/. Embryol. exp. Morph. Vol. 38, pp. 125-138 1977
Printed in Great Britain
125
Recent findings on oogenesis of Drosophila
melanogaster
II. Further evidence on the origin of yolk platelets
By F. GIORGI 1 AND J. JACOB
From the Institute of Animal Genetics, University of Edinburgh, Scotland
SUMMARY
Vitellogenic ovaries from Drosophila melanogaster flies have been exposed, either in in vivo
or in vitro conditions, to various extracellular tracers in an attempt to determine the possible
route of entry of the yolk precursors.
Ruthenium red and lanthanum nitrate have been shown to gain access to the oocyte surface
by initially passing through the intercellular spaces of the follicle layer. Both these tracers,
however, never attain an intracellular location within any of the cells forming the ovarian
chamber.
Colloidal Thorotrast when injected into adult females has never been detected within any
of the ovarian chambers examined, irrespective of their stage.
Vitellogenic oocytes exposed to peroxidase in in vivo conditions exhibit the oolemma and
all the structural elements present in the cortical ooplasm well labelled within a very short
time after the injection. Moreover, with gradually increasing exposure times to peroxidase,
the labelled yolk platelets increase progressively in number. At each time interval after the
injection, the label over the yolk platelets remains restricted to the superficial layer and never
gets into the associated body.
The pattern of tritiated lysine incorporation into vitellogenic oocytes has been studied over
a period of 20 h. A few hours after injection of the radioactive tracer, the silver grains located
over the ooplasm appear distributed at random. A predominant labelling of the yolk platelets
as compared to the rest of the ooplasm, becomes evident only with a 6 h delay since the
time of injection.
When analysed by electrophoresis and isolectrofocusing, the vitellogenic ovary is seen to
exhibit a number of protein bands which are common to those of other tissues as, for instance,
haemolymph and fat body.
The evidence obtained in the present study is discussed in relation to the hypothesis of an
extraovarian origin of the yolk precursors and their sequestration into forming yolk platelets.
INTRODUCTION
In the previous paper (Giorgi & Jacob, 1977) we have shown how the ultrastructure of the cortical ooplasm in developing oocytes of Drosophila melanogaster suggests a mechanism for the extraovarian origin of yolk material. It
was assumed that yolk precursors were taken in from the external medium
by means of oolemma-derived vesicles and tubules and that these led to the
1
Present address: Istituto di Istologia ed Embriologia, Via A. Volta 4, 56100 Pisa, Italy.
9
EMB 38
126
F. GIORGI AND J. JACOB
formation of yolk platelets. However, the limitations arising from a study of
purely static pictures constitute a well known problem, and it would be desirable
to substantiate any hypothesis so formulated by means of an experimental
approach. First of all it seems necessary to demonstrate that vitellogenic
oocytes can take in material from the external medium and sequester it into
forming yolk platelets. Further, if indeed such a route of entry exists in
vitellogenic oocytes, then it is also important to show that the material which
comes to be present in the yolk platelets has actually been taken up as such and
is not processed by the protein synthesizing machinery of the oocyte. Finally,
as further corroboration, it may be useful to check if there is a similarity in the
protein content of the yolk and that of the haemolymph.
The experiments reported in the present paper were designed to clarify the
points mentioned above. A description of these follows.
MATERIAL AND METHODS
Cytochemical experiments
In order to establish a possible route of entry in vitellogenic oocytes, ovaries
from 2- to 3-day-old flies were fixed for 2 h at 4 °C in 5 % glutaraldehyde in
0-1 M cacodylate buffer at pH 7-2 with ruthenium red or lanthanum nitrate
added (0-1 g/lOml of fixative). After fixation the ovaries were rinsed several
times in the buffer and then postfixed for 3 h in 1 % osmium tetroxide made up
in 0-1 M cacodylate buffer at pH 7-2 with or without the same tracers added. The
postfixed ovaries were then dehydrated in a graded series of alcohols, passed
through propylene oxide and finally embedded in Epon-Araldite mixture.
In another series of experiments, a 1 % solution of thorium dioxide (Thorotrast, Taab) in Drosophiia Ringer was injected into 2- to 3-day-old flies; a minimum
amount of 0-5 /A was injected into each fly. At various intervals after the
injection, the ovaries were dissected out and fixed as before. A few ovaries were
exposed to 1 % solution of thorium dioxide in 30 % acetic acid after fixation in
glutaraldehyde. The ovaries treated this way were then postfixed, dehydrated
and embedded as before.
To follow the process of uptake of exogenous material in vitellogenic oocytes
in physiological conditions, a 0-5 % solution of peroxidase (Sigma) in Drosophiia
Ringer was injected into 2- to 3-day-old flies; each fly received at least 0-5 /d of
the solution. Alternatively a few ovaries were freshly dissected and exposed in
in vitro conditions (either in Drosophiia Ringer or cultured in a medium recently
worked out by Shields, Dubendorfer & Sang, 1975) to a peroxidase solution
which has the same concentration as in vivo. At various intervals of time after
the injection or after the beginning of the culture experiment, the ovaries were
fixed in 5 % glutaraldehyde in 0-1 M cacodylate buffer at pH 7-2 for 2h at 4 °C.
This was then followed by a 30-min incubation in the diamino-benzidine (DAB)
medium in Tris-HCl (Graham & Karnovsky, 1966) at pH 7-6 with 0-1 ml of H2O2
Oogenesis of Drosophila. / /
127
added. Following several changes in the same buffer, the ovaries were finally
postfixed in 1 % osmium tetroxide in 0-1 M cacodylate buffer, pH 7-2 for 3 h
at 4 °C, dehydrated in alcohol and embedded as usual in Epon-Araldite
mixture.
Autoradiographic experiments
3
[ H] Lysine (Radiochemical Centre, Amersham) was incorporated into developing oocytes by two different methods: in vitro incubation and in vivo microinjection. In the first case, ovaries dissected from 2-to 3-day-old flies were
cultured in tissue culture medium 199 (Bio-cult) which contained radioactive
lysine at the concentration of 100/*Ci/ml (specific activity 4-6 Ci/mM). The
ovaries were cultured in the radioactive medium for 30, 60 or 120 min and then
fixed in 5 % formaldehyde in 0-1 M cacodylate buffer, pH 7-2. In a few cases, the
ovaries were transferred to a non-radioactive medium containing lysine and
chased for various lengths of time. In vivo experiments were performed by
injecting a minimum amount of 0-5 JLL\ per fly of [3H]lysine in Drosophila Ringer
at the concentration of 1 /tCi//d. Ovaries were then dissected at various intervals
and fixed as in the previous case. Following postfixation in 1 % osmium
tetroxide and embedment in Epon-Araldite mixture, thick sections were cut and
placed on glass slides for light microscope autoradiography. The slides were
then coated with liquid llford L4 emulsion and exposed for various lengths of
time in a light-proof box.
For electron microscope autoradiography, thin sections were cut in a MT-1
Porter-Blum ultramicrotome and collected on nickel grids previously covered
with Formvar and carbon. The sections were then covered with a pregelled
layer of liquid llford L4 emulsion according to the coating technique of Caro &
Van Tubergen (1962) as modified in more recent years (Jacob & Budd, 1975).
After coating, the grids were stored in a light-proof box for periods of time
ranging from 1 week to 2 months. At the end of the exposure periods, the grids
were developed in Kodak D 19b developer and fixed in freshly prepared
Kodak F-24 fixative. Following double staining in uranyl acetate and lead citrate,
the grids were observed in a AEI-EM6 electron microscope working at 60 kV.
The auto radiographs were studied by computing the grain density for each
selected area as a function of the duration of exposure to the tracer. Two areas
were distinguished in each oocyte: the yolk and the yolk-free ooplasm. The
ovarian chambers exposed to the radioactive lysine were staged at the time of
fixation. Therefore, the actual stage of the ovarian chamber at the beginning of
the experiment could only be determined by using the time scale chart of
ovarian development of Drosophila melanogaster worked out by David &
Merle (1968).
9-2
128
F. GIORGI AND J. JACOB
Electrophoresis and isoelectrofocusing experiments
For the electrophoretic analysis of various tissues of Drosophila flies, the disc
electrophoretic system on acrylamide gels (Davis, 1964) was used. Both the
ovary and the fat body were homogenized in a 0-5 ml capacity homogenizer in
a 20 % sucrose solution. The haemolymph was collected from a number of
adult females or third instar larvae by puncturing the specimens with dissecting
needles and collecting the released liquid in a 20 % sucrose solution.
For isoelectrofocusing, samples were collected in much the same way as in the
previous case, but once homogenized, they were placed in a solubilizing solution
made according to the formula of Miner & Heston (1972). The acrylamide gels
contained an ampholine carrier with a range of pH from 3-5 to 10; the gels were
run overnight at an average voltage of 500 V. In one experiment, both ovary and
haemolymph were collected from flies previously injected with [14C]amino
acids mixture (Radiochemical Centre, Amersham). Acrylamide gels obtained
after running the radioactive samples were cut, dried and autoradiographed
according to the technique of Fairbanks, Levinthal & Reeder (1965) using
Kodirex films. Autoradiograms and gels were then scanned with a Joyce
microdensitometer and the resulting profiles presented diagrammatically.
RESULTS
Following fixation in glutaraldehyde with ruthenium red or lanthanum nitrate
added, ovarian chambers of all stages exhibited electron-dense deposits in
various places. These were the interspaces between adjacent follicle cells (Fig. 1),
between nurse cells and also at the follicle-oocyte border (Fig. 2). While the
follicle cell interspaces were filled with the tracer at all the stages examined, the
follicle-oocyte border becomes labelled only in stages preceding stage 10, that is
the stage when the formation of the vitelline membrane is completed. In no case,
however, were the tracers detected intracellularly in any of the cells forming the
ovarian chamber.
When Thorotrast was used on glutaraldehyde-fixed ovarian chambers, electrondense deposits of this colloidal tracer appeared along the outer margin of the
basement lamina of all chamber stages (Fig. 3). In no case was Thorotrast
observed within the chamber, i.e. the interspaces of the follicle layer or the
follicle-oocyte border. Even in chambers that were fixed several hours after the
injection of Thorotrast there was no indication of Thorotrast in any location
within the chamber. In contrast to this, two types of cells appeared to acquire
Thorotrast intracellularly within a few hours of in vivo exposure to the tracer.
These are the lumen cells or plasmatocytes and the follicle cells which are known
to remain at the bottom of the ovary during deposition of the mature egg (Fig. 4).
Both these cells, it may be pointed out, are characterized by the lack of a basement lamina.
Oogenesis o/Drosophila. / /
129
«
Fig. 1. The follicle layer from a stage-9 ovarian chamber fixed according to the
lanthanum nitrate method. Note the presence of heavy deposits of lanthanum in the
follicle cell interspaces (arrows), x 8000.
Fig. 2. The follicle-oocyte border from a stage-9 ovarian chamberfixedaccording to
the procedure of ruthenium red. Note the presence of a densely stained layer over
the oocyte surface, x 12000.
Fig. 3. Part of the anterior region of a stage-10 ovarian chamber fixed after in vitro
incubation with colloidal thorium dioxide. Colloidal particles adhere to the surface
of the epithelial sheath (Es) and the basement lamina (bl). Follicle cell (FC); Nurse
cell (NC). x 10000.
Fig. 4. Part of the cytoplasm of a follicle cell at the base of the ovary. The ovary was
fixed 30 h after the injection of thorium dioxide. Note the heavy deposits of colloidal
thorium within the residual bodies (rb). x 10000.
130
F. GIORGI AND J . J A C O B
Fig. 5. The cortical ooplasm of a stage-8 ovarian chamberfixed30 min after the injection of peroxidase. The follicle-oocyte border contains the tracer. Several coated
vesicles (cv) and small yolk platelets (y) are labelled. Other large yolk platelets are
not labelled. Follicle cell (FC); ooplasm (00). Unstained section, x 12000.
Fig. 6. The central ooplasm from a stage-10 ovarian chamber fixed 24 h after the
injection of peroxidase. Note that all the yolk platelets are labelled. The tracer is
restricted to the superficial layer of the yolk platelets. Unstained section, x 12000.
Fig. 7. A yolk platelet from the central ooplasm of a stage-10 ovarian chamber fixed
24 h after the injection of peroxidase. The associated body (Ab) is devoid of the
tracer, x 10000.
Fig. 8. The cortical ooplasm from a stage-12 ovarian chamber fixed 30 min after the
injection of peroxidase. Note the peroxidase reaction products in the endoplasmic
reticulum {ER). x 10000.
Oogenesis of Drosophila. / /
131
The physical continuity which is envisaged after the use of ruthenium red and
lanthanum nitrate must be interpreted cautiously. In fact, the lack of tracer in
any intracellular location may well be attributed to the fact that these tracers
were used during fixation. Under these conditions, a cell cannot be expected to
be capable of active uptake. A substance which overcomes these limitations is
peroxidase which can be supplied to the tissue in in vivo conditions, and can then
be detected cytochemically following fixation.
In ovarian chambers of vitellogenic stages, the reaction products due to the
peroxidase are observed not only along the follicle-oocyte border, but also in
several micropinocytotic vesicles which apparently are not in continuity with
the oolemma. In addition, small yolk platelets in the cortical ooplasm also
became labelled with peroxidase (Fig. 5). It could also be noted that with
progressively longer periods of in vivo exposure to the peroxidase a larger number
of yolk platelets became labelled. There was, however, no indication of a regular
layer of labelled yolk platelets. Instead, they were randomly distributed among
unlabelled platelets. Such a distribution is presumably a consequence of the
rhythmical contractions of the epithelial and peritoneal sheaths which bring
about a reshuffling of the yolk platelets. In stage-10 ovarian chambers, examined
in ovarioles that were exposed to peroxidase for 24 h, practically every yolk
platelet contained peroxidase reaction products (Fig. 6). This might be taken to
indicate that a 24-h period covers the period of vitellogenesis from its inception
in stage 8 to stage 10. This observation also tends to confirm that the process of
pinocytosis per se is unaffected by exposure to peroxidase.
As described in the previous paper of the present series (Giorgi & Jacob,
1976), the yolk platelets in stage-10-12 chambers are predominantly of the type
referred to as alpha-1. When these yolk platelets were examined in ovarioles
exposed for 24 h to peroxidase, the reaction products were found to be confined
to their superficial layer. In sections passing through the associated body, it was
obvious that this body is largely free of peroxidase reaction products (Fig. 7).
In ovarian chambers of stages later than 10, no reaction products could be
detected in the yolk platelets of the cortical ooplasm. This may be taken to
indicate that chambers in stages 8-10 are the only ones to absorb proteins from
the haemolymph and to sequester them into forming yolk platelets. In fact,
when the vitelline membrane is fully formed, no further uptake seems to occur.
Nurse cells do not exhibit peroxidase reaction products for the major part of
oogenesis, that is up to stage 10. During this period the tracer is restricted to the
intercellular spaces between these cells. Reaction products, however, appear in
the endoplasmic reticulum of nurse cells and oocytes (Fig. 8) in ovarian
chambers from stage 11 onwards. A similar reaction is observable even in stage-11
and later ovarian chambers when incubation is carried out in the diaminobenzidinemedium without priorexposure to peroxidase. It may thus be concluded
that the reaction observed in ovarian chambers of these stages is due to an
endogenous activity. In similar control experiments of vitellogenic stages,
132
F. G I O R G I AND J. JACOB
10
Fig. 9. Light microscope autoradiography of the ooplasm from a stage-10 ovarian
chamber fixed 1 h after injection of tritiated lysine. Note the heavy localization of
grains along the vitelline membrane (Vm). Follicle cells (FC); ooplasm (00).
x 1500.
Fig. 10. An EM autoradiographic preparation of the central ooplasm of a stage-10
ovarian chamber taken from an ovariole fixed 20 h after the injection of tritiated
lysine. Note that all yolk platelets are heavily labelled. The autoradiographic
exposure was 7 days, x 8000.
reaction products were not observed in any of the sites found to be labelled
after in vivo exposure to peroxidase.
It may be recalled here that ovarian chambers of vitellogenic stages exposed
in vitro to peroxidase did not yield any labelled yolk platelet even when treated
for periods of time that in in vivo conditions resulted in a high number of labelled
platelets. Negative results were obtained when the ovaries were incubated in
Ringer or in complete culture medium (Shields et al. 1975). Further experiments
are required to clarify the differences noted between the in vitro and in vivo
experiments with peroxidase.
The results obtained so far indicate that vitellogenic oocytes may acquire
yolk precursors from the haemolymph. To obtain further corroboration, the
pattern of tritiated lysine incorporation into vitellogenic oocytes was studied by
light and electron microscope autoradiography. Experiments were carried out
both in in vivo and in vitro conditions.
In stage-9-10 ovarian chambers examined 1 h after the injection of the tracer,
the grain density over the ooplasm was low and the silver grains were distributed
Oogenesis of Drosophila. / /
133
Table 1. The density of autoradiographic grains in stage-10 ovarian
chambers exposed to tritiated lysine for various lengths of time
The values of grain density for each time are expressed as number of grains counted
over an area measured in /*rn2. The standard error for each value is indicated.
Region of cell
Time (h)
Yolk platelets in
ooplasm (dy)
Yolk-free
ooplasm (do)
1
2
6
10
16
20
208 ±7-8
41-70±23-8
76-62 ±25-4
lll-99±26-6
54-11 ±13-8
7000 ±16-6
4-39±3-0
19-47 ± 5 0
6-44 ±2-6
16-99 ±2-4
2-78±l-8
3-45 + 2-0
i
*
{
10
16
X -dy
O -do
1 2
20
Fig. 11. Diagrammatical representation of the data given in Table 1. The points are
plotted with a ±2x standard error, (x), dy; (O), do.
at random (Fig. 9). By contrast, the cytoplasm of the follicle cells of these
chambers exhibited a higher density of silver grains with the highest concentration over the vitelline membrane. Grains were distributed fairly uniformly
over yolk platelets and yolk-free ooplasm even 2 h after the initial injection of
tritiated lysine. From 2 to 6 h, however, the number of grains began to increase
134
LHm
F. GIORGI AND J. JACOB
AHm
Ov
FB
009-010
0 16-021
0-37
0-50-0-58
0-55-0-59
0-77-0-83
AR&
Ov
12
13
Fig. 12. Electrophoretic analysis of several tissues from larvae and adults of
Drosophila. {LHm), Larval haemolymph; {AHm), adult haemolymph; (Ov) ovary;
{FB) Fat body; {RF) ratio between the distance calculated for each major band from
the origin of the gel and the distance of the track dye band. Protein bands are
stained with Coomassie blue.
Fig. 13. Isoelectrofocusing of the haemolymph {AHm) and ovary (Ov) extracted
from adults of Drosophila 24 h after injection of a mixture of [14C]labelled amino
acids. {ARG) Autoradiography on the dried gels.
predominantly over the yolk platelets of stage-9-10 chambers. Further, the
number of yolk platelets which become labelled increased steadily for up to
about 10 h after injection of the tracer (Table 1).
Ovarian chambers of stage 9-10 fixed 16 or 20 h after injection showed most
of the yolk platelets labelled (Fig. 10). The overall grain density over the yolk
continued to be markedly higher than that on the yolk-free ooplasm, but by
this time there was a noticeable decline compared with that at the 10 h time
interval (Fig. 11). The reason for this drop in grain density remains unclear;
a plausible explanation will be proffered in the next paper of this series.
Oogenesis 0/Drosophila. / /
135
The pattern of labelling in vitellogenic oocytes (stage-9-10 chambers) exposed
for up to about 2 h in in vitro conditions to [3H]lysine was more or less similar
to that observable in corresponding stages exposed in vivo for the same length
of time. However, in ovaries that were cultured for even up to 6 h the grain
density over the yolk area never became statistically higher than that over the
rest of the ooplasm. The autoradiographical data, therefore, indicate that the
yolk platelets become specifically well labelled only in in vivo conditions and
with a minimum time lapse of 6 h following injection.
In order to examine whether any of the proteins present in the ovary has an
extraovarian counterpart, it seemed useful to attempt an electrophoretic analysis
(Davis, 1964) of the soluble proteins of various tissues from larvae and adult
females of Drosophila. The results of this analysis are shown in Fig. 12. It can
be seen that among the protein bands which are discernible in the haemolymph
of the adult female, several of them present an electrophoretic mobility similar
to that of the proteins of the ovary and fat body.
When ovarian extracts were analysed by isolectrofocusing according to the
procedure of Miner & Heston (1972), numerous sharp bands could be resolved
within the pH range of 3-5-100. Although the number of protein bands visible
in the haemolymph was lower than that of the ovary (a situation opposite to that
obtained following disc electrophoretic analysis), in this case too a similarity in
the protein content of the two tissues was evident (Fig. 13). Autoradiographs of
acrylamide gels further show that proteins of haemolymph and ovary become
almost equally labelled 24 h after injection of labelled lysine.
DISCUSSION
The present study has shown that tracers such as ruthenium red and lanthanum
nitrate gain access to the oocyte surface by initially passing through the intercellular spaces of the follicle layer. Such a route of entry was previously
demonstrated in the oocyte of the cecropia moth by the use of fluoresceinlabelled antibodies (Telfer, 1961). Similar results have also been obtained in
vitellogenic oocytes of Aedes aegypti (Anderson & Spielman, 1971).
When Thorotrast was used as an extracellular tracer, it was found that this
colloidal tracer does not penetrate ovarian chambers of any stage, either in
in vivo or in vitro conditions. On the other hand, this tracer has been shown to
gain access to certain cells of the ovary which are devoid of the basement lamina.
It may thus be reasonably assumed that the basement lamina normally prevents
diffusion of large sized-molecules such as those of the colloidal Thorotrast. In
their study of the vitellogenesis of Aedes aegypti, Anderson & Spielman (1971)
noted that among the various tracers they used, Thorotrast was the one which
penetrated more slowly into the ooplasm. The fact that in Drosophila ovary this
tracer does not penetrate at all may be due to the fact that in this species the
basement lamina has different sieving properties from those of Aedes aegypti.
Tt must be noted that the unavoidable necessity to use the tracers mentioned
136
F. GIORGI AND J. JACOB
above, apart from Thorotrast, along with fixatives limits the possibility of
a kinetic approach to the study of absorption of substances into vitellogenic
oocytes. However, this limitation has been overcome in the present study by the
use of peroxidase in in vivo conditions. The peroxidase detected within the
vitellogenic oocytes (from stage 8 up to stage 10, which marks the completion of
the vitelline membrane) is associated with the coated vesicles, tubules and small
yolk platelets in the cortical ooplasm. With progressively longer periods of
exposure to tracer, the number of yolk platelets which become labelled gradually
increases. These data strongly suggest that proteins from the external medium
(haemolymph) can enter the oocyte at the region of the pits of the oolemma.
They further provide evidence that the tubules present in the cortical ooplasm
are oolemma-derived entities and that, in all probability, they are formed by
progressive fusion of the coated vesicles. There was never any indication of
peroxidase in the cisternae of the endoplasmic reticulum of the oocyte at the
time of pinocytotic uptake. This makes untenable the suggestion of Cummings &
King (1970) that the tubules of the cortical ooplasm in Drosophila oocytes are
derived from the endoplasmic reticulum. The latter show positive cytochemical
reaction for peroxidase in both nurse cells and oocyte only in stages later than
11. Further, control experiments have shown that the reaction in question is due
to an endogenous activity.
A further clarification of the mechanism of uptake of yolk proteins was
sought in autoradiographic experiments with tritiated lysine. Analysis of autoradiographs made at various intervals of time reveal that there is a distinctly
heavy labelling of the yolk platelets from 6 h after the injection of the tracer. In
his work on protein synthesis in the ovary of Musca domestica, Bier (1963) also
noted a considerable time-lag in the labelling of the yolk platelets. Similarly
in the honey-bee ovary, Engels (1972) showed that the yolk fraction is labelled
after a considerable delay following the labelling of other ovarian proteins. By
comparison, the level of labelling of the yolk-free ooplasm during the time-lag
discussed above is very low and this labelling could be attributed to protein
containing material, presumably ribonucleoproteins, which are transferred to
the oocyte from the nurse cells. In view of the possibility of this transfer, it is
difficult to find out if there is a low level of synthetic activity in the ooplasm.
By contrast, the cytoplasm of the follicle cells shows a heavy incorporation of
amino acids even 1 h after the injection. The labelling patterns observed in the
ovarian chambers demonstrate that, at any rate, there is no synthesis of proteins
in the ooplasm which is even remotely comparable to that observable in the
follicle cells. Further, the delayed appearance of label over the yolk platelets
could conceivably be attributed to the time required for the yolk proteins to be
synthesized in an extraovarian tissue(s) and then transferred to the oocyte
through the blood stream. The plausibility of this conclusion is also increased
by our findings that in vitro the yolk platelets never attain a grain density
comparable to that obtained in in vivo experiments.
Oogenesis of Drosophila. / /
137
The similarities in the electrophoretic patterns of the various tissues examined,
taken together with the evidence obtained from other experiments, are clearly
indicative of the occurrence of a process of uptake whereby the ovary can
sequester proteins from the haemolymph. Support for this view has also come
from the isoelectrofocusing analysis of solubilized proteins from the ovary and
the haemolymph.
As to the actual site of synthesis of yolk proteins, recent evidence obtained
from several Drosophila species (Gelti-Douka, Gingeras & Kambysellis, 1974)
would indicate that this is at the level of fat body. This organelle, in fact, has
been shown to be capable of synthesizing and realizing into the culture medium
a few proteins which are precipitated by yolk antibodies.
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