control condition. In contrast, addition of orthovanadate resulted in a 76% increase in the
phosphorylation of the 85-kDa protein over control values. This is
similar to the results observed for genistein and orthovanadate on HRP
uptake (Fig. 1).
|
The phosphorylation of the 85-kDa protein in rat pancreatic acini as a
function of pH of the medium is shown quantitatively in Fig.
6. After incubation in KHB buffered at pH
7.4, phosphorylation of the 85-kDa protein was maximal at 15 min.
Dephosphorylation was observed over the subsequent 30 min to levels
approaching that observed for the pH 6.0 condition. The increase in
phosphorylation of the 85-kDa protein at pH 7.4 compared with pH 6.0 is
similar to the results observed for Src kinase activity (Fig. 3). Note that some increase in the phosphorylation of this substrate occurs in
acini incubated in KHB buffered at pH 6.0 over the initial 20 min of
incubation. Assuming there are multiple sites of phosphorylation as is
typical for many phosphorylated proteins, the explanation for this
phenomenon may be as follows. Phosphorylation at specific sites on the
85-kDa protein under conditions of incubation in KHB at pH 6.0 may be
unrelated to endocytosis, or, alternatively, phosphorylation at certain
sites on this molecule may inhibit endocytosis in a manner similar to
activation or inhibition of Src kinase, depending on which sites are
phosphorylated.
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The results of this study demonstrate that endocytosis at the APM of the pancreatic acinar cell is regulated through activation of tyrosine kinase(s). The tyrosine-dependent phosphorylation of principally one substrate correlated with the activation of HRP uptake. This protein, with an apparent relative molecular mass of 85 kDa, was present in both mouse and rat pancreas. Although this protein was localized to the particulate fraction, its identity remains unknown.
Does phosphorylation of the 85-kDa protein mediate apical endocytosis and is this process regulated through Src kinase activation? Our data are correlative, but the fact that genistein inhibited and orthovanadate stimulated phosphorylation of the 85-kDa protein to an extent similar to that observed for HRP uptake suggests a relationship between these two processes. Activation of Src kinase activity was maximal within 15 min, correlating with peak phosphorylation of the 85-kDa protein. Src kinase activity returned to pH 6.0 levels within 30 min of incubation. Coincident with this, the 85-kDa protein dephosphorylated, although with slower kinetics that are perhaps related to the regulation of phosphatase activity.
Other signaling pathways were not identified. We have previously shown that modulation of calcium-dependent pathways by measurement of fura 2 or the addition of phorbol esters or diacylglycerol had no effect on the activation of HRP uptake in this system (12). Similarly, increasing intracellular cAMP had no effect on HRP uptake. The inhibition of HRP uptake by cytochalasin D is consistent with the finding in other polarized epithelial cell systems that actin filaments are critical in the endocytic process (7, 13, 20, 23). Of note, the inhibition of HRP uptake by cytochalasin D in our study was much greater than that observed for genistein. This is most likely explained by the fact that genistein only partially inhibits tyrosine kinase activity at this dose (14), although we cannot exclude that HRP uptake may occur through other constitutive pathways.
Because the cleavage of GP2 from the APM appears to be the signal that activates HRP uptake in these cells (12), activation of tyrosine kinases may be the transduction pathway through which GP2 mediates this endocytic process. Tyrosine kinase activity has been shown to colocalize with immunoprecipitates of GPI-anchored proteins. In human T lymphocytes, CD59, CD55, and CD48 coimmunoprecipitate with p56lck (25). However, the mechanism by which a GPI-anchored protein, which is confined to the exoplasmic leaflet of the lipid bilayer, can lead to activation of Src kinases on the endoleaflet remains unknown. A likely explanation is that a transmembrane protein also exists, which transduces the signal across the lipid bilayer. Solomon et al. (24) have found that heterotrimeric GTP binding proteins colocalize to these complexes. These authors postulate that clustering of the GPI-anchored proteins on the ectoleaflet of the membrane leads to aggregation of heterotrimeric GTP binding proteins with the tyrosine kinases p59fyn and p56lck on the endoleaflet of the membrane, resulting ultimately in cell proliferation and cytokine production.
Shenoy-Scaria et al. (21) have examined the mechanism by which cross-linking the GPI-linked protein decay-accelerating factor (DAF) leads to lymphocyte proliferation. EL-34 murine thymoma cells were transfected with the cDNA encoding either DAF or a transmembrane form of DAF (DAF-TM). Only the GPI-linked form led to cellular proliferation, activation of tyrosine kinases (p59fyn and p56lck), and phosphorylation of substrates, including an 85-kDa protein. In one study, palmitoylation of an amino-terminal cysteine motif of p59fyn and p56lck was required for colocalization with immunoprecipitates of DAF (22), although another study found palmitoylation was not required (18). However, this cysteine residue at position 3 appears to be critical in forming a complex with GPI-linked proteins. This is absent in p60src, which does not coimmunoprecipitate with DAF.
GPI-anchored proteins are generally clustered within specific lipid domains enriched in glycosphingolipids on the cell surface (4, 5). The cholesterol within this lipid domain is essential for the clustering of GPI-anchored proteins (19) and may contain caveolin (1). Because of the glycosphingolipid composition, these complexes can be isolated as Triton X-100-insoluble complexes from cell lysates. Colocalization of the nonreceptor tyrosine kinase p62yes to these detergent-insoluble membrane complexes has been demonstrated in MDCK cells (2).
Taken together, GPI-anchored proteins appear to exist in a complex with nonreceptor tyrosine kinases in lymphocytes as well as polarized epithelial cells such as MDCK cells. These lipid microdomains on the cell surface, by virtue of other components such as heterotrimeric GTP binding proteins or caveolin, would contain the machinery necessary for endocytosis. In addition, GPI-anchored complexes are associated with patches of polymerized actin that are critical in the early events in endocytosis (7). The current findings that 1) apical endocytosis in the pancreatic acinar cell is regulated by tyrosine kinase activation and 2) our previous studies showing that GPI anchor cleavage is the signal that activates this endocytic process suggest that GP2 may be complexed with tyrosine kinases such as Src. The cleavage of GP2 from the membrane would appear to be the signal that activates membrane internalization through tyrosine kinase signaling, a process that subsequently is dependent on actin filaments.
![]() |
ACKNOWLEDGEMENTS |
---|
This study was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grant R01 DK-52765 (to S. D. Freedman).
![]() |
FOOTNOTES |
---|
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Address for reprint requests: S. D. Freedman, Beth Israel Deaconess Medical Center, Dana 501, 330 Brookline Ave., Boston, MA 02215.
Received 19 March 1998; accepted in final form 13 October 1998.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Anderson, R. G. W.
Caveolae: where incoming and outgoing messengers meet.
Proc. Natl. Acad. Sci. USA
90:
10909-10913,
1993[Abstract].
2.
Arreaza, G.,
K. A. Melkonian,
M. LaFevre-Bernt,
and
D. A. Brown.
Triton X-100-resistant membrane complexes from cultured kidney epithelial cells contain the src family protein tyrosine kinase p62yes.
J. Biol. Chem.
269:
19123-19127,
1994
3.
Bradbury, N. A.,
T. Jilling,
K. L. Kirk,
and
R. J. Bridges.
Regulated endocytosis in a chloride secretory epithelial cell line.
Am. J. Physiol.
262 (Cell Physiol. 31):
C752-C759,
1992
4.
Brown, D. A.,
B. Crise,
and
J. K. Rose.
Mechanism of membrane anchoring affects polarized expression of two proteins in MDCK cells.
Science
245:
1499-1501,
1989[Medline].
5.
Brown, D. A.,
and
J. K. Rose.
Sorting of GPI-anchored proteins to glycolipid enriched membrane subdomains during transport to apical cell surface.
Cell
67:
523-544,
1992.
6.
De Camilli, P.,
D. Peluchett,
and
J. Meldolesi.
Dynamic changes of the luminal plasmalemma in stimulated parotid acinar cells. A freeze fracture study.
J. Cell Biol.
70:
59-74,
1976[Abstract].
7.
Deckert, M.,
M. Ticchioni,
and
A. Bernard.
Endocytosis of GPI-anchored protein in human lymphocytes: role of glycolipid-based domains, actin cytoskeleton, and protein kinases.
J. Cell Biol.
133:
791-799,
1996[Abstract].
8.
Eker, P.,
K. H. Holm,
B. van Deurs,
and
K. Sandvig.
Selective regulation of apical endocytosis in polarized Madin-Darby canine kidney cells by mastoparan and cAMP.
J. Biol. Chem.
269:
18607-18615,
1994
9.
Fallon, R. J.,
M. Danaher,
R. L. Saylors,
and
A. Saxena.
Defective asialoglycoprotein receptor endocytosis mediated by tyrosine kinase inhibitors.
J. Biol. Chem.
269:
11011-11017,
1994
10.
Freedman, S. D.,
H. F. Kern,
and
G. A. Scheele.
Acinar lumen pH regulates endocytosis, but not exocytosis, at the apical plasma membrane of pancreatic acinar cells.
Eur. J. Cell Biol.
75:
153-162,
1998[Medline].
11.
Freedman, S. D.,
H. F. Kern,
and
G. A. Scheele.
Apical membrane trafficking during regulated pancreatic exocrine secretionrole of alkaline pH in the acinar lumen and enzymatic cleavage of GP2, a GPI-linked protein.
Eur. J. Cell Biol.
65:
354-365,
1994[Medline].
12.
Freedman, S. D.,
H. F. Kern,
and
G. A. Scheele.
Cleavage of GPI-anchored proteins from the plasma membrane activates apical endocytosis in pancreatic acinar cells.
Eur. J. Cell Biol.
75:
163-173,
1998[Medline].
13.
Gottlieb, T. A.,
I. E. Ivanov,
M. Adesnik,
and
D. D. Sabatini.
Actin microfilaments play a critical role in endocytosis at the apical but not the basolateral surface of polarized epithelial cells.
J. Cell Biol.
120:
695-710,
1993[Abstract].
14.
Hatakeyama, N.,
D. Mukhopadhyay,
R. K. Goyal,
and
H. I. Akbarali.
Tyrosine kinase-dependent modulation of calcium entry in rabbit colonic muscularis mucosae.
Am. J. Physiol.
270 (Cell Physiol. 39):
C1780-C1789,
1996
15.
Holen, I.,
P. E. Stromhaug,
P. B. Gordon,
M. Fengsrud,
T. O. Berg,
and
P. O. Seglen.
Inhibition of autophagy and multiple steps in asialoglycoprotein endocytosis by inhibitors of tyrosine protein kinases (tyrophostins).
J. Biol. Chem.
270:
12823-12831,
1995
16.
Holm, P. K.,
P. Eker,
K. Sandvig,
and
B. van Deurs.
Phorbol myristate acetate selectively stimulates apical endocytosis via protein kinase C in polarized MDCK cells.
Exp. Cell Res.
217:
157-168,
1995[Medline].
17.
Li, S.,
R. Seitz,
and
M. P. Lisanti.
Phosphorylation of caveolin by src tyrosine kinases.
J. Biol. Chem.
271:
3863-3868,
1996
18.
Rodgers, W.,
B. Crise,
and
J. K. Rose.
Signals determining protein tyrosine kinase and glycosyl phosphatidylinositol anchored protein targeting to a glycolipid-enriched membrane fraction.
Mol. Cell. Biol.
14:
5384-5391,
1994[Abstract].
19.
Rothberg, K. G.,
Y.-S. Ying,
B. A. Kamen,
and
R. G. W. Anderson.
Cholesterol controls the clustering of the glycophospholipid-anchored membrane receptor for 5-methyltetrahydrofolate.
J. Cell Biol.
111:
2931-2938,
1990[Abstract].
20.
Schelling, J. R.,
A. S. Hanson,
R. Marzed,
and
S. L. Linas.
Cytoskeleton-dependent endocytosis is required for apical type 1 angiotensin II receptor-mediated phospholipase C activation in cultured rat proximal tubule cells.
J. Clin. Invest.
90:
2472-2480,
1992[Medline].
21.
Shenoy-Scaria, A. M.,
J. Kwong,
T. Fujita,
M. W. Olszowy,
A. S. Shaw,
and
D. M. Lublin.
Signal transduction through decay-accelerating factor.
J. Immunol.
149:
3535-3541,
1992
22.
Shenoy-Scaria, A. M.,
L. K. Gauen,
J. Kwong,
A. S. Shaw,
and
D. M. Lublin.
Palmitylation of an amino-terminal cysteine motif of protein tyrosine kinases p56lck and p59fyn mediates interaction with glycosyl-phosphatidylinositol-anchored proteins.
Mol. Cell. Biol.
13:
6385-6392,
1993[Abstract].
23.
Shurety, W.,
N. A. Bright,
and
J. P. Luzio.
The effects of cytochalasin D and phorbol myristate acetate on the apical endocytosis of ricin in polarised Caco-2 cells.
J. Cell Sci.
109:
2927-2935,
1996
24.
Solomon, K. R.,
C. E. Rudd,
and
R. W. Finberg.
The association between glycosylphosphatidylinositol anchored proteins and heterotrimeric G protein subunits in lymphocytes.
Proc. Natl. Acad. Sci. USA
93:
6053-6058,
1996
25.
Stefanova, I.,
V. Horejsi,
I. J. Ansotegui,
W. Knapp,
and
H. Stockinger.
GPI-anchored cell-surface molecules complexed to tyrosine kinases.
Science
254:
1016-1019,
1991[Medline].