Actin filament cables in Drosophila nurse cells are composed of modules that slide passively past one another during dumping - PubMed (original) (raw)
Actin filament cables in Drosophila nurse cells are composed of modules that slide passively past one another during dumping
G M Guild et al. J Cell Biol. 1997.
Abstract
At a late stage in Drosophila oogenesis, nurse cells rapidly expel their cytoplasm into the oocyte via intracellular bridges by a process called nurse cell dumping. Before dumping, numerous cables composed of actin filaments appear in the cytoplasm and extend inward from the plasma membrane toward the nucleus. This actin cage prevents the nucleus, which becomes highly lobed, from physically blocking the intracellular bridges during dumping. Each cable is composed of a linear series of modules composed of approximately 25 cross-linked actin filaments. Adjacent modules overlap in the cable like the units of an extension ladder. During cable formation, individual modules are nucleated from the cell surface as microvilli, released, and then cross-linked to an adjacent forming module. The filaments in all the modules in a cable are unidirectionally polarized. During dumping as the volume of the cytoplasm decreases, the nucleus to plasma membrane distance decreases, compressing the actin cables that shorten as adjacent modules slide passively past one another just as the elements of an extension ladder slide past one another for storage. In Drosophila, the modular construction of actin cytoskeletons seems to be a generalized strategy. The behavior of modular actin cytoskeletons has implications for other actin-based cytoskeletal systems, e.g., those involved in Listeria movement, in cell spreading, and in retrograde flow in growth cones and fibroblasts.
Figures
Figure 1
Diagram of nurse cell dumping during the late stages of Drosophila oogenesis. During stage 10B (4 h), the collective nurse cell cytoplasm (dark stippling) accounts for half the volume of the egg chamber, whereas the oocyte accounts for the other half (light stippling). The polytene nurse cell nuclei are shown as circles whereas the nurse cell plasma membranes are omitted for clarity. During stage 11 (30 min), the nurse cells contract and dump their cytoplasmic contents into the oocyte. As a result, nurse cell volume is reduced relative to oocyte volume. Stage 12 egg chambers (2 h) are characterized by a relatively small nurse cell component still containing nuclei.
Figure 2
Portions of wild-type stage 10B egg chambers stained with rhodamine-conjugated phalloidin and examined by confocal microscopy. (a) Cortical actin staining outlines 7 of the 15 nurse cells (left) and the anterior portion of the oocyte (right). Two ring canals (rc) that connect the oocyte to the nurse cells are also indicated (arrows). The nurse cells contain prominent arrays of actin bundles extending from the plasma membrane toward the nucleus (dark regions within the nurse cells). These cables are located on plasma membranes adjacent to the oocyte (1), adjacent to other nurse cells with a common ring canal (2), adjacent to other nurse cells without a ring canal connection (3), and adjacent to follicle cells surrounding the egg chamber (4). The densities of these cables along the nurse cell plasma membrane are indicated in Table I. This is a 14-μm optical section. (b) The actin cables originate from the plasma membrane and extend toward and outline the nucleus (dark region). The striated actin cables exhibit gaps at low (leftward arrows), medium (upward arrowheads), and high (downward arrowheads) frequencies. In addition, the F-actin staining intensity in the gaps between the striations is often well above background. Also, the fluorescent intensity of adjacent modules in the same cable can be quite different (leftward arrows) and probably reflects the presence of overlapping actin bundles. The oocyte (not shown) of this egg chamber is connected to this nurse cell. This is a 6-μm optical section. Bars: (a) = 100 μm; (b) 10 μm.
Figure 3
Portions of singedX2 stage 10 egg chambers stained with rhodamine-conjugated phalloidin and examined by confocal microscopy. (a) The cortical actin and two ring canals between three nurse cells are shown. There is a wide variety of ring canal morphologies in snX2 egg chambers (unpublished observations). The ring canals shown here look nearly wild type in appearance. (b) The cortical actin and striated actin cables between two nurse cells are shown. For comparison, a wild-type nurse cell at approximately the same stage is shown in Fig. 2_b._ These are 6- (a) and 7-μm (b) optical sections. Bars: (a) 10 μm; (b) 5 μm.
Figure 4
Thin-sections through wild-type and mutant nurse cell nuclei. (A) At stage 10B a wild-type nucleus is nearly round and contains large dense masses that superficially resemble nucleoli. (B) In contrast, at stage 11 the wild-type nuclear surface is indented and lobed. The nucleoplasm is much denser with numerous areas of heterochromatin. Printed at the same magnification as a. (C) Thin section through a portion of two nurse cells from the singedX2 mutant at stage 11. In this section a large lobe from the nucleus (N) has entered the ring canal, whose margins are indicated by the arrows. This nuclear lobe has effectively plugged the canal. In the inset we have diagrammed what this looks like at lower magnification as in the micrograph. The margins of the ring canals are indicated by the arrows. Bars: (A) 10 μm; (C) 1 μm.
Figure 5
Thin-sections through a portion of the surface of a wild-type nurse cell. (A) Extending basally from a microvillus (arrow) is an actin filament bundle. This bundle, as it extends basally, is connected to other actin bundles that run parallel to it. (B) Thin-section cut deeper in the cytoplasm. This micrograph illustrates that actin bundles associate laterally with other bundles. Although it cannot be proven from these two micrographs, individual bundles appear short (e.g., 2–3 μm) but overlap like the units of an extension ladder. Both A and B are cut from stage 11 follicles and printed at the same magnification. Bar, 1 μm.
Figure 6
Serial thin-sections through a portion of the cytoplasm of a wild-type stage 11 nurse cell. Of interest here is that short actin bundles associate laterally to form a single continuous cable. One end of the cable extends basally from a microvillus and associates with a second bundle that, in turn, associates with a third. From these serial sections and one not included, we know that bundle 2 begins and ends within the cytoplasm proper and is not attached to the membrane. The drawing on the right illustrates the bundles and how they are associated with each other. Bar, 1 μm.
Figure 7
Higher magnification views of thin-sections through cables composed of small bundles. These sections are cut from wild-type stage 11 follicles. All are printed at the same magnification. (A) The horizontal striations present on each bundle are due to the fascin crossbridges as described by Tilney et al. (1995). The striations of adjacent bundles are in transverse register (arrows). (B) Three nearby bundles seen in transverse section. (C) Another cable composed of a small bundle, a medium-sized bundle, and a large bundle. We suspect the large bundle may be composed of two cross-linked small bundles. Bar, 0.1 μm.
Figure 8
Bar graphs illustrating the number of actin filaments per bundle. Filament number was determined by counting individual filaments in electron micrographs similar to those shown in Figs. 7 (B and C) and 12 B. (a) The number of actin filaments in 55 bundles from wild-type stage 11 egg chambers was counted and their distribution was plotted. The average number of filaments per bundle ± SEM is given. (b) The number of actin filaments in 42 bundles from wild-type stage 12 egg chambers was counted and their distribution was plotted. Peaks at approximately 50, 75, and 100 filaments per bundle reflect the highly overlapping arrangement of these bundles.
Figure 9
Thin-section through a portion of a nurse cell from a wild-type stage 11 follicle that was detergent extracted and decorated with S1 of myosin before fixation. It is interesting to note that adjacent bundles have an identical polarity as determined by the S1 arrow. In short, the barbed ends are found nearest the plasma membrane. The arrows on the micrographs reflect the polarity of the myosin S1 arrow and point toward the pointed end (plus end) of the actin filaments. Bar, 0.5 μm.
Figure 10
Portions of wild-type egg chambers stained with rhodamine-conjugated phalloidin and examined by confocal microscopy. Regions containing many overlapping bundles are shown. (a) Stage 10B. Gaps emphasizing the striations (arrows). (b) Stage 11. Gaps emphasizing the striations (arrows) are more frequent per unit length than in earlier stages. In addition, F-actin staining intensity in the gaps is well above background. (c) Stage 12. Here many actin bundles have been compressed and have coalesced to form high density and relatively striation-free bundles. Bars, 5 μm.
Figure 11
(A) Portion of a kelchneo stage 10 egg chamber stained with rhodamine-conjugated phalloidin and examined by confocal microscopy. A number of striated actin bundles can be seen (arrowheads). Of interest here are the distorted regions characterized by angular intersections of relatively straight bundles (arrows). Similar distortions can be seen in wild-type egg chambers, especially where the actin bundles contact the nuclear envelope (e.g., Fig. 2_b_). This is a 3-μm optical section. (B) Thin-section through the portion of the nurse cell cytoplasm from a kelchneo stage 11 egg follicle. Of interest is that the cable appears broken. This occurs at the point of overlap of two modules. Bars: (A) 5 μm; (B) 0.1 μm.
Figure 12
Thin-sections both longitudinal (A) and nearly transverse (B) of the actin filament bundles and/or cables that reside in nurse cells after dumping. These are wild-type stage 12 follicles. Note that the bundles lie nearly parallel to each other. Because the volume is so reduced after dumping, adjacent bundles overlap to a large extent.
Figure 13
Extension ladder model for shortening actin cables in Drosophila nurse cells. During stage 10B of oogenesis (top), overlapping actin bundles (cables) extend from the plasma membrane toward the lobed nucleus. These cables are in their extended form much like an extension ladder at its maximum length. During nurse cell dumping (stage 11, middle), cell volume is reduced but nuclear volume is not. As the distance between the nuclear envelope and the plasma membrane is reduced, individual bundles in the actin cables are forced to slide past one another, reducing the overall length of the cable. After dumping (bottom), the nuclear envelope to plasma membrane distance is reduced to a minimum, resulting in maximal overlaps among the actin bundles. These cables are in their most compressed form much like an extension ladder in its most retracted form where modules exhibit maximum overlap.
Figure 14
Shortening actin cables keep ring canals obstruction-free during nurse cell dumping. Wild-type nurse cells (left) are characterized at stage 10 by the formation of overlapping actin bundles extending from the plasma membrane toward the nucleus. During stage 10B the nucleus becomes lobed. During nurse cell dumping (stage 11), cytoplasmic volume is reduced considerably by export through the ring canals (arrows). The nuclear lobes are prevented from entering and plugging the ring canals by the passively collapsible network of actin cables. Nurse cells in singed egg chambers fail to assemble this network of actin cables. As a result, cytoplasmic transport leads to nuclear plugging of the ring canals, reducing the amount of cytoplasm transferred to the oocyte.
References
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