Characterization of erythrocyte cytoskeletal regulation by monitoring membrane-spanning protein diffusion
Abstract
The structural strength and exceptional deformability of red blood cells (RBCs) can be attributed to an underlying cytoskeleton and protein tethers that anchor it to the membrane. Band 3, a membrane spanning anion transporter, serves as one of the major tethers between membrane and cytoskeleton and RBCs with a deficiency or defect in band 3, or the proteins that interact with it, have reduced deformability and lifespan, often resulting in a hemolytic anemia. Glycophorin A (GPA) is another membrane spanning protein suspected to interact with band 3 and possibly with the cytoskeleton under certain stimuli. While it is clear that band 3 and the cytoskeletal proteins are crucial for red blood cell survival and function, their organization and regulation is not well understood. Single particle tracking (SPT) provides a molecular-level view of the mobility of a protein and can reveal information about its interactions with the underlying cytoskeleton. In order to understand the formation and maintenance of these interactions, band 3 diffusion was measured using SPT on intact RBCs from transgenic mice with cytoskeletal defects. The diffusion of band 3 and GPA were also studied on intact, normal human RBCs and on RBCs with increased band 3 phosphorylation. Hyperphosphorylation of band 3 increased its diffusion ∼2 orders of magnitude revealing a novel mechanism of regulation of the band 3-cytoskeleton interaction. On the other hand, band 3 phosphorylation had no effect on the mobility of GPA suggesting they do not form a tight association though GPA may have other binding partners in the membrane.
Degree
Ph.D.
Advisors
Low, Purdue University.
Subject Area
Biochemistry|Biophysics
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