Dynamic Localization of the Actin-Bundling Protein Cortexillin I during Cell Migration

Abstract

Cortexillins are actin-bundling proteins that play a critical role in regulating cell morphology and actin cytoskeleton reorganization in Dictyostelium. Here, we investigated dynamic subcellular localization of cortexillin I in chemotaxing Dictyostelium cells. Most of the cortexillin I was enriched on the lateral sides of moving cells. Upon chemoattractant stimulation, cortexillin I was rapidly released from the cortex followed by a transient translocation to the cell cortex with a peak at ~5 s and a subsequent decrease to basal levels, indicating that localization of cor-texillin I at the cortex in chemotaxing cells is controlled by two more signaling components, one for the initial delocalization from the cortex and another for the translocation to the cortex ~5 s after chemoattractant stimulation. Loss of cortexillins leads to reduced cell polarity and an in-creased number of lateral pseudopodia during chemotaxis, suggesting that cortexillins play an inhibitory role in producing pseudopodia along the lateral sides of the cell. Cells lacking cortexillins displayed extended chemoattrac-tantmediated Arp2/3 complex translocation kinetics to the cortex. Our present study provides a new insight into the function of cortexillins during reorganization of the actin cytoskeleton and cell migration.

Actin cross-linking proteins cortexillin I and II are required for cAMP signaling duringDictyosteliumchemotaxis and development

“…The different results from the two laboratories might be due to the different assays used and/or differences in the parental cell strains. Cha and Jeon (2011) also observed that ctxA − /B − cells are flatter and rounder and chemotax more slowly than WT cells and that aggregation is inhibited and development does not proceed to completion in ctxA − /B − cells.…”

The I/LWEQ Domain in RapGAP3 Required for Posterior Localization in Migrating Cells

“…It is known that cells form actin foci at their bottoms, which are exhibited by GFP-coronin, a marker protein for newly formed F-actin ( Fig. 2B ) ( Cha and Jeon, 2011 ; Jeon et al, 2007b ). GFPLD was found at foci at the bottom of cells, similar to the actin foci shown by GFP-coronin.…”

“…The myosin II , gCA/sGC , and gbp a/b null strains were obtained from the DictyBase Stock Center. The expression plasmids for GFP-coronin and RFP-coronin were described previously ( Cha and Jeon, 2011 ; Jeon et al, 2007b ). For expression of GFP-RapGAP3, the full coding sequence of rapGAP3 cDNA was generated by RT-PCR, cloned into the Bgl II- Xho I site of pBluescript KS (−), sequenced, and subcloned into the expression vector pEXP-4(+) containing a GFP fragment ( Jeon et al, 2009 ).…”

“…The subcellular localization of proteins in response to chemoattractant stimulation was examined as described previously ( Cha and Jeon, 2011 ; Jeon et al, 2007b ). Vegetative cells were washed twice with Na/K phosphate buffer, resuspended at a density of 5 × 10 6 cells/ml in Na/K phosphate buffer, and pulsed with 30 nM cAMP at 6-min intervals for 5 h. The pulsed cells were placed on glass-bottomed microwell plates.…”

Cytoskeletal Regulation of Inflammation and Its Impact on Skin Blistering Disease Epidermolysis Bullosa Acquisita

“…The actin cytoskeleton is involved in an array of vital cellular functions and has a pronounced influence on many aspects of skin biology [ 14 , 15 , 16 ]. Despite the structural role of the cytoskeleton, it is highly dynamic and can be rapidly modified to facilitate changes in cell structure during vesicle-organelle transport, cell-cell interactions, cell-extracellular matrix interactions and cell adhesion and motility [ 17 , 18 , 19 ]. The cytoskeleton comprises a network of filamentous (F)-actin, microtubules, intermediate filaments and stress fibres, all working together to mediate the continual remodelling, assembly and severing needed to generate the mechanical force required for cellular contraction, adhesion and motility [ 20 ].…”