Authors: Yukio Nishimura¹; Kiyoko Yoshioka²; Ora Bernard³; Masaru Himeno¹; Kazuyuki Itoh²

Source: European Journal of Cell Biology, Volume 83, Number 7, July 2004 , pp. 369-380(12)

Publisher: Urban & Fischer

Abstract:

LIM kinase (LIMK) plays a critical role in stimulus-induced remodeling of the actin cytoskeleton by linking signals from the Rho family GTPases to changes in cofilin activity. Recent studies have shown an important role for LIMK1 signaling in tumor cell invasion through regulating actin dynamics. In this study, we investigate the role of LIMK1 in intracellular vesicle trafficking of lysosomes/endosomes. We analyzed by confocal immunofluorescence microscopy the cellular distribution of lysosomal proteins and the endocytosis of an endocytic tracer, epidermal growth factor (EGF), in LIMK1-transfected cells. We found in these cells an abnormal dispersed translocation of lysosomes stained for LIMPII and cathepsin D throughout the cytoplasm. The small punctate structures that stained for these lysosomal proteins were redistributed to the periphery of the cell. Computational 3D-image analysis of confocal immunofluorescence micrographs further demonstrated that these vesicles did not colocalize with the transferrin receptor, an early endosomal marker. Furthermore, LIMPII-positive lysosomes did not colocalize with early endosomes labeled with endocytosed Texas red-transferrin. These results indicate that there is no mixing between dispersed lysosomes and early endosomes in the LIMK1-transfected cells. Moreover, LIMK1 overexpression resulted in a marked retardation in the receptor-mediated internalization of Texas red-labeled EGF in comparison with mock-transfected cells. At 30 min after internalization, most of the Texas red-EGF staining overlapped with LIMPII-positive late endosomes/lysosomes in mock-transfected cells, whereas in LIMK1 transfectants only a small fraction of internalized EGF colocalized with LIMPII-positive structures in the perinuclear region. Taken together, the findings presented in this paper suggest that LIMK1 has a role in regulating vesicle trafficking of lysosomes and endosomes in invasive tumor cells.

Document Type: Research article

DOI: http://dx.doi.org/10.1078/0171-9335-00382

Affiliations: 1: Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812 – 8582, Japan 2: Department of Biology, Osaka Medical Center for Cancer and Cardiovascular Diseases, 1-3-2 Nakamichi, Higashinari-ku, Osaka 537 – 8511, Japan 3: The Walter and Eliza Hall Institute of Medical Research, 1G Royal Parade, Parkville, Victoria 3050, Australia

Publication date: 2004-07-01

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