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Localized depolymerization of the major sperm protein cytoskeleton correlates with the forward movement of the cell body in the amoeboid movement of nematode sperm.

Italiano JE, Stewart M, Roberts TM - J. Cell Biol. (1999)

Bottom Line: At pH 6.35, the cytoskeleton pulled away from the leading edge and receded through the lamellipodium as its disassembly at the cell body continued.The cytoskeleton disassembled rapidly and completely in cells treated at pH 5.5, but reformed when the cells were washed with physiological buffer.These results indicate that cell body retraction is mediated by tension in the cytoskeleton, correlated with MSP depolymerization at the base of the lamellipodium.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Science, Florida State University, Tallahassee, Florida 32306, USA.

ABSTRACT
The major sperm protein (MSP)-based amoeboid motility of Ascaris suum sperm requires coordinated lamellipodial protrusion and cell body retraction. In these cells, protrusion and retraction are tightly coupled to the assembly and disassembly of the cytoskeleton at opposite ends of the lamellipodium. Although polymerization along the leading edge appears to drive protrusion, the behavior of sperm tethered to the substrate showed that an additional force is required to pull the cell body forward. To examine the mechanism of cell body movement, we used pH to uncouple cytoskeletal polymerization and depolymerization. In sperm treated with pH 6.75 buffer, protrusion of the leading edge slowed dramatically while both cytoskeletal disassembly at the base of the lamellipodium and cell body retraction continued. At pH 6.35, the cytoskeleton pulled away from the leading edge and receded through the lamellipodium as its disassembly at the cell body continued. The cytoskeleton disassembled rapidly and completely in cells treated at pH 5.5, but reformed when the cells were washed with physiological buffer. Cytoskeletal reassembly occurred at the lamellipodial margin and caused membrane protrusion, but the cell body did not move until the cytoskeleton was rebuilt and depolymerization resumed. These results indicate that cell body retraction is mediated by tension in the cytoskeleton, correlated with MSP depolymerization at the base of the lamellipodium.

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The pattern of recovery of locomotion after treatment with HKB-acetate, pH 5.5, shows that cytoskeletal depolymerization at the base of the lamellipodium is required for movement of the cell body. In this sequence, a cell, moving in the direction indicated by the black arrow in a, was perfused with pH 5.5 buffer to cause its cytoskeleton to disassemble (b). The white arrow indicates a stationary mark on the substrate. When the cell was washed with HKB buffer, MSP fiber complexes began to reassemble along the periphery of the lamellipodium, causing membrane protrusion (c, arrows). Within 15 s (d) fiber complexes regrowing from the right side of the lamellipodium reached the cell body and started to treadmill (solid black arrows); as indicated by the change in position relative to the stationary mark, the cell body moved in the direction of these treadmilling fiber complexes. At this point, there was still a gap between the fiber complexes growing from the left side and the cell body (open arrow). 15 s later (e), more of the fiber complexes on the right side were fully rebuilt (solid arrows), and treadmilling and movement of the cell body toward that side continued. By 60 s after washing, the cell locomotion resumed (f) in a new direction (bold black arrow) corresponding to the direction of recovery of cell body movement. Note that due to the movement of the cell, the position of the frame in f differs from that in a–e. Bar, 10 μm.
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Figure 5: The pattern of recovery of locomotion after treatment with HKB-acetate, pH 5.5, shows that cytoskeletal depolymerization at the base of the lamellipodium is required for movement of the cell body. In this sequence, a cell, moving in the direction indicated by the black arrow in a, was perfused with pH 5.5 buffer to cause its cytoskeleton to disassemble (b). The white arrow indicates a stationary mark on the substrate. When the cell was washed with HKB buffer, MSP fiber complexes began to reassemble along the periphery of the lamellipodium, causing membrane protrusion (c, arrows). Within 15 s (d) fiber complexes regrowing from the right side of the lamellipodium reached the cell body and started to treadmill (solid black arrows); as indicated by the change in position relative to the stationary mark, the cell body moved in the direction of these treadmilling fiber complexes. At this point, there was still a gap between the fiber complexes growing from the left side and the cell body (open arrow). 15 s later (e), more of the fiber complexes on the right side were fully rebuilt (solid arrows), and treadmilling and movement of the cell body toward that side continued. By 60 s after washing, the cell locomotion resumed (f) in a new direction (bold black arrow) corresponding to the direction of recovery of cell body movement. Note that due to the movement of the cell, the position of the frame in f differs from that in a–e. Bar, 10 μm.

Mentions: Treatment of sperm with HKB-acetate at pH 5.5 caused the entire MSP cytoskeleton to disassemble rapidly and the lamellipodium to round up (Fig. 5; see also Roberts and King 1991). This effect was completely reversible. The pattern by which the cytoskeleton was rebuilt and locomotion resumed showed that cytoskeletal assembly and disassembly produce independent forces and that both are required for locomotion. When cells treated with pH 5.5 buffer (Fig. 5, a and b) were washed with HKB buffer without acetate, fiber complexes began to form around the periphery of the lamellipodium (Fig. 5 c). This localized assembly resulted in formation of protrusions from the cell surface, but the cell body remained stationary. As these new fiber complexes continued to elongate, those emanating from the side of the lamellipodium reached the cell body and began depolymerization and treadmilling. In the cell shown in Fig. 5, the fiber complexes on the right side of the lamellipodium reached the cell body before those from the other side. When these fiber complexes began to treadmill, the cell body was pulled to that side (Fig. 5 d). Soon, additional fiber complexes growing from the right side of the lamellipodium reached the cell body and the entire cell began to move to the right before the fiber complexes from the other side were completely rebuilt (Fig. 5 e). This asymmetry in cytoskeletal reconstruction, with treadmilling resuming earlier on one side of the lamellipodium than the other, resulted in a 60° change in the direction of locomotion, compared with that observed before acid treatment. The behavior of this unusual cell emphasized that movement, first of the cell body and then the whole cell, was determined by the location where cytoskeletal depolymerization resumed along the cell body–lamellipodium junction. In most cells recovering from this acid treatment, cytoskeletal reconstruction was symmetric and the directions of movement before and after treatment were similar. In each, however, movement of the cell body did not occur until the onset of depolymerization of the reconstructed fiber complexes.


Localized depolymerization of the major sperm protein cytoskeleton correlates with the forward movement of the cell body in the amoeboid movement of nematode sperm.

Italiano JE, Stewart M, Roberts TM - J. Cell Biol. (1999)

The pattern of recovery of locomotion after treatment with HKB-acetate, pH 5.5, shows that cytoskeletal depolymerization at the base of the lamellipodium is required for movement of the cell body. In this sequence, a cell, moving in the direction indicated by the black arrow in a, was perfused with pH 5.5 buffer to cause its cytoskeleton to disassemble (b). The white arrow indicates a stationary mark on the substrate. When the cell was washed with HKB buffer, MSP fiber complexes began to reassemble along the periphery of the lamellipodium, causing membrane protrusion (c, arrows). Within 15 s (d) fiber complexes regrowing from the right side of the lamellipodium reached the cell body and started to treadmill (solid black arrows); as indicated by the change in position relative to the stationary mark, the cell body moved in the direction of these treadmilling fiber complexes. At this point, there was still a gap between the fiber complexes growing from the left side and the cell body (open arrow). 15 s later (e), more of the fiber complexes on the right side were fully rebuilt (solid arrows), and treadmilling and movement of the cell body toward that side continued. By 60 s after washing, the cell locomotion resumed (f) in a new direction (bold black arrow) corresponding to the direction of recovery of cell body movement. Note that due to the movement of the cell, the position of the frame in f differs from that in a–e. Bar, 10 μm.
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Related In: Results  -  Collection

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Figure 5: The pattern of recovery of locomotion after treatment with HKB-acetate, pH 5.5, shows that cytoskeletal depolymerization at the base of the lamellipodium is required for movement of the cell body. In this sequence, a cell, moving in the direction indicated by the black arrow in a, was perfused with pH 5.5 buffer to cause its cytoskeleton to disassemble (b). The white arrow indicates a stationary mark on the substrate. When the cell was washed with HKB buffer, MSP fiber complexes began to reassemble along the periphery of the lamellipodium, causing membrane protrusion (c, arrows). Within 15 s (d) fiber complexes regrowing from the right side of the lamellipodium reached the cell body and started to treadmill (solid black arrows); as indicated by the change in position relative to the stationary mark, the cell body moved in the direction of these treadmilling fiber complexes. At this point, there was still a gap between the fiber complexes growing from the left side and the cell body (open arrow). 15 s later (e), more of the fiber complexes on the right side were fully rebuilt (solid arrows), and treadmilling and movement of the cell body toward that side continued. By 60 s after washing, the cell locomotion resumed (f) in a new direction (bold black arrow) corresponding to the direction of recovery of cell body movement. Note that due to the movement of the cell, the position of the frame in f differs from that in a–e. Bar, 10 μm.
Mentions: Treatment of sperm with HKB-acetate at pH 5.5 caused the entire MSP cytoskeleton to disassemble rapidly and the lamellipodium to round up (Fig. 5; see also Roberts and King 1991). This effect was completely reversible. The pattern by which the cytoskeleton was rebuilt and locomotion resumed showed that cytoskeletal assembly and disassembly produce independent forces and that both are required for locomotion. When cells treated with pH 5.5 buffer (Fig. 5, a and b) were washed with HKB buffer without acetate, fiber complexes began to form around the periphery of the lamellipodium (Fig. 5 c). This localized assembly resulted in formation of protrusions from the cell surface, but the cell body remained stationary. As these new fiber complexes continued to elongate, those emanating from the side of the lamellipodium reached the cell body and began depolymerization and treadmilling. In the cell shown in Fig. 5, the fiber complexes on the right side of the lamellipodium reached the cell body before those from the other side. When these fiber complexes began to treadmill, the cell body was pulled to that side (Fig. 5 d). Soon, additional fiber complexes growing from the right side of the lamellipodium reached the cell body and the entire cell began to move to the right before the fiber complexes from the other side were completely rebuilt (Fig. 5 e). This asymmetry in cytoskeletal reconstruction, with treadmilling resuming earlier on one side of the lamellipodium than the other, resulted in a 60° change in the direction of locomotion, compared with that observed before acid treatment. The behavior of this unusual cell emphasized that movement, first of the cell body and then the whole cell, was determined by the location where cytoskeletal depolymerization resumed along the cell body–lamellipodium junction. In most cells recovering from this acid treatment, cytoskeletal reconstruction was symmetric and the directions of movement before and after treatment were similar. In each, however, movement of the cell body did not occur until the onset of depolymerization of the reconstructed fiber complexes.

Bottom Line: At pH 6.35, the cytoskeleton pulled away from the leading edge and receded through the lamellipodium as its disassembly at the cell body continued.The cytoskeleton disassembled rapidly and completely in cells treated at pH 5.5, but reformed when the cells were washed with physiological buffer.These results indicate that cell body retraction is mediated by tension in the cytoskeleton, correlated with MSP depolymerization at the base of the lamellipodium.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Science, Florida State University, Tallahassee, Florida 32306, USA.

ABSTRACT
The major sperm protein (MSP)-based amoeboid motility of Ascaris suum sperm requires coordinated lamellipodial protrusion and cell body retraction. In these cells, protrusion and retraction are tightly coupled to the assembly and disassembly of the cytoskeleton at opposite ends of the lamellipodium. Although polymerization along the leading edge appears to drive protrusion, the behavior of sperm tethered to the substrate showed that an additional force is required to pull the cell body forward. To examine the mechanism of cell body movement, we used pH to uncouple cytoskeletal polymerization and depolymerization. In sperm treated with pH 6.75 buffer, protrusion of the leading edge slowed dramatically while both cytoskeletal disassembly at the base of the lamellipodium and cell body retraction continued. At pH 6.35, the cytoskeleton pulled away from the leading edge and receded through the lamellipodium as its disassembly at the cell body continued. The cytoskeleton disassembled rapidly and completely in cells treated at pH 5.5, but reformed when the cells were washed with physiological buffer. Cytoskeletal reassembly occurred at the lamellipodial margin and caused membrane protrusion, but the cell body did not move until the cytoskeleton was rebuilt and depolymerization resumed. These results indicate that cell body retraction is mediated by tension in the cytoskeleton, correlated with MSP depolymerization at the base of the lamellipodium.

Show MeSH
Related in: MedlinePlus