Recently, p24A and p23 (also termed Tmp21), two members of the p24 protein family, have been proposed to function as integral receptors for the COPI-vesicle coat. This study describes the intracellular localization and trafficking of p24A in comparison to p23. For immunolocalization of p24A and p23, strong reduction and denaturation conditions were necessary to allow antibody interaction. Both p24A and p23 cycle continuously between intermediate compartment (IC) elements and the cis-Golgi network. In vivo trafficking of p24A and p23 tagged to green fluorescent protein (GFP) revealed that both proteins travel by large (up to 1 micrometer in length) microtubule-dependent pre-Golgi carriers with a maximum speed of up to 1.6 micrometer s-1 from the IC to the Golgi cisternae. Aluminum fluoride, a general activator of heterotrimeric G-proteins, blocked peripheral pre-Golgi movements of GFP-p24A/p23 and inhibited fluorescence recovery after photobleaching in the perinuclear Golgi area. p24A and p23 are predominantly colocalized. Overexpression of GFP-p24A, to an extent which did not destroy the Golgi complex, induced delocalization of part of the proteins into ER elements. This study therefore gives new insights into the localization and trafficking behavior of the two COPI-binding proteins p24A and p23.

Anderson
K. L.
,
McNiven
M. A.
(
1995
).
Vesicle dynamics during regulated secretion in a novel pancreatic acinar cell in vitro model.
Eur. J. Cell Biol
66
,
25
38
Aridor
M.
,
Bannykh
S. I.
,
Rowe
T.
,
Balch
W. E.
(
1995
).
Sequential coupling between COPII and COPI vesicle coats in endoplasmic reticulum to Golgi transport.
J. Cell Biol
131
,
875
893
Balch
W. E.
,
McCaffery
J. M.
,
Plutner
H.
,
Farquhar
M. G.
(
1994
).
Vesicular stomatitis virus glycoprotein is sorted and concentrated during export from the endoplasmic reticulum.
Cell
76
,
841
852
Barlowe
C.
,
Orci
L.
,
Yeung
T.
,
Hosobuchi
M.
,
Hamamoto
S.
,
Salama
N.
,
Rexach
M. F.
,
Ravazzola
M.
,
Amherdt
M.
,
Schekman
R.
(
1994
).
COPII: a membrane coat formed by sec proteins that drive vesicle budding from the endoplasmic reticulum.
Cell
77
,
895
907
Bednarek
S. Y.
,
Ravazzola
M.
,
Hosobuchi
M.
,
Amherdt
M.
,
Perrelet
A.
,
Schekman
R.
,
Orci
L.
(
1995
).
COPI-and COPII-coated vesicles bud directly from the endoplasmic reticulum in yeast.
Cell
83
,
1183
1196
Belden
W. J.
,
Barlowe
C.
(
1996
).
Erv25p, a component of COPII-coated vesicles, forms a complex with Emp24p that is required for efficient endoplasmic reticulum to Golgi transport.
J. Biol. Chem
271
,
26939
26946
Bloom
G. S.
,
Goldstein
L. S. B.
(
1998
).
Cruising along microtubule highways: How membranes move through the secretory pathway (Commentary).
J. Cell Biol
140
,
1277
1280
Blum
R.
,
Feick
P.
,
Puype
M.
,
Vandekerckhove
J.
,
Klengel
R.
,
Nastainczyk
W.
,
Schulz
I.
(
1996
).
Tmp21 and p24A, two type I proteins enriched in pancreatic microsomal membranes, are members of a protein family involved in vesicular trafficking.
J. Biol. Chem
271
,
17183
17189
Cole
N. B.
,
Smith
C. L.
,
Sciaky
N.
,
Terasaki
M.
,
Edidin
M.
,
Lippincott-Schwartz
J.
(
1996
).
Diffusional mobility of Golgi proteins in membranes of living cells.
Science
273
,
797
801
Cosson
P.
,
Letourneur
F.
(
1994
).
Coatomer interaction with di-lysine endoplasmic reticulum retention motifs.
Science
263
,
1629
1631
Donaldson
J. G.
,
Lippincott-Schwartz
J.
,
Bloom
G. S.
,
Kreis
T. E.
,
Klausner
R. D.
(
1990
).
Dissociation of a 110-kDa peripheral membrane protein from the Golgi apparatus is an early event in brefeldin A action.
J. Cell Biol
111
,
2295
306
Donaldson
J. G.
,
Lippincott-Schwartz
J.
,
Klausner
R. D.
(
1991
).
Guanine nucleotides modulate the effects of brefeldin A in semipermeable cells: regulation of the association of a 110-kD peripheral membrane protein with the Golgi apparatus.
J. Cell. Biol
112
,
579
588
Donaldson
J. G.
,
Finazzi
D.
,
Klausner
R. D.
(
1992
).
Brefeldin A inhibits Golgi membrane-catalysed exchange of guanine nucleotide onto ARF protein.
Nature
360
,
350
352
Fiedler
K.
,
Veit
M.
,
Stamnes
M. A.
,
Rothman
J. E.
(
1996
).
Bimodal binding of coatomer with the p24 family of putative cargo receptors.
Science
273
,
1396
1399
Helms
J. B.
(
1995
).
Role of heterotrimeric GTP binding proteins in vesicular protein transport: indications for both classical and alternative G protein cycles.
FEBS Lett
369
,
84
88
Helms
J. B.
,
Helms-Brons
D.
,
Brugger
B.
,
Gkantiragas
I.
,
Eberle
H.
,
Nickel
W.
,
Nurnberg
B.
,
Gerdes
H. H.
,
Wieland
F. T.
(
1998
).
A putative heterotrimeric G-protein inhibits the fusion of COPI-coated vesicles.
J. Biol. Chem
273
,
15203
15208
Ktistakis
N. T.
,
Roth
M. G.
,
Bloom
G. S.
(
1991
).
PtK1 cells contain a nondiffusable, dominant factor that makes the Golgi apparatus resistant to Brefeldin A.
J. Cell. Biol
113
,
1009
1023
Kuismanen
E.
,
Saraste
J.
(
1989
).
Low temperature-induced transport blocks as tools to manipulate membrane traffic.
Methods Cell Biol
32
,
257
274
Letourneur
F.
,
Gaynor
E. C.
,
Hennecke
S.
,
Demolliere
C.
,
Duden
R.
,
Emr
S. D.
,
Riezman
H.
,
Cosson
P.
(
1994
).
Coatomer is essential for retrieval of dilysine-tagged proteins to the endoplasmic reticulum.
Cell
79
,
1199
1207
Lewis
M. J.
,
Pelham
H. R.
(
1992
).
Ligand-induced redistribution of a human KDEL-receptor from the Golgi complex to the endoplasmic reticulum.
Cell
68
,
353
364
Lippincott-Schwartz
J.
,
Yuan
L. C.
,
Bonifacino
J. S.
,
Klausner
R. D.
(
1989
).
Rapid redistribution of Golgi proteins into the ER in cells treated with brefeldin A: evidence for membrane cycling from Golgi to ER.
Cell
56
,
801
813
Lippincott-Schwartz
J.
,
Donaldson
J. G.
,
Schweizer
A.
,
Berger
E. G.
,
Hauri
H. P.
,
Yuan
L. C.
,
Klausner
R. D.
(
1990
).
Microtubule-dependent retrograde transport of proteins into the ER in the presence of brefeldin A suggests an ER recycling pathway.
Cell
60
,
821
836
Mittal
R.
,
Ahmadian
M. R.
,
Goody
R. S.
,
Wittinghofer
A.
(
1996
).
Formation of a transition-state analog of the Ras GTPase reaction by Ras-GDP, tetrafluoroaluminate and GTPase-activating proteins.
Science
273
,
115
117
Munro
S.
,
Pelham
H. R.
(
1987
).
A C-terminal signal prevents secretion of luminal ER proteins.
Cell
48
,
899
907
Nickel
W.
,
Sohn
K.
,
Bunning
C.
,
Wieland
F. T.
(
1997
).
p23, a major COPI-vesicle membrane protein, constitutively cycles through the early secretory pathway.
Proc. Natl. Acad. Sci. USA
94
,
11393
11398
Orci
L.
,
Glick
B. S.
,
Rothman
J. E.
(
1986
).
A new type of coated vesicular carrier that appears not to contain clathrin: its possible role in protein transport within the Golgi stack.
Cell
46
,
171
184
Orci
L.
,
Tagaya
M.
,
Amherdt
M.
,
Perrelet
A.
,
Donaldson
J. G.
,
Lippincott-Schwartz
J.
,
Klausner
R. D.
,
Rothman
J. E.
(
1991
).
Brefeldin A, a drug that blocks secretion, prevents the assembly of non-clathrin-coated buds on Golgi cisternae.
Cell
64
,
1183
1195
Orci
L.
,
Perrelet
A.
,
Rothman
J. E.
(
1998
).
Vesicles on strings: Morphological evidence for processive transport within the Golgi stack.
Proc. Natl. Acad. Sci. USA
95
,
2279
2283
Palade
G. E.
(
1975
).
Intracellular aspects of the process of protein synthesis.
Science
189
,
347
358
Pepperkok
R.
,
Scheel
J.
,
Horstmann
H.
,
Hauri
H. P.
,
Griffiths
G.
,
Kreis
T. E.
(
1993
).
-COP is essential for biosynthetic membrane transport from the endoplasmic reticulum to the Golgi complex in vivo.
Cell
74
,
71
82
Peter
F.
,
Plutner
H.
,
Zhu
H.
,
Kreis
T. E.
,
Balch
W.
(
1993
).
Beta-COP is essential for transport of protein from the endoplasmic reticulum to the Golgi in vitro.
J. Cell Biol
122
,
1155
1167
Prasher
D. C.
,
Eckenrode
V. K.
,
Ward
W. W.
,
Prendergasr
F. G.
,
Cormier
M. J.
(
1992
).
Primary structure of the Aequorea victoria green fluorescent protein.
Gene
111
,
320
323
Presley
J. F.
,
Cole
N. B.
,
Schroer
T. A.
,
Hirschberg
K.
,
Zaal
J. M.
,
Lippincott-Schwartz
J.
(
1997
).
ER-to-Golgi transport visualized in living cells.
Nature
389
,
81
85
Rojo
M.
,
Pepperkok
R.
,
Emery
G.
,
Kellner
R.
,
Stang
E.
,
Parton
R. G.
,
Gruenberg
J.
(
1997
).
Involvement of the transmembrane protein p23 in biosynthetic protein transport.
J. Cell Biol
139
,
1119
1135
Rothman
J. E.
,
Wieland
F. T.
(
1996
).
Protein sorting by transport vesicles.
Science
272
,
227
234
Saraste
J.
,
Kuismanen
E.
(
1984
).
Pre-and post-Golgi vacuoles operatein the transport of Semliki Forest virus membrane glycoproteins to the cell surface.
Cell
38
,
535
549
Scales
S. J.
,
Pepperkok
R.
,
Kreis
T. E.
(
1997
).
Visualization of ER-to-Golgi transport in living cells reveals a sequential mode of action for COPII and COPI.
Cell
90
,
1137
1148
Schekman
R.
,
Mellman
I.
(
1997
).
Does COPI go both ways?.
Cell
90
,
197
200
Schimmöller
F.
,
Singer-Kruger
B.
,
Schröder
S.
,
Kruger
U.
,
Barlowe
C.
,
Riezman
H.
(
1995
).
The absence of Emp24p, a component of ER-derived COPII-coated vesicles, causes a defect in transport of selected proteins to the Golgi.
EMBO J
14
,
1329
1339
Schweizer
A.
,
Fransen
J. A.
,
Matter
K.
,
Kreis
T. E.
,
Ginsel
L.
,
Hauri
H. P.
(
1990
).
Identification of an intermediate compartment involved in protein transport from the endoplasmic reticulum to Golgi apparatus.
Eur. J. Cell Biol
53
,
185
196
Sohn
K.
,
Orci
L.
,
Ravazzola
M.
,
Amherdt
M.
,
Bremser
M.
,
Lottspeich
F.
,
Fiedler
K.
,
Helms
J. B.
,
Wieland
F. T.
(
1996
).
A major transmembrane protein of Golgi-derived COPI-coated vesicles involved in coatomer binding.
J. Cell. Biol
135
,
1239
1248
Stamnes
M. A.
,
Craighead
M. W.
,
Hoe
M. H.
,
Lampen
N.
,
Geromanos
S.
,
Tempst
P.
,
Rothman
J. E.
(
1995
).
An integral membrane component of coatomer-coated transport vesicles defines a family of proteins involved in budding.
Proc. Natl. Acad. Sci. USA
92
,
8011
8015
This content is only available via PDF.