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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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MSP cytoskeletal dynamics in crawling Ascaris sperm. The long branched elements that extend from the leading edge to the base of the lamellipodium are the fiber complexes, each a dense meshwork of MSP filaments. The cytoskeleton flows retrograde as the fiber complexes are assembled at the leading edge and disassembled at the cell body. Because the rates of cytoskeletal flow and locomotion are coupled, morphological markers in the cytoskeleton, such as the branch in the fiber complex indicated by the arrowhead, remain nearly stationary relative to the substrate. The fields of view are identical in each frame and the interval between frames is 10 s. Over the 30-s interval from a–d, both the leading edge and the cell body advanced by 6.5 μm while the lamellipodium maintained a length of 25 μm. This illustrates the balance between the rates of cytoskeletal assembly/leading edge protrusion and cytoskeletal disassembly/cell body retraction during sperm locomotion. Bar, 10 μm.
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Figure 1: MSP cytoskeletal dynamics in crawling Ascaris sperm. The long branched elements that extend from the leading edge to the base of the lamellipodium are the fiber complexes, each a dense meshwork of MSP filaments. The cytoskeleton flows retrograde as the fiber complexes are assembled at the leading edge and disassembled at the cell body. Because the rates of cytoskeletal flow and locomotion are coupled, morphological markers in the cytoskeleton, such as the branch in the fiber complex indicated by the arrowhead, remain nearly stationary relative to the substrate. The fields of view are identical in each frame and the interval between frames is 10 s. Over the 30-s interval from a–d, both the leading edge and the cell body advanced by 6.5 μm while the lamellipodium maintained a length of 25 μm. This illustrates the balance between the rates of cytoskeletal assembly/leading edge protrusion and cytoskeletal disassembly/cell body retraction during sperm locomotion. Bar, 10 μm.

Mentions: The dynamics of the cytoskeleton of Ascaris sperm can be observed directly in crawling cells using DIC microscopy and indicate that both MSP assembly and disassembly are important in their amoeboid motility. The MSP-based cytoskeleton in these cells form fiber complexes, each comprised of a meshwork of MSP filaments, that can be observed in living cells (Fig. 1; see also Sepsenwol et al. 1989; Roberts and King 1991). Distinctive features, such as branches in the fiber complexes, can be followed and show that the cytoskeleton treadmills as sperm locomote (see Sepsenwol et al. 1989; Roberts and King 1991). When sperm crawl, filaments assemble into fiber complexes along the leading edge and flow retrograde to the base of the lamellipodium, where they disassemble. The rates of cytoskeletal assembly and disassembly are balanced and are coupled to the pace of locomotion. Thus, as illustrated by analysis of morphological markers in the cell shown in Fig. 1, the lamellipodium maintains its shape over time while the cytoskeleton flows rearward with respect to the cell, but does not move, or moves very slowly, with respect to the substrate. This pattern of lamellipodial dynamics has also been measured by computer-assisted microscopy (Royal et al. 1995). For example, difference pictures comparing the shape of the lamellipodium of crawling sperm at two-second intervals showed that the area of the zone of expansion at the leading margin was similar to the area lost due to retraction at the base of the lamellipodium. This study also showed that the average instantaneous velocity of sperm crawling on glass was 30 μm/min, but that the cells surged forward about every 0.35 min, increasing their velocity by an average of 47%. The cells maintained their lamellipodial shape during these surges, indicating that cytoskeletal assembly and disassembly remain balanced even when crawling speed changes (Royal et al. 1995).


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)

MSP cytoskeletal dynamics in crawling Ascaris sperm. The long branched elements that extend from the leading edge to the base of the lamellipodium are the fiber complexes, each a dense meshwork of MSP filaments. The cytoskeleton flows retrograde as the fiber complexes are assembled at the leading edge and disassembled at the cell body. Because the rates of cytoskeletal flow and locomotion are coupled, morphological markers in the cytoskeleton, such as the branch in the fiber complex indicated by the arrowhead, remain nearly stationary relative to the substrate. The fields of view are identical in each frame and the interval between frames is 10 s. Over the 30-s interval from a–d, both the leading edge and the cell body advanced by 6.5 μm while the lamellipodium maintained a length of 25 μm. This illustrates the balance between the rates of cytoskeletal assembly/leading edge protrusion and cytoskeletal disassembly/cell body retraction during sperm locomotion. Bar, 10 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169480&req=5

Figure 1: MSP cytoskeletal dynamics in crawling Ascaris sperm. The long branched elements that extend from the leading edge to the base of the lamellipodium are the fiber complexes, each a dense meshwork of MSP filaments. The cytoskeleton flows retrograde as the fiber complexes are assembled at the leading edge and disassembled at the cell body. Because the rates of cytoskeletal flow and locomotion are coupled, morphological markers in the cytoskeleton, such as the branch in the fiber complex indicated by the arrowhead, remain nearly stationary relative to the substrate. The fields of view are identical in each frame and the interval between frames is 10 s. Over the 30-s interval from a–d, both the leading edge and the cell body advanced by 6.5 μm while the lamellipodium maintained a length of 25 μm. This illustrates the balance between the rates of cytoskeletal assembly/leading edge protrusion and cytoskeletal disassembly/cell body retraction during sperm locomotion. Bar, 10 μm.
Mentions: The dynamics of the cytoskeleton of Ascaris sperm can be observed directly in crawling cells using DIC microscopy and indicate that both MSP assembly and disassembly are important in their amoeboid motility. The MSP-based cytoskeleton in these cells form fiber complexes, each comprised of a meshwork of MSP filaments, that can be observed in living cells (Fig. 1; see also Sepsenwol et al. 1989; Roberts and King 1991). Distinctive features, such as branches in the fiber complexes, can be followed and show that the cytoskeleton treadmills as sperm locomote (see Sepsenwol et al. 1989; Roberts and King 1991). When sperm crawl, filaments assemble into fiber complexes along the leading edge and flow retrograde to the base of the lamellipodium, where they disassemble. The rates of cytoskeletal assembly and disassembly are balanced and are coupled to the pace of locomotion. Thus, as illustrated by analysis of morphological markers in the cell shown in Fig. 1, the lamellipodium maintains its shape over time while the cytoskeleton flows rearward with respect to the cell, but does not move, or moves very slowly, with respect to the substrate. This pattern of lamellipodial dynamics has also been measured by computer-assisted microscopy (Royal et al. 1995). For example, difference pictures comparing the shape of the lamellipodium of crawling sperm at two-second intervals showed that the area of the zone of expansion at the leading margin was similar to the area lost due to retraction at the base of the lamellipodium. This study also showed that the average instantaneous velocity of sperm crawling on glass was 30 μm/min, but that the cells surged forward about every 0.35 min, increasing their velocity by an average of 47%. The cells maintained their lamellipodial shape during these surges, indicating that cytoskeletal assembly and disassembly remain balanced even when crawling speed changes (Royal et al. 1995).

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