Protein 4.1R-dependent multiprotein complex: new insights into the structural organization of the red blood cell membrane.
Protein 4.1R (4.1R) is a multifunctional component of the red cell membrane. It forms a ternary complex with actin and spectrin, which defines the nodal junctions of the membrane-skeletal network, and its attachment to the transmembrane protein glycophorin C creates a bridge between the protein network and the membrane bilayer. We now show that deletion of 4.1R in mouse red cells leads to a large diminution of actin accompanied by extensive loss of cytoskeletal lattice structure, with formation of bare areas of membrane. Whereas band 3, the preponderant transmembrane constituent, and proteins known to be associated with it are present in normal or increased amounts, glycophorin C is missing and XK, Duffy, and Rh are much reduced in the 4.1R-deficient cells. The inference that these are associated with 4.1R was borne out by the results of in vitro pull-down assays. Furthermore, whereas Western blot analysis showed normal levels of band 3 and Kell, flow cytometric analysis using an antibody against the extracellular region of band 3 or Kell revealed reduction of these two proteins, suggesting a conformational change of band 3 and Kell epitopes. Taken together, we suggest that 4.1R organizes a macromolecular complex of skeletal and transmembrane proteins at the junctional node and that perturbation of this macromolecular complex not only is responsible for the well characterized membrane instability but may also remodel the red cell surface.
Red Cell Physiology Laboratory and Membrane Biochemistry Laboratory, New York Blood Center, New York, NY 10065, USA.
Marcela
M
Salomao
Xihui
X
Zhang
Yang
Y
Yang
Soohee
S
Lee
John H
JH
Hartwig
Joel Anne
JA
Chasis
Narla
N
Mohandas
Xiuli
X
An
D000818:Animals; D000918:Antibody Specificity; D001789:Blood Group Antigens / Q000378:metabolism; D001798:Blood Proteins / Q000737:chemistry / Q000172:deficiency / Q000378:metabolism*; D015153:Blotting, Western; D003598:Cytoskeletal Proteins / Q000378:metabolism; D004910:Erythrocyte Membrane / Q000737:chemistry* / Q000648:ultrastructure; D005434:Flow Cytometry; D017353:Gene Deletion; D015151:Immunoblotting; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008840:Microfilament Proteins; D008954:Models, Biological; D046912:Multiprotein Complexes / Q000378:metabolism*; D011485:Protein Binding
D001789:Blood Group Antigens; D001798:Blood Proteins; D003598:Cytoskeletal Proteins; C514956:Epb41 protein, mouse; D008565:Membrane Proteins; D008840:Microfilament Proteins; D046912:Multiprotein Complexes