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KMID : 0613820170270101199
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Journal of Life Science
2017 Volume.27 No. 10 p.1199 ~ p.1206
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Lymphotoxin β Receptor Stimulation Is Linked to MLCK Activity and Suppresses Stress Fiber Formation in Agonistic Anti-LTβR Antibody-stimulated Fibroblastic Reticular Cells
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Kim Min-Hwan
Lee Jong-Hwan
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Abstract
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The lymphotoxin β receptor (LTβR), a member of the tumor necrosis factor receptor family, plays an important role in lymphoid tissue’s architecture and organogenesis. We found that LTβR stimulation induced changes in stress fibers (SFs) in fibroblastic reticular cells (FRCs). MLCK and ROCK play critical roles in the regulation of SF formation in cells. The present study was performed to investigate the antifibrotic effects on SF regulation of LTβR signaling, with a focus on MLCK inhibition. The effect of LTβR on the SF change was analyzed using immunoblot and fluorescence assays and agonistic anti-LTβR antibody-treated FRCs. In addition, we checked the level of Rho-guanosine diphosphate (GDP)/guanosine triphosphate (GTP) exchange activity with FRC lysate. Phospho-ezrin proteins acting as membrane-cytoskeleton linkers completely de-phosphorylated in agonistic anti-LTβR antibody-treated FRCs. The actin bundles rearranged into SFs, where phospho-myosin light chain (p-MLC) co-localized in FRCs. ML7-treated FRCs completely blocked SFs and showed retraction and shrinkage processes comparable to those observed in agonistic anti-LTβR antibody-treated cells. Inhibition of ROCK activity induced changes in the actin cytoskeleton organization; however, some SFs remained in the cells, while they were completely disrupted by MLCK inhibition with ML7. We showed that the phosphorylation of MLC was completely abolished with LTβR stimulation in FRCs. When LTβR was stimulated with the agonistic anti-LTβR antibody, the Rho-GDP/GTP exchange activity was reduced, however, the activity was not completely abolished. Collectively, the results illustrated that MLCK was potently responsible for the SF regulation triggered via LTβR signaling in FRCs.
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KEYWORD
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FRC (fibroblastic reticular cell), MLCK, LTβR (lymphotoxin β receptor), ROCK, SF (stress fiber)
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