The localization and activity of sphingosine kinase 1 are coordinately-regulated with actin cytoskeletal dynamics in macrophages

Authors:
Kusner DJ, Thompson CR, Melrose NA, Pitson SM, Obeid LM, Iyer SS
In:
Source: J Biol Chem
Publication Date: (2007)
Issue: 282(32): 23147-62
Research Area:
Immunotherapy / Hematology
Cells used in publication:
THP-1
Species: human
Tissue Origin: blood
RAW 264.7
Species: mouse
Tissue Origin: blood
Platform:
Nucleofectorâ„¢ I/II/2b
Abstract
The physiologic and pathologic functions of sphingosine kinase (SK)1 require translocation to specific membrane compartments. We tested the hypothesis that interactions with actin filaments regulate the localization of SK1 to membrane surfaces; including the plasma membrane (PM) and phagosome. Macrophage activation is accompanied by a marked increase in association of SK1 with actin filaments. Catalytically-inactive (CI)- and phosphorylation-defective (PD)-SK1 mutants exhibited reductions in PM translocation, colocalization with cortical actin filaments, membrane ruffling, and lamellipodia formation, compared to wild-type (WT)-SK1. However, translocation of CI- and PD-SK1 to phagosomes were equivalent to WT-SK1. SK1 exhibited constitutive- and stimulus-enhanced association with actin filaments and F-actin-enriched membrane fractions in both intact macrophages and a novel in vitro assay. In contrast, SK1 bound G-actin only understimulated conditions. Actin inhibitors disrupted SK1 localization and modulated its activity. Conversely, reduction of SK1 levels or activity via RNA interference or specific chemical inhibition resulted in dysregulation of actin filaments. Thus, the localization and activity of SK1 are coordinately regulated with actin dynamics during macrophage activation.