Effect of overexpression of β- and γ-actin isoforms on actin cytoskeleton organization and migration of human colon cancer cells

Histochemistry and Cell Biology, Sep 2014

Actins are eukaryotic proteins, which are involved in diverse cellular functions including muscle contraction, cell motility, adhesion and maintenance of cell shape. Cytoplasmic actin isoforms β and γ are ubiquitously expressed and essential for cell functioning. However, their unique contributions are not very well understood. The aim of this study was to determine the effect of β- and γ-actin overexpression on the migration capacity and actin cytoskeleton organization of human colon adenocarcinoma BE cells. In cells overexpressing β- or γ-actin, distinct cytoskeletal actin rearrangements were observed under the laser scanning confocal microscope. Overexpressed actins localized at the submembranous region of the cell body, especially near to the leading edge and on the tips of pseudopodia. The cells transfected with plasmids containing cDNA for β- or γ-actin were characterized by increased migration and invasion capacities. However, the migration velocity was statistically significantly higher only in the case of γ-actin overexpressing cells. In conclusion, the increased level of β- or γ-actin leads to actin cytoskeletal remodeling followed by an increase in migration and invasion capacities of human colon BE cells. These data suggest that expression of both actin isoforms has an impact on cancer cell motility, with the subtle predominance of γ-actin, and may influence invasiveness of human colon cancer.

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Effect of overexpression of β- and γ-actin isoforms on actin cytoskeleton organization and migration of human colon cancer cells

Histochem Cell Biol Effect of overexpression of β‑ and γ‑actin isoforms on actin cytoskeleton organization and migration of human colon cancer cells Aleksandra Simiczyjew 0 Antonina Joanna Mazur 0 Agnieszka Popow‑Woz´niak 0 Maria Malicka‑Błaszkiewicz 0 Dorota Nowak 0 0 a. Simiczyjew · a. J. Mazur · a. Popow-Woz ́niak · M. Malicka-Błaszkiewicz · D. nowak ( 1 ) Department of Cell Pathology, Faculty of Biotechnology, University of Wrocław , Joliot-Curie 14a, 50-383 Wrocław , Poland actins are eukaryotic proteins, which are involved in diverse cellular functions including muscle contraction, cell motility, adhesion and maintenance of cell shape. Cytoplasmic actin isoforms β and γ are ubiquitously expressed and essential for cell functioning. However, their unique contributions are not very well understood. The aim of this study was to determine the effect of β- and γ-actin overexpression on the migration capacity and actin cytoskeleton organization of human colon adenocarcinoma Be cells. In cells overexpressing β- or γ-actin, distinct cytoskeletal actin rearrangements were observed under the laser scanning confocal microscope. Overexpressed actins localized at the submembranous region of the cell body, especially near to the leading edge and on the tips of pseudopodia. The cells transfected with plasmids containing cDna for β - or γ-actin were characterized by increased migration and invasion capacities. However, the migration velocity was statistically significantly higher only in the case of γ-actin overexpressing cells. In conclusion, the increased level of β- or γ-actin leads to actin cytoskeletal remodeling followed by an increase in migration and invasion capacities of human colon Be cells. These data suggest that expression of both actin isoforms has an impact on cancer cell motility, with the subtle predominance of γ-actin, and may influence invasiveness of human colon cancer. actin isoforms; Cancer cells invasion; Migration Introduction The actin family consists of highly conservative proteins, abundant in all eukaryotic cells. actin plays different roles in cell functioning including cell motility, contractile ring formation during cytokinesis, maintenance of cell shape, signal transduction, cell adhesion, transcription and muscle contraction (Perrin and ervasti 2010) . In vertebrates, six actin isoforms are expressed. They have been classified by both isoelectric point and primary tissue or cellular localization and comprise β- and γ-cytoplasmic, α-skeletal, α-cardiac, and α- and γ-smooth muscle isoactins (Vandekerckhove and Weber 1978). Muscle actins are tissue specific, whereas β- and γ-cytoplasmic actins are ubiquitously present in almost all cell types and are essential for cell survival (Harborth et al. 2001) . actin isoforms are products of separate genes, although there is a high homology among their nucleotide sequences resulting in very similar protein primary structure. actin isoforms mirror tissue but not species specificity (Sheterline et al. 1995; Khaitlina 2001) . The differences between actin isoforms occur especially in the most variable n-terminal region of the actin molecule. The two cytoplasmic actin isoforms—β and γ—differ only by four amino acids located at positions 2, 3, 4 and 10. β-actin contains asp-asp-asp at the n-terminus and Val at position 10 of the polypeptide chain, whereas γ-actin possesses n-terminal tripeptide glu-glu-glu and Ile at position 10 (Vandekerckhove and Weber 1978). The proportion of cytoplasmic actins varies and depends on the cell type (Vandekerckhove and Weber 1978; Sheterline et al. 1995; nowak and Malicka-Błaszkiewicz 1999) . In most cells, regardless of their different origins, β- and γ-actin isoforms are found in a ratio of approximately 2:1 (Khaitlina 2007; Bergeron et al. 2010) . However, the β/γ isoform ratio in different rat tissues is: 1:1 in testicles, 2.5:1 in liver and 6:1 in aorta (Vandekerckhove and Weber 1978; nowak and MalickaBłaszkiewicz 1999) . Mammalian erythrocytes contain only β-actin (Pinder and gratzer 1983) , whereas γ-actin predominates in chicken auditory hair cells (Höfer et al. 1997 ). The changes in actin isoform levels in cells are often connected with pathological processes. The changed levels of β- and γ-actins were shown to accompany many tumor types such as chemically induced melanoma, hepatoma, lymphoma and breast cancer (nowak and MalickaBłaszkiewicz 1999; Toh et al. 1977; gabbiani et al. 1984; Brittingham et al. 1997) . Cytoplasmic β-actin seems to be overexpressed in many tumors, especially in actively moving cancer cells. This isoform level was also remarkably increased in the case of selected, highly invasive variants of colon cancer cells (nowak et al. 2005) , MDCK cells transformed by Moloney sarcoma virus (MSV) (le et al. 1998) or melanoma T1C1 cells (goidin et al. 2001) . However, the published data are often controversial. For example, lymphocytes synthesize β-actin in remarkabl (...truncated)


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Aleksandra Simiczyjew, Antonina Joanna Mazur, Agnieszka Popow-Woźniak, Maria Malicka-Błaszkiewicz, Dorota Nowak. Effect of overexpression of β- and γ-actin isoforms on actin cytoskeleton organization and migration of human colon cancer cells, Histochemistry and Cell Biology, 2014, pp. 307-322, Volume 142, Issue 3, DOI: 10.1007/s00418-014-1199-9