Abstract
Oxidative stress can lead to T cell hyporesponsiveness. A reducing micromilieu (e.g. provided by dendritic cells) can rescue T cells from such oxidant-induced dysfunction. However, the reducing effects on proteins leading to restored T cell activation remained unknown. One key molecule of T cell activation is the actin-remodeling protein cofilin, which is dephosphorylated on serine 3 upon T cell costimulation and has an essential role in formation of mature immune synapses between T cells and antigen-presenting cells. Cofilin is spatiotemporally regulated; at the plasma membrane, it can be inhibited by phosphatidylinositol 4,5-bisphosphate (PIP2). Here, we show by NMR spectroscopy that a reducing milieu led to structural changes in the cofilin molecule predominantly located on the protein surface. They overlapped with the PIP2- but not actin-binding sites. Accordingly, reduction of cofilin had no effect on F-actin binding and depolymerization and did not influence the cofilin phosphorylation state. However, it did prevent inhibition of cofilin activity through PIP2. Therefore, a reducing milieu may generate an additional pool of active cofilin at the plasma membrane. Consistently, in-flow microscopy revealed increased actin dynamics in the immune synapse of untransformed human T cells under reducing conditions. Altogether, we introduce a novel mechanism of redox regulation: reduction of the actin-remodeling protein cofilin renders it insensitive to PIP2 inhibition, resulting in enhanced actin dynamics.
Highlights
Cofilin is a key molecule for actin dynamics whose activity can be locally inhibited by PIP2
Reducing Treatment Rescues Cofilin from PIP2 Inhibition— After ensuring that the F-actin-depolymerizing function of cofilin was not influenced by reduction, we considered that the observed structural changes might alter cofilin interaction with other binding partners
A Reducing Micromilieu Lowers the F-actin Enrichment in the Immune Synapse—Having established the fact that PIP2 inhibition of cofilin is prevented by reduction of cofilin, we investigated what effect a reducing milieu might have on the actin cytoskeleton during T cell activation
Summary
Cofilin is a key molecule for actin dynamics whose activity can be locally inhibited by PIP2. We show by NMR spectroscopy that a reducing milieu led to structural changes in the cofilin molecule predominantly located on the protein surface They overlapped with the PIP2- but not actin-binding sites. Reduction of cofilin had no effect on F-actin binding and depolymerization and did not influence the cofilin phosphorylation state It did prevent inhibition of cofilin activity through PIP2. This is achieved with the help of dendritic cells, which can provide a reducing intracellular milieu for nearby T cells by up-regulating thiols in the T cells. We show that the protein structure of cofilin is influenced by a reducing milieu, which prevents cofilin inactivation by PIP2, quantitatively enhancing its actin-remodeling activity in the cell
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