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Transport of casein submicelles and formation of secretion granules in the golgi apparatus of epithelial cells of the lactating mammary gland of the rat.

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THE ANATOMICAL RECORD 235:363-373 (1993)
Transport of Casein Submicelles and Formation of Secretion
Granules in the Golgi Apparatus of Epithelial Cells of the Lactating
Mammary Gland of the Rat
Departments of Anatomy (Y.C., L.H.) and Animal Sciences (L.X., J.D.T.) McGill
University, Montreal, Quebec, Canada; Ddpartment de biologie cellulaire et moldculaire,
Centre d'itudes nucldaires, France (A.R.)
Lactating mammary glands fixed by perfusion with 5% glutaraldehyde subsequently were postfixed with potassium ferrocyanide reduced osmium or were treated with tannic acid. Stained thin sections were
examined with the electron microscope and stereopairs were prepared.
The distribution of casein submicelles was analyzed in the various components of the Golgi apparatus. The Golgi stacks were composed of five or six
elements, all of which contained casein submicelles 20 nm in diameter. The
cis-tubular network or cis-element, as well as the underlying three or four
midsaccules, showed these casein submicelles either attached to their membrane or free in the lumen. The trans-most element of the stacks formed
distended prosecretory granules in which both isolated or clustered casein
submicelles were suspended in an electron-lucentfluid. These micellar aggregates increased in size and became progressively more compact to form
spherical dense bodies or casein micelles, in which the individual 20 nm
particles could easily be resolved. Casein micelles were seen in secretory
granules in addition to a wispy material of low density. The numerous small
spherical vesicles (80 nm or larger) seen on the cis, lateral, or trans aspects
of the stacks did not appear to contain free casein submicelles. This raises
questions regarding the role of these vesicles in the transport of casein
macromolecules through the Golgi stacks. It was noticeable that in this
Golgi apparatus a trans-Golgi network was limited to a few small residual
tubules free from casein submicelles. It thus appears that the greater part
of the trans-most Golgi element gives rise to the large prosecretory granules. After leaving the Golgi region and prior to exocytosis, the secretory
granules often fuse to form larger granules before exocytosis.
0 1993 Wiley-Liss, Inc.
Key words: Prosecretory granules, Trans-Golgi network, Exocytosis
The structural complexity and variability of the
Golgi apparatus of various types of fully differentiated
cells in vivo has already been highlighted as a result of
the use of stereoelectron microscopy (Rambourg and
Clermont, 1990). During the past decade, biochemical,
cytochemical, and genetic studies have added substantial information on the composition and possible functions of the cis-, mid- and trans-elements of this organelle, as reviewed recently by Mellman and Simons
(1992). These authors indicated, however, th a t many
questions concerning the structural and functional relationships of Golgi components remain to be answered, in particular the exact nature of the mechanisms by which the glycosylated proteins are
transported from the cis to the trans face of the Golgi
stacks (Mellman and Simons, 1992).
Numerous vesicles of various sizes are seen associated with the stacks of Golgi saccules and these have
generally been considered as carriers of proteins not
only from the proximal cisternae of the endoplasmic
reticulum to the cis aspect of the Golgi stacks but also
from one saccule to the next in a cis-trans direction
until the proteins are packaged within vesicles budding from the trans-most Golgi element or trans-Golgi
network (Palade, 1975; Farquhar, 1983, 1985; Farquhar and Palade, 1981; Goldfischer, 1982; Rothman,
1981, 1985; Dunphy and Rothman, 1985; Pfeffer and
Rothman, 1987; Griffith and Simons, 1986; Wattenberg, 1991). According to this vesicular transport
model, the vesicles would be mobile, while the other
elements of the Golgi stacks would be comparatively
stable. This model of protein transport across the Golgi
Received June 16, 1992; accepted July 24, 1992
stacks is still debated, however, and alternate models
are now being considered (Mellman and Simons, 1992).
As indicated by these authors, one piece of information
that would favor the vesicular transport model would
be to demonstrate that the Golgi vesicles seen on the
edges of the saccules do contain the proteins present
within the saccules themselves. In the absence of such
data, the role of these particular vesicles remains uncertain.
It is the purpose of the present study to investigate
the three dimensional structure of the Golgi apparatus
of the secretory cells of mammary glands and, more
specifically, to examine by three-dimensional electron
microscopy the distribution of casein submicelles
within this organelle, with the hope of clarifying the
respective role of saccules and vesicles in their transport from the cis to the trans face of the stacks. Indeed
the formation of casein micelles, which form by aggregation of submicelles, has been well studied via electron microscopy (reviewed in Mather and Keenan,
1983; Hollman, 1974; Saacke and Heald, 1974; Keenan
et al., 1974; Wellings, 1969; Bargmann and Welsch,
1969). However, the exact distribution of casein submicelles within the various elements of the Golgi apparatus, including the associated small vesicles, has
received comparatively little attention. The use of reduced osmium as a postfixation proposed by Karnovsky
(1971) or the treatment with tannic acid to improve the
contrast of some structures as suggested by Simionescu
and Simionescu (1976) on tissues fixed by perfusion
was found to be particularly useful not only to determine the distribution of casein submicelles within the
Golgi apparatus but also better to illustrate the steps
in the formation of the casein micelles that are present
in the secretory granules.
The mammary glands of ten lactating SpragueDawley rats (350-450 g) whose pups were 8 days old at
the time of the experiment were fixed by perfusion
through the abdominal aorta in an anterograde direction with 5% glutaraldehyde buffered in sodium cacodylate (0.1 MI containing 0.05%CaC1, at pH 7.4. Ten
minutes later, the mammary glands were removed, cut
into small 1 mm3 pieces, and placed in the same fixative for 1 h r a t 4°C. The tissue was then washed in
several changes of 0.1 M sodium cacodylate buffer and
left overnight in the same buffer a t 4°C. On the following day, the mammary gland tissue was postfixed in
one of two ways. In the case of six animals, the tissue
was postfixed for 1 h r a t 4°C in reduced osmium (1:l
mixture of 2%aqueous osmium tetroxide and 3% aqueous potassium ferrocyanide; Karnovsky, 1971). To increase the contrast of casein micelles and submicelles
and coated vesicles, tissue from four other lactating
females were postfixed according to the method of Simionescu and Simionescu (1976). This consisted of
washing the tissues several times in 0.1 M sodium cacodylate buffer, followed by immersions in 1%buffered
tannic acid (3 x 20 min each, pH 7.0) at room temperature. The reaction was arrested in 1% buffered
Na,SO, (20 min, pH 7.4) also a t room temperature.
After postfixation, the tissues were dehydrated in
ethanol and propylene oxide and embedded in Epon.
Thick sections (0.5 Fm) were cut and stained with tolu-
idine blue to locate the glandular cells and evaluate the
quality of fixation. Thin sections of selected areas of the
blocks were cut with a diamond knife and stained with
both uranyl acetate (5 min) and lead citrate (2 min). All
thin sections were examined with a Philips 400 or 400T
electron microscope.
For stereoscopy, grids were placed on the goniometric stage of the electron microscope, and stereopairs
were obtained by taking pictures of the same field after
tilting the specimen at -12" and + 12" from the original 0" position. A three-dimensional magnified image
of the structure was obtained by looking at properly
adjusted pairs of such photographs with a stereoscopic
binocular lens.
General Appearance of Glandular Cells
The cuboidal or pyramidal glandular epithelial cells
showed a spherical nucleus surrounded by stacked cisternae of rough endoplasmic reticulum (ER) (Fig. 1).
Secretory granules variable in size, and appearing as
vacuoles containing the electron dense casein micelles,
were seen toward the apex of the cell but their number
varied markedly from cell to cell (Fig. 1). Occasionally,
lipid droplets were also seen in the apical cytoplasm. In
thin sections and at low magnification, the Golgi apparatus appeared as stacks of more or less distended
saccules, widely separated from each other, and located
in the juxta- and supranuclear region of the cell (Fig.
1). In addition to mitochondria distributed throughout
the cytoplasm, a few dense bodies identified as lysosomes were also present (Fig. 1). The apical cell surface
showed numerous long microvilli, while a t the base
numerous folds of the plasma membrane were underlined by a continuous basement membrane (Fig. 1).
The Golgi Apparatus
In thin sections, the stacks of saccules were separated from each other, and intersaccular regions were
not readily evident. However, a three-dimensional
analysis of the Golgi apparatus of these cells has shown
that it is a continuous network formed by a branching
and anastomotic irregular ribbon (Dylewski et al.,
1984) as observed in other glandular cells (Rambourg
and Clermont, 1990).
Each stack of saccules consisted of five or six closely
apposed elements that presented the following structural characteristics from the cis to the trans face of the
pile. In sections perpendicular to the Golgi ribbon, the
first or cis-element appeared as a series of more or less
distended elongated or ovoid membranous profiles connected by bridges (Figs. 2-5). These images corresponded to sections through a network of anastomosed
tubules or a highly porous or fenestrated saccule as
seen in face views of the stack (see Fig. 7). The ciselement characteristically contained spherical particulates, 20 nm in diameter, identified as casein submicelles (Figs. 3, 6). No such particulates were present in
the lumen of the ER cisternae seen at proximity or in
the small vesicles seen between the ER and the ciselement (Figs. 2-5). Underlying the cis-element were
four or five saccules showing the following similar features. In transverse sections, they showed a lumen
which was variable in size and presented few disconti-
Fig. 1. Low-power photograph showing a glandular cell with its
centrally located nucleus (N). The cell apex shows microvilli (M) protruding into the acinar lumen and at the base a n irregular folded
plasma membrane is seen facing the basement membrane (BM). Numerous stacks of cisternae of the rough endoplasmic reticulum (ER)
are seen on the basolateral aspects of the nucleus. Several large vacuolated secretory granules (Sg) containing the dense casein micelles
are seen at the apex of the cell. Some secretory granules are seen
proximal to stacks of Golgi saccules (S). Mitochondria (m) are seen
throughout the cytoplasm. x 11,200.
nuities or pores. The greater part of the saccule had a
lumen that was comparatively narrower than that of
the cis-element. They contained casein submicelles
that were close to the saccular membrane or free in the
lumen (Figs. 2-4). Flattened portions of these saccules
(i.e., 10 nm in width) too narrow to contain casein
submicelles alternated with slightly distended portions, which contained the secretion product (Figs. 3,
6). Other portions of the saccules particularly toward
their edges were often slightly distended (Figs. 5, 6).
The saccules on the trans-side of the stack had a tendency to show swellings containing casein submicelles
suspended in an electron-lucent fluid (Figs. 2, 3). Occasionally, the cis-element and the three or four underlying saccules were interrupted along their length by
large fenestrations in register forming typical panshaped cavities and containing several small (80 nm)
vesicles (Figs. 5, 7). These vesicle-containing cavities
corresponded to the “wells” originally described in the
Golgi stacks of spermatids (Hermo et al., 1980) and
Fig. 2. Golgi region of a glandular cell showing several stacks of
Golgi saccules ( S ) and the various steps in the formation of secretory
granules (Sg). The various components of the stacks contain small
particles, 20 nrn in diameter, corresponding to casein submicelles
(small straight arrows). On the trans aspect of the stacks, distended
prosecretory granules (Pg) contain submicelles forming linear clusters or irregular compact aggregates (large curved arrows). These
prosecretory granules frequently show coated buds at their surface
(small curved arrows). Secretion granules (Sg)show typical spherical
casein micelles (asterisks). Small vesicles (V) are seen in the Golgi
region a few of them being coated (CV). A few short membranous
tubules (TI are also seen on the trans aspect of the Golgi stack. ER,
endoplasrnic reticulum; m, mitochondrion. x 54,000.
Fig. 3. High magnification of a Golgi stack. This material was
treated with tannic acid to stain the casein submicelles. These particulates, 20 nm in diameter (arrows), are seen in all elements of the
stack including the cis-element (CE). The aggregation of casein submicelles (white asterisk) is observed in the distended prosecretory
granule (Pg) on the trans aspect of the stack. On the cis and trans
aspects of the stack, some densifications of the cytoplasmic matrix
(black asterisks) are noticeable. ER, cisternae of endoplasmic reticulum. V, vesicles. x 80,000.
Fig. 4. Stack of Golgi elements showing the cis-element (CE) and the
underlying saccules (S) all containing casein submicelles. A portion of
a saccule is flattened (curved arrow). The preparation was not treated
with tannic acid, but the pale stained casein submicelles are still
visib!e within the distended portions of the saccules and in a prose-
cretory granule (Pg), in which they form a dense aggregate (white
asterisk). The small vesicle (v) seen at the edges of the saccules are
free from casein submicelles. x 60,000.
Fig. 5. Stacks of (S) saccules showing casein submicelles in the lumen of each and in a distended prosecretory granule (Pg) on the trans
aspect. On the cis face, clusters of small vesicles (V) are seen between
the cisternae of endoplasmic reticulum (ER) and the stack of saccules.
Some of these vesicles are seen associated with a densification of the
cytoplasmic matrix (white asterisk). Some small vesicles are also seen
in a “well” (W) or pan-shaped cavity within the stack. The cis-element
(CE)with a portion of it seen in face view (black asterisk) is porous. At
lower right, a compact casein micelle (open arrow) within a secretory
granule (Sg) shows a characteristic honeycombed substructure. N,
nucleus. x 60,000.
since then in the Golgi apparatus of a variety of cell
types (Ichikawa and Ichikawa, 1987; Rambourg and
Clermont, 1990). On the trans aspect of the stacks,
markedly distended vesicular structures, referred t o as
prosecretory granules, contained casein submicelles either attached to the membrane or loosely bound to each
other and suspended in an abundant electron-lucent
fluid (Figs. 2, 4). These large prosecretory granules
varied in shape; they were occasionally hemispherical,
with their flattened part applied to the overlying saccule (Fig. 6) or spheroidal and partly separated from
the stack (Figs. 2-5). In prosecretory granules, the
casein submicelles formed loose linear aggregates,
which became progressively denser and more irregular
in shape (Fig. 1).The large prosecretory granules frequently showed coated buds at their surface (Fig. 2).
Finally, in the Golgi region but separated from the
Golgi stack, spherical secretory granules, contained
spherical dense bodies composed of closely packed
casein submicelles which gave to these bodies a characteristic honeycombed texture (Figs. 5,9b). Such
spherical bodies, or casein micelles, varied in size, the
largest being 200-300 nm in diameter. They were suspended in an abundant electron-lucent fluid, which
also contained a pale wispy precipitate but rare free
casein submicelles (Fig. 9a). All transitions between
the prosecretory and the secretory granules were observed, but such images were rare suggesting that the
transformation of the large prosecretory granules into
secretory granules must be rapid.
Very large secretory granules were also seen outside
the Golgi region and close to the apex of the cell (Figs.
1,9a). Such an increase in volume of some of the secretory granules seemingly resulted from the fusion of
smaller secretory granules with each other before exocytosis. Indeed, images of fusion between granules
were frequently encountered (Fig. 9a).
Other Membrane-Bound Elements Associated With the
Various Golgi Elements
Spherical vesicles were seen on the cis, lateral, and
trans aspects of the Golgi ribbon. On the cis aspect of
the Golgi stack, between the ER cisternae and the ciselement, small vesicles 80 nm in diameter were present
(Figs. 4,5). Similar clusters of small vesicles were also
seen along the edges of the saccules (Fig. 5). The 20 nm
casein submicelles were rarely if ever seen within
these vesicles (Figs. 4, 7, 8). This was particularly evident when such vesicles were examined in stereopairs
(Figs. 7,8).
On the trans aspect of the stack, the vesicles were
variable in size and some presented a distinct coat (Fig.
2). The latter were similar in diameter to coated buds
seen at the surface of the prosecretory granules (Fig. 2).
All these vesicles were free from casein submicelles.
Occasionally, some membranous profiles had the appearance of short tubules and possibly represented residual elements detached from the prosecretory granules (Fig. 2). In the trans region of the Golgi apparatus,
in addition to various prosecretory granules, secretory
granules, and some cisternae of the rough ER, there
were also multivesicular bodies containing flattened
vesicles (Fig. 6).
Steps in the Formation of Casein Micelles
The present study confirmed that the casein micelles
form as a result of aggregation of small particulates or
casein submicelles within prosecretory granules seen
on the trans aspect of the Golgi stacks or in the transGolgi region as reviewed by various authors (Keenan
et al., 1974; Wellings, 1969; Hollman, 1974; Bargmann
and Welsch, 1969). There appears to be three distinct
steps in the formation of casein micelles. First, there is
the formation within the Golgi elements of casein submicelles, a macromolecular aggregate of several different casein molecules [ a , p , K ] previously synthesized in
the ER. Such casein submicelles, which are 20 nm in
diameter in our electron micrographs, were equivalent
in size to the casein submicelles of cow’s milk (Walstra
and Jenness, 1984). They were found within all elements of the Golgi stack, including the cis-element
(Fig. 10). They were rarely if ever seen within the
small vesicles associated with the Golgi stack. Attached or not to the saccular membrane, the submicelles were distributed throughout the saccules, except
in their flattened portions, where the saccular lumen
was too narrow to accommodate them. Second, the initial aggregation of casein submicelles, as linear and
then irregular larger compact aggregates, was observed only in the distended trans-most Golgi element
or prosecretory granules (Fig. 10). Therefore, the binding of casein submicelles to each other via calcium
phosphate bridges (Walstra and Jenness, 1984) does
not appear to take place within the Golgi saccules
themselves but occurs within the distended vacuolated
prosecretory granules, which soon separate from the
trans-aspect of the stack (Fig. 10). Third, the presence
of the typical spherical compact casein micelles, in
which the casein submicelles form a regular pattern
(Bargmann and Welsch, 1989), and in our material
show a typical honeycombed texture, was observed
only in the secretory granules present in the Golgi region but distal from the stack (Fig. 10). Thus the micelle formation occurs subsequent to the post translational events of phosphorylatiodglycosylation and to
the intrasaccular presence of calcium and inorganic
phosphate (Larson, 1985).
Within the prosecretory and secretory granules, the
casein submicelles and micelles were suspended in an
electron-lucent fluid which presumably contains lactose and other milk proteins (a-lactalbumin, p-lactoglobulin), the latter being either a t too low a concentration or poorly preserved during tissue processing. As
indicated by Holt (1983) and Mather and Keenan
(19831, the marked swelling of the trans-most Golgi
elements may be due to the presence of lactose accompanied or followed by an influx of water, a major constituent of milk, to maintain the osmotic equilibrium.
The marked swelling mentioned above appears to
mobilize the greater part of the membrane of the transmost Golgi element, and in these as in other glandular
cells, the trans-Golgi network is considerably reduced
in size or absent (Fig. 10). Indeed the trans-Golgi network, which is extensive in nonglandular cells, is less
developed in exocrine secretory cells (Rambourg and
Clermont, 1990). In some secretory cells, such as in
prolactin cells (Rambourg et al., 1992) or the epithelial
Fig. 6. Portion of a Golgi stack in tannic acid-treated tissue, showing
casein submicelles in the saccules. The saccules are generally distended, at the edges in particular, the parts of the saccules are flattened and too narrow to contain the casein submicelles (arrows). A
multivesicular body (MV) is seen next to a prosecretory granule (Pg).
A secretory granule (Sg) with a casein micelle (open arrow) is also
visible. Dense cytoplasmic matrix (asterisk). x 80,000.
Figs. 7-8. Stereopairs of small portions of the Golgi apparatus. A
single magnified stereoscopic image may be obtained by utilizing a
properly adjusted (at 65 mm) binocular lens.
Fig. 7. Tangential section through the Golgi apparatus with a face
view of some Golgi elements.The cis-element (asterisk) is highly porous. The numerous small vesicles seen around the Golgi saccules are
free of casein submicelles (arrow). W, face view of a “well” containing
small vesicles. N, nucleus. x 58,500.
Fig. 8. Edges of a Golgi stack with slightly distended saccules (S)
containing casein submicelles and at proximity some small vesicles
(arrow).These vesicles do not contain casein submicelles. The asterisk
indicates a densification of the cytoplasmic matrix surrounding the
small vesicles located between the endoplasmic reticulum (ER) and
the cis-element of the stack. x 48,750.
Fig. 9. a: Low-power photograph of the apex of a glandular cell next to the lumen (Lu) showing large
secretory granules containing casein micelles suspended in a fluid containing a wispy material. Two
granules present an image of fusion (FG).A small portion of a is magnified in b to show the honeycombed
texture of the casein micelles (arrows) within a secretion granule. a, X 35,100; b, x 55,000.
cells of the seminal vesicle, in which the secretory granules seen on the trans side of the stack may be
granules are particularly large (Clermont et al., 19921, considered (Fig. 10). First, the 20 nm submicelles may
the trans-Golgi network is limited to a few residual be transported by means of the small 80 nm vesicles
membranous tubules as observed here in the glandular seen a t the edges of the saccules as postulated by the
cells of lactating mammary gland. The corollary of this vesicular transport hypothesis (see Introduction). Secobservation is that the whole trans-most Golgi element ond, the submicelles may be transported by the sacmust be continually and rapidly renewed as the forma- cules themselves, as the latter would migrate from the
tion of secretion granules in these cells has been shown cis to the trans face of the stack as originally postulated
to take place in <1h r (Hollman, 1974; Keenan et al., by several electron microscopists in early studies on
the Golgi apparatus of various cell types (Morre et al.,
Finally, the large prosecretory granules appear to 1971, 1979; Morre and Mollenhauer, 1974; and see relose some membrane via the formation of coated buds views Whaley and Dauwalder, 1979; Goldfischer, 1982;
(Fig. 2), which would explain their slight reduction in Keenan et al., 1974; Mather and Keenan, 1983). The
volume as they transform into secretory granules. The possibility that casein submicelles flow from one saceventual increase in size of some secretory granules a s cule to the next via intermittent connections between
they approach the apex of the cell appears to be the saccules or a s recently proposed by Mellman and Siconsequence of their fusion with one another (Fig. 10). mons (1992) via tubules bridging adjacent stacks and
connecting saccules a t different levels within these
Transport of Submicelles From the Cis to the
stacks will not be considered for the time being. Indeed,
Trans Aspect of the Golgi Stack
intersaccular connections in the cis-trans axis were not
Two pathways for the intra-Golgi transport of casein readily observed in thin sections, and intersaccular tusubmicelles from the cis-element to the prosecretory bular regions similar to those already described for the
Fig. 10. Diagram summarizing the observations on the general appearance of the various components of a Golgi stack of glandular cells
of a lactating mammary gland. The casein submicelles (CSM) were
observed in the cis-element (CE) and the underlying Golgi saccules
(GS). On the trans aspect of the stack, a distended prosecretory granule (Pg) shows linear aggregates of casein submicelles and denser
irregular compact aggregates (CSMA). In the secretory granules (Sg),
the casein submicelles are compacted within spherical casein micelles
(CM), which have a characteristic honeycombed texture. Some secretory granules fuse (FSg) as they migrate toward the cell apex to be
exocytosed. Characteristically, the small vesicles (v), 80 nm in diameter or larger, are free from casein submicelles. Some coated vesicles
(CV) seemingly bud or fuse with prosecretory granules. The “well”
(W) is a pan-shaped cavity within the stack of saccules formed by
fenestrations in register with the mouth directed toward the cis side
of the stack. These wells, which are rare in the present Golgi apparatus, usually contain small vesicles. The fact that the small 80 nm
vesicles seen a t the edges of the saccules do not seemingly contain
casein submicelles suggest that these vesicles do not serve as carriers
of submicelles from one saccule to the next in a cis-trans direction. As
a corollary, the saccules themselves may transport the casein submicelles from the cis to the trans face of the stack and be turned over in
the process. ER, cisternae of the rough endoplasmic reticulum.
Golgi apparatus of a wide variety of cell types (Rambourg and Clermont, 1990) were seemingly absent
from the Golgi apparatus of secretory cells of lactating
mammary glands.
The fact that the small vesicles seen at the margins
of the saccules were rarely if ever found to contain
casein submicelles, as verified by stereoscopy, appears
to disqualify such structures as potential carriers of
casein submicelles. The possibility that casein submicelles would disintegrate into their molecular subcomponents to be transported by the vesicles and then re-
constitute as a submicelle in the next saccule, and this
occurring repetitiously, could always be considered but
does not seem to be plausible. It is thus suggested that
the saccules themselves serve as carriers of caseins
from the cis to the trans side of the stack (Fig. 10).
A similar conclusion was reached in a recent electron
microscopic study of the formation of secretory granules in glandular epithelial cells of the seminal vesicle
(Clermont et al., 1992). In this particular Golgi apparatus, the proteins that form the dense core of the definitive secretory granule appeared in the cis-element
of the stack in close association with its membrane, a n
association maintained in all subjacent saccules and
the secretory granules themselves. These dense proteinaceous condensations were thus considered a s a
membrane marker. The analysis of their distribution
in the Golgi saccules and complete absence from the
small 80 nm vesicles led to the suggestion that the
saccules themselves were involved in their transport
from the cis side of the stack to the trans-located secretory granules (Clermont et al., 1992).
Such a conclusion is also supported by several electron microscopic studies of scale formation in the Golgi
apparatus of algae (McFadden and Melkonian, 1966;
McFadden et al., 1986; Melkonian et al., 1991). In these
cells, the numerous saccules of the Golgi stacks all contain scales a t various stages of formation, while the
vesicles seen a t their edges were found to be free from
the electron-dense scale material. These various observations, including those of the present study, therefore
favor the model of saccular migration to explain the
transport of secretory proteins from the cis-element to
the trans-located prosecretory granule.
Regarding the role of the small vesicles seen in association with the Golgi apparatus, it is likely that
some vesicles seen on the cis face of the stack may serve
as carriers of proteins from the proximal ER cisternae
to the cis-element, either directly or through a n intermediate compartment, while others may serve to retrieve membrane from the Golgi saccules to the ER (as
reviewed by Mellman and Simons, 1992). The role of
the vesicles seen a t the margin of the saccules is more
problematical (Mellman and Simons, 1992). In the case
of the glandular cells of lactating mammary glands,
they may be involved in the delivery of other milk proteins to the saccules or serve in the retrieval of membrane by the ER. Finally, the coated or uncoated vesicles seen on the trans aspect of the Golgi stack may
represent residual elements that budded from the
prosecretory granules and either be recycled to the ER
or be eliminated by their incorporation into the multivesicular bodies, autophagosomal in nature, seen in
the Golgi region. In conclusion, the present study presents morphological observations that would favor the
saccular migration model and as a consequence would
indicate that all elements of the stack are actively renewed in the process of secretory granule formation.
This work was supported by grants of the MRC to
Y.C. and L.H. and of the NSERC of Canada to J.D.T.
L.X. was supported by The National Education of the
Republic of China. The technical assistance of Ms.
Jeannie Mui and Ms. Matilda Cheung is gratefully acknowledged.
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