Ubiquitination of spectrin regulates the erythrocyte spectrin-protein-4.1-actin ternary complex dissociation: implications for the sickle cell membrane skeleton.
It has been demonstrated by our laboratory that the irreversibly sickled cell (ISC) spectrin-4.1-actin complex dissociates slowly as compared to ternary complexes formed out of control (AA) and reversibly sickle cell (RSCs) core skeletons. These studies indicated that the molecular basis for the inability of irreversibly sickled cells (ISCs) to change shape is a skeleton that disassembles, and therefore reassembles, very slowly. The present study is based on the following observations: a) alpha-spectrin repeats 20 and 21 contain ubiquitination sites, and b) The spectrin repeats beta-1 and beta-2 are in direct contact with spectrin repeats alpha-20 and alpha-21 during spectrin heterodimer formation, and contain the protein 4.1 binding domain. We demonstrate here that alpha-spectrin ubiquitination at repeats 20 and 21 increases the dissociation of the spectrin-protein-4.1-actin ternary complex thereby regulating protein 4.1's ability to stimulate the spectrin-actin interaction. Performing in vitro ternary complex dissociation assays with AA control and sickle cell SS spectrin (isolated from high-density sickle cells), we further demonstrate that reduced ubiquitination of alpha-spectrin is, in part, responsible for the locked membrane skeleton in sickle cell disease.
Ghatpande
S S
SS
Department of Molecular and Cell Biology, University of Texas at Dallas, Richardson, TX, USA.
Goodman
S R
SR
D000199:Actins / Q000378:metabolism*; D000755:Anemia, Sickle Cell / Q000378:metabolism / Q000473:pathology*; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D004910:Erythrocyte Membrane / Q000378:metabolism*; D006801:Humans; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000378:metabolism*; D011485:Protein Binding; D013049:Spectrin / Q000737:chemistry / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism*
D000199:Actins; D003598:Cytoskeletal Proteins; D046911:Macromolecular Substances; D008565:Membrane Proteins; D025801:Ubiquitin; C028841:erythrocyte membrane band 4.1 protein; D013049:Spectrin