Lateral mobility of band 3 in the human erythrocyte membrane studied by fluorescence photobleaching recovery: Evidence for control by cytoskeletal interactions

Golan, David E.; Veatch, William R.

doi:10.1073/pnas.77.5.2537

Cited by 206 publications

(130 citation statements)

References 30 publications

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“…Whereas band 3 mobility in intact normal RBCs under physiologic conditions is relatively consistent from study to study, lateral mobility in ghost membranes is highly variable. Fractional mobilities ranging from 0 to 70% and diffusion coefficients from 0 to 80 x 10-" cm2/s have been reported ( 12,13,(87)(88)(89). We have observed that small changes in experimental conditions can dramatically affect protein lateral mobility in RBC ghost membranes (data not shown).…”

Section: Discussionmentioning

confidence: 52%

“…Intact cells are likely to be more physiologic than ghost membrane preparations in elucidating molecular alterations that mediate removal ofdense sickle and normal RBCs from the circulation. Band 3 lateral mobility has been studied in both RBC ghosts and intact cells ( 12,13,17,64,86). Whereas band 3 mobility in intact normal RBCs under physiologic conditions is relatively consistent from study to study, lateral mobility in ghost membranes is highly variable.…”

Section: Discussionmentioning

confidence: 99%

“…Glycophorin A, present in -5 x 105 copies per cell (5), is the major sialoglycoprotein. The membrane skeleton determines RBC stability, deformability, and shape (6-9) and appears to regulate lateral and rotational mobility of band 3 and glycophorin A in the plane ofthe membrane (10)(11)(12)(13)(14)(15)(16)(17)(18). The major skeletal protein, spectrin, is attached to the overlying bilayer through interactions involving: ankyrin, which links spectrin to a portion ( 10-15%) of band 3; protein 4.1, which links spectrin to glycophorin C and possibly to glycophorin A and band 3; and, possibly, inner leaflet aminophospholipids, including phosphatidylserine and phosphatidylethanolamine (2).…”

Section: Introductionmentioning

confidence: 99%

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Band 3 and glycophorin are progressively aggregated in density-fractionated sickle and normal red blood cells. Evidence from rotational and lateral mobility studies.

Corbett¹,

Golan²

1993

J. Clin. Invest.

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Band 3 aggregation in the plane of the red blood cell (RBC) membrane is postulated to be important in the pathophysiology of hemolysis of dense sickle and normal RBCs. We used the fluorescence photobleaching recovery and polarized fluorescence depletion techniques to measure the lateral and rotational mobility of band 3, glycophorins, and phospholipid analogues in membranes of density-separated intact RBCs from seven patients with sickle cell disease and eight normal controls. The fractions of laterally mobile band 3 and glycophorin decreased progressively as sickle RBC density increased. Normal RBCs also showed a progressive decrease in band 3 fractional mobility with increasing buoyant density. Rapidly rotating, slowly rotating, and rotationally immobile forms of band 3 were observed in both sickle and normal RBC membranes. The fraction of rapidly rotating band 3 progressively decreased and the fraction of rotationally immobile band 3 progressively increased with increasing sickle RBC density. Changes in the fraction of rotationally immobile band 3 were not reversible upon hypotonic swelling of dense sickle RBCs, and normal RBCs osmotically shrunken in sucrose buffers failed to manifest band 3 immobilization at median cell hemoglobin concentration values characteristic of dense sickle RBCs. We conclude that dense sickle and normal RBCs acquire irreversible membrane abnormalities that cause transmembrane protein immobilization and band 3 aggregation. Band 3 aggregates could serve as cell surface sites of autologous antibody binding and thereby lead to removal of dense sickle and normal (senescent)

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Section: Discussionmentioning

confidence: 52%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Band 3 and glycophorin are progressively aggregated in density-fractionated sickle and normal red blood cells. Evidence from rotational and lateral mobility studies.

Corbett¹,

Golan²

1993

J. Clin. Invest.

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“…Compared to N-4-nitrobenzo-2-oxa-1,3-diazole phosphatidylethanolamine, it diffuses at a rate near the predicted theoretical limit for a freely diffusing membrane protein (32)(33)(34) . Similar diffusion coefficients have been measured for rhodopsin in amphibian rod outer segment disks (35,36), and for plasma membrane proteins in membranes in which an underlying cytoskeleton may be modified or is separated from the membrane (37)(38)(39)(40)(41)(42) . It is not known whether a membrane-associated cytoskeleton is present in the posterior tail of guinea pig sperm .…”

Section: Rapid Communicationsmentioning

confidence: 48%

A localized surface protein of guinea pig sperm exhibits free diffusion in its domain.

Myles¹,

Primàkoff²,

Koppel³

1984

The Journal of Cell Biology

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Using the technique of fluorescence redistribution after photobleaching, we are studying the cellular mechanisms involved in localizing surface molecules to particular domains . A number of antigens localized to discrete surface regions have been identified with monoclonal antibodies on guinea pig sperm cells (Primakoff, P., and D . G . Myles, 1983, Dev . Biol ., 98 :417-428) . One of these monoclonal antibodies, PT-1, binds exclusively to the posterior tail region of the sperm cell surface . PT-1 recognizes an integral membrane protein that in complex with n-octyl-ß-D-glucopyranoside has a sedimentation coefficient of 6 .8S in sucrose density gradients . Fluorescence redistribution after photobleaching measurements reveal that within its surface domain the PT-1 antigen diffuses rapidly (D = 2 .5 x 10 -9 Cm 2/s) and completely (>90% recovery after bleaching) . These results rule out for this membrane protein all models that invoke immobilization as a mechanism for maintaining localization . We propose that the mechanism for localization of the PT-1 antigen may be a barrier to diffusion at the domain boundary .The restriction of cell surface molecules into specific domains has been well established for a wide variety of mammalian cell types . These include acetylcholine receptors at the synapse on muscle cells (1), low density lipoprotein and asialoglycoprotein receptors in coated pits of fibroblasts (2) and hepatocytes (3) and more extensive localized domains on the surface of sperm (4, 5), intestinal epithelial cells-(6, 7), and hepatocytes (8, 9). Although topographical localization of various surface receptors has been widely described, little is known concerning the mechanisms that maintain surface protein localizations . Immobilization in the membrane is a potential mechanism for maintaining a non-uniform distribution of surface molecules, and in the case of the acetylcholine receptor, it has been shown that clustered receptors are immobilized (10).In the current study we have addressed the question whether there are mechanisms other than immobilization to localize surface molecules . We have used the guinea pig sperm cell . Previously, the existence of five different antigen domains on the surface of the guinea pig sperm was established with monoclonal antibodies directed to surface antigens. These antigens may be restricted to the anterior head, posterior head, whole head, whole tail, or posterior tail cell surface (11). In some cases, patching of localized sperm surface molecules can be induced, suggesting that these molecules may be mobile in the plane of the membrane (12-15) . However, ability to patch may not be a reliable indicator of mobility of a receptor in the membrane (16-18) and gives no measure of the diffusion rate or percent mobile fraction. Therefore, we RAPID COMMUNICATIONS have directly measured these parameters using fluorescence redistribution after photobleaching of a localized sperm surface antigen that can be patched . This antigen, identified by the monoclonal antibody PT-...

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“…More importantly, it implies that the target of the photodamage involved in invagination formation is unable to diffuse away from the laser focus. Spectrin molecules are part of a membrane-associated protein network, and at room temperature 60% of band 3 proteins are immobile due to their interaction with ankyrin (Golan & Veatch, 1980). Therefore, both proteins are indeed mostly immobile membrane components.…”

Section: Discussionmentioning

confidence: 99%

Localized photodamage of the human erythrocyte membrane causes an invagination as a precursor of photohaemolysis

Wong

Ng‐Kamstra

Chen

et al. 2007

Journal of Microscopy

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SummaryFluorescence excitation can result in the formation of reactive oxygen species and free radicals damaging to live cells. In the case of erythrocytes, reaction of these reactive oxygen species with membrane components causes large-scale morphological changes followed by cell haemolysis. In an effort to understand the origin of these morphological changes, we have studied the consequences of localized photodamage on the erythrocyte membrane. For this, we irradiated a small area of the cell membrane using a focused laser beam in the presence of an external photosensitizer. We observed the rapid formation of an invagination (approximately 1 μm deep) at the laser focus, long before photohaemolysis occurred. We measured the rate of invagination formation and the rate of cell haemolysis, using a combination of fluorescence contrast imaging (to detect the membrane position) with fluorescence correlation spectroscopy (to measure photosensitizer concentration). We found that the kinetics of both processes depend in a similar manner on light energy flux, fluorophore concentration and the presence of oxygen scavenger. This leads us to the conclusion that the observed invagination is due to the photooxidation of membrane-associated proteins, representing a precursor of cellular photohaemolysis. We then discuss two different molecular mechanisms (conformational change of the protein band 3 and detachment of the spectrin cytoskeleton from the lipid membrane) that may explain how the photodamage of membraneassociated proteins can lead to a deformation of the lipid bilayer.

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Lateral mobility of band 3 in the human erythrocyte membrane studied by fluorescence photobleaching recovery: Evidence for control by cytoskeletal interactions

Cited by 206 publications

References 30 publications

Band 3 and glycophorin are progressively aggregated in density-fractionated sickle and normal red blood cells. Evidence from rotational and lateral mobility studies.

Band 3 and glycophorin are progressively aggregated in density-fractionated sickle and normal red blood cells. Evidence from rotational and lateral mobility studies.

A localized surface protein of guinea pig sperm exhibits free diffusion in its domain.

Localized photodamage of the human erythrocyte membrane causes an invagination as a precursor of photohaemolysis

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