Project Details
Projekt
Targeting spectrin redox switches to regulate the mechanoproperties of red blood cells.
Subject Area
Biochemistry
Term
from 2017 to 2021
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 386517575
The mechanical properties of red blood cells (RBCs) are fundamental for their physiology. To transport oxygen, RBCs need to survive the hemodynamic changes in the vascular tree, to dynamically contribute to the flow for decreasing vascular resistance, and to deform during the passage through narrower vessels. RBC mechanoproperties are conferred mainly by the structural characteristics of their cytoskeleton, which mainly consists of a spectrin scaffold connected to nodes of ankyrin and alpha-tubulin. Changes in redox state and treatment with thiol-targeting molecules decrease deformability of RBCs, and affect the structure/stability of the spectrin cytoskeleton. However, structural characteristics, molecular localization of redox switches and their functional link to RBC mechanoproperties, blood viscosity and tissue perfusion are unknown. We hypothesize that cysteine redox switches (CRS) on the spectrin cytoskeleton are targeted by redox signaling in RBCs and modulate RBC mechanical properties, thus, contributing to the modulation of blood viscosity, blood flow, and tissue perfusion. Hence, in this project we aim to (1.) to identify, experimentally validate and analyze the effects of putative CRS in erythrocytic spectrin; (2.) to analyze the effects of CRS modification on RBC mechanical properties; (3.) to investigate the functional significance of spectrin CRS in the physiology of RBCs in vivo. First, by integrative modeling, we will generate a structural model of spectrin at the atomistic level, which will allow us to identify and localize putative CRS in spectrin and to investigate how their chemical modification impacts elastic properties of spectrin. In conjunction, the impact of CSR modifications on structure/functional characteristics of spectrin in cells, including stability, mechanical properties, and protein/protein interactions with other cytoskeletal proteins will be experimentally analyzed. Second, RBC will be treated with physiological stimuli inducing intracellular production of reactive species (i.e., NO, O2-/H2O2, sulfide/persulfide/polysulfide) under normoxia and hypoxia, or changes in the cellular redox state. We will then analyze spectrin CSR modification, and their effects on RBC morphology and RBC mechanical properties, including RBC deformability, osmotic stability, and blood viscosity. Third, we will analyze the effects of (1.) pharmacological application of NO or sulfide, (2.) changes in redox state by phenylhydrazine, or (3.) genetic modulation of endogenous NO synthesis in RBC in mice on spectrin CRS modification, and their effects on RBC functional properties, gas exchange, and tissue perfusion. Structural and functional characterization of putative spectrin CRS will allow to shed light on the role of redox regulation on RBC mechanical properties, and may provide novel functional targets to modulate RBC function, blood flow, and tissue perfusion.
DFG Programme
Priority Programmes
