Institute of Molecular Medicine and Genetics, Medical College of Georgia, Augusta, Georgia 30912
ACTIN DYNAMICS ARE CENTRAL to a number
of basic physiologies in polarized cells, including polarized cell
domains, cell shape, and intracellular trafficking and signaling. The
study by Ammar and colleagues, the current article in focus (Ref.
1, see p. C407 in this issue), provides evidence in
gastric parietal cells for the regulation of actin pools of varying
stability and turnover. The investigators have utilized latrunculins
that sequester G-actin monomers, thereby reducing the pool of
polymerizable actin. This mechanism is substantially different from
cytochalasin D, which leads to fragmentation of F-actin filaments. Thus
latrunculin is not expected to alter stable F-actin structures.
However, in situations where there is turnover of F-actin, latrunculin
treatment can deplete cellular F-actin. In support of this concept,
these investigators have observed little effect of latrunculin B
treatment on resting parietal cells. In contrast, significant effects
were observed in cells stimulated with increases in cAMP. In cultured parietal cells, the investigators were able to follow two separate cAMP-stimulated actin-based functions that were associated with actin
turnover: lamellapodial spreading along the Matrigel substratum and
vesicle fusion into canalicular vacuoles that are lined by F-actin.
Although both functions could be altered by treatment with latrunculin
B, alteration of canalicular F-actin and inhibition of aminopyrine
accumulation (an indirect assay of tubulovesicle fusion) required at
least 10-fold higher concentrations of latrunculin than that which is
needed for inhibition of lamellapodial spreading. These results suggest
the presence of distinct pools of basolateral and apical F-actin
filaments in parietal cells.
The presence of separable pools of actin in polarized epithelial cells
underscores the subcellular specializations required for a
physiological response to global signals such as increases in
intracellular cAMP. Yao and colleagues (11) have
previously demonstrated that the apical canalicular membrane in
parietal cells is underlined by F-actin filaments that contain
The parietal cell recycling of the H+-K+-ATPase
remains the most dramatic example of apical membrane recycling
(7). Although this process is massively amplified in
parietal cells, it is a fundamental component of polarized cell
function in all epithelial cell systems. Recent studies have
demonstrated that many of the candidate regulators found on parietal
cell tubulovesicles, including Rab11 small GTPases,
soluble N-ethylmaleimide-sensitive factor attachment
protein target receptor proteins (SNAREs), and secretory carrier
membrane proteins (SCAMPs), are also present on vesicles associated
with general apical recycling systems in cultured cell systems such as
Madin-Darby canine kidney (MDCK) cells (2-4, 6, 10).
However, in contrast with parietal cells, investigations of general
apical recycling systems in cultured cell line systems have emphasized
the important role of microtubules in the coordinated processes of
apical recycling and basolateral-to-apical transcytosis (2). Disruption of microtubules in these systems markedly
inhibits both transcytosis and apical recycling. In contrast, the
present investigations, as well as previous studies in parietal cells (11), all emphasize a prominent role of the actin-based
cytoskeleton in the apical recycling of the
H+-K+-ATPase. Microtubule disruption appears to
a have a relatively minor effect on parietal cell function.
How can one rationalize these disparate results in different cell
systems? First, recent investigations have demonstrated that an
actin-based motor, myosin Vb, associates with Rab11a and is required
for cargo exit from the apical recycling system in MDCK cells
(9). Thus the actin-based cytoskeleton may be a general
regulator of movement of recycled cargoes out from apical recycling
systems toward eventual fusion with apical membrane targets. Second, it
is possible that the parietal cell, in addition to amplifying the
apical recycling system, has also specialized specifically in this
process to the exclusion of other vesicle trafficking functions
possessed by most polarized epithelial cells. Many model systems, such
as MDCK cells, utilize the apical recycling system for processing of
both apically recycling and transcytosing cargoes, with the
transcytotic pathway accounting for the majority of the membrane
trafficked (2). In contrast, the parietal cell likely
processes little if any transcytotic traffic. Thus the microtubule-based dependence of the apical recycling system in MDCK
cells may reflect the requirement for microtubules for the entry of the
predominant source of transcytosing membranes into the system. No
studies have examined the effects of latrunculins on apical recycling
or transcytosis in MDCK cells. Still, it is important to note that the
trafficking of another apically recycled protein,
Na+/H+ exchanger 3, is altered by latrunculin
treatment (8). The absence of microtubule dependence in
parietal cell recycling may reflect its purely actin-based recycling
pathways without any need for membrane pools that require microtubules
for entry into the recycling system.
The studies presented in the present report establish the polarization
of separate actin pools with differentiable functions at the
basolateral and apical membranes. Future clarification of the
requirements for actin-based vesicle movements out from apical
recycling systems in cell lines such as MDCK should provide an
increased understanding of the general role of the actin cytoskeleton in regulating membrane recycling. Similarly, further investigations must now address how the elements of the actin cytoskeleton interact with tubulovesicle proteins to mediate and modulate vectorial fusion of
H+-K+-ATPase-containing vesicles with the
parietal cell secretory canaliculus.
ARTICLE
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REFERENCES
-actin, while the basolateral membrane contains
-actin.
Separation of these two pools of F-actin containing different actin
isoforms accounts for the polarized distribution of actin-binding
proteins such as ezrin, which is predominantly associated with the
apical membranes through its binding with
-actin filaments
(11). It is of note that evidence does exist for movement
of proteins between these two pools of F-actin in parietal cells. Chew
and colleagues (5) have noted in parietal cells the
redistribution of Lasp-1, a multidomain linker protein, from
basolateral
-actin filaments to the secretory canaliculus in
response to increases in intracellular cAMP. The effects of
latrunculins observed in the present report could therefore reflect
either differential binding of latrunculins to different
-actin
monomer isoforms of actin or differences in the rates of F-actin turnover.
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FOOTNOTES |
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Address for reprint requests and other correspondence: J. R. Goldenring, Institute of Molecular Medicine and Genetics, CB-2803, Medical College of Georgia, 1120 Fifteenth St., Augusta, GA 30912 (E-mail: jgolden{at}mail.mcg.edu).
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