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Regulation mechanism of the lateral diffusion of band 3 in erythrocyte membranes by the membrane skeleton.

Tomishige M, Sako Y, Kusumi A - J. Cell Biol. (1998)

Bottom Line: When the membrane skeletal network was dragged and deformed/translated using optical tweezers, band 3 molecules that were undergoing hop diffusion were displaced toward the same direction as the skeleton.Mild trypsin treatment of ghosts, which cleaves off the cytoplasmic portion of band 3 without affecting spectrin, actin, and protein 4.1, increased the intercompartmental hop rate of band 3 by a factor of 6, whereas it did not change the corral size and the microscopic diffusion rate within a corral.These results indicate that the cytoplasmic portion of band 3 collides with the membrane skeleton, which causes temporal confinement of band 3 inside a mesh of the membrane skeleton.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Science, Graduate School of Science, Nagoya University, Chikusa-ku, Nagoya 464-8602, Japan.

ABSTRACT
Mechanisms that regulate the movement of a membrane spanning protein band 3 in erythrocyte ghosts were investigated at the level of a single or small groups of molecules using single particle tracking with an enhanced time resolution (0.22 ms). Two-thirds of band 3 undergo macroscopic diffusion: a band 3 molecule is temporarily corralled in a mesh of 110 nm in diameter, and hops to an adjacent mesh an average of every 350 ms. The rest (one-third) of band 3 exhibited oscillatory motion similar to that of spectrin, suggesting that these band 3 molecules are bound to spectrin. When the membrane skeletal network was dragged and deformed/translated using optical tweezers, band 3 molecules that were undergoing hop diffusion were displaced toward the same direction as the skeleton. Mild trypsin treatment of ghosts, which cleaves off the cytoplasmic portion of band 3 without affecting spectrin, actin, and protein 4.1, increased the intercompartmental hop rate of band 3 by a factor of 6, whereas it did not change the corral size and the microscopic diffusion rate within a corral. These results indicate that the cytoplasmic portion of band 3 collides with the membrane skeleton, which causes temporal confinement of band 3 inside a mesh of the membrane skeleton.

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(a and b) Representative trajectories of band 3 in the  erythrocyte membrane with a temporal resolution of 33 ms for 10 s.  Band 3 that is (a) or is not (b) undergoing macroscopic diffusion.  The color is changed simply to indicate the passage of time. (c)  Interconversion between diffusing and stationary phases occurred during an observation time over 10 min. The band 3 molecule was first undergoing apparent simple Brownian diffusion  (red line), and then suddenly became stationary (blue line). The  band 3 molecule stayed in that area for ∼1 min (orange line).  Then the molecule started to move again (green line) to resume  apparent simple Brownian diffusion (magenta line). Bar, 500 nm.
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Figure 2: (a and b) Representative trajectories of band 3 in the erythrocyte membrane with a temporal resolution of 33 ms for 10 s. Band 3 that is (a) or is not (b) undergoing macroscopic diffusion. The color is changed simply to indicate the passage of time. (c) Interconversion between diffusing and stationary phases occurred during an observation time over 10 min. The band 3 molecule was first undergoing apparent simple Brownian diffusion (red line), and then suddenly became stationary (blue line). The band 3 molecule stayed in that area for ∼1 min (orange line). Then the molecule started to move again (green line) to resume apparent simple Brownian diffusion (magenta line). Bar, 500 nm.

Mentions: Approximately two-thirds of the gold particles attached to band 3 were undergoing simple Brownian diffusion when they were observed with a time resolution of 33 ms (video rate) (Fig. 2 a). The remaining one-third of the particles exhibited oscillatory movements within small regions (∼100 nm-φ) during the observation period of 10 s (Fig. 2 b). These particles are likely to represent band 3 molecules that are bound to the membrane skeleton. Blackman et al. (1996) suggested a possibility that ∼25% of band 3 molecules are aggregated to form ∼5,000 mers (and each aggregate is rotationally mobile). These aggregates may also contribute as the band 3 fraction that do not undergo long-range diffusion in the present measurement.


Regulation mechanism of the lateral diffusion of band 3 in erythrocyte membranes by the membrane skeleton.

Tomishige M, Sako Y, Kusumi A - J. Cell Biol. (1998)

(a and b) Representative trajectories of band 3 in the  erythrocyte membrane with a temporal resolution of 33 ms for 10 s.  Band 3 that is (a) or is not (b) undergoing macroscopic diffusion.  The color is changed simply to indicate the passage of time. (c)  Interconversion between diffusing and stationary phases occurred during an observation time over 10 min. The band 3 molecule was first undergoing apparent simple Brownian diffusion  (red line), and then suddenly became stationary (blue line). The  band 3 molecule stayed in that area for ∼1 min (orange line).  Then the molecule started to move again (green line) to resume  apparent simple Brownian diffusion (magenta line). Bar, 500 nm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2132872&req=5

Figure 2: (a and b) Representative trajectories of band 3 in the erythrocyte membrane with a temporal resolution of 33 ms for 10 s. Band 3 that is (a) or is not (b) undergoing macroscopic diffusion. The color is changed simply to indicate the passage of time. (c) Interconversion between diffusing and stationary phases occurred during an observation time over 10 min. The band 3 molecule was first undergoing apparent simple Brownian diffusion (red line), and then suddenly became stationary (blue line). The band 3 molecule stayed in that area for ∼1 min (orange line). Then the molecule started to move again (green line) to resume apparent simple Brownian diffusion (magenta line). Bar, 500 nm.
Mentions: Approximately two-thirds of the gold particles attached to band 3 were undergoing simple Brownian diffusion when they were observed with a time resolution of 33 ms (video rate) (Fig. 2 a). The remaining one-third of the particles exhibited oscillatory movements within small regions (∼100 nm-φ) during the observation period of 10 s (Fig. 2 b). These particles are likely to represent band 3 molecules that are bound to the membrane skeleton. Blackman et al. (1996) suggested a possibility that ∼25% of band 3 molecules are aggregated to form ∼5,000 mers (and each aggregate is rotationally mobile). These aggregates may also contribute as the band 3 fraction that do not undergo long-range diffusion in the present measurement.

Bottom Line: When the membrane skeletal network was dragged and deformed/translated using optical tweezers, band 3 molecules that were undergoing hop diffusion were displaced toward the same direction as the skeleton.Mild trypsin treatment of ghosts, which cleaves off the cytoplasmic portion of band 3 without affecting spectrin, actin, and protein 4.1, increased the intercompartmental hop rate of band 3 by a factor of 6, whereas it did not change the corral size and the microscopic diffusion rate within a corral.These results indicate that the cytoplasmic portion of band 3 collides with the membrane skeleton, which causes temporal confinement of band 3 inside a mesh of the membrane skeleton.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Science, Graduate School of Science, Nagoya University, Chikusa-ku, Nagoya 464-8602, Japan.

ABSTRACT
Mechanisms that regulate the movement of a membrane spanning protein band 3 in erythrocyte ghosts were investigated at the level of a single or small groups of molecules using single particle tracking with an enhanced time resolution (0.22 ms). Two-thirds of band 3 undergo macroscopic diffusion: a band 3 molecule is temporarily corralled in a mesh of 110 nm in diameter, and hops to an adjacent mesh an average of every 350 ms. The rest (one-third) of band 3 exhibited oscillatory motion similar to that of spectrin, suggesting that these band 3 molecules are bound to spectrin. When the membrane skeletal network was dragged and deformed/translated using optical tweezers, band 3 molecules that were undergoing hop diffusion were displaced toward the same direction as the skeleton. Mild trypsin treatment of ghosts, which cleaves off the cytoplasmic portion of band 3 without affecting spectrin, actin, and protein 4.1, increased the intercompartmental hop rate of band 3 by a factor of 6, whereas it did not change the corral size and the microscopic diffusion rate within a corral. These results indicate that the cytoplasmic portion of band 3 collides with the membrane skeleton, which causes temporal confinement of band 3 inside a mesh of the membrane skeleton.

Show MeSH