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Silencing of STRN4 suppresses the malignant characteristics of cancer cells.

Wong M, Hyodo T, Asano E, Funasaka K, Miyahara R, Hirooka Y, Goto H, Hamaguchi M, Senga T - Cancer Sci. (2014)

Bottom Line: We previously reported that STRN4 directly associated with protein kinases, such as MINK1, TNIK and MAP4K4, which are associated with tumor suppression or tumor progression.However, it remains unclear whether STRN4 is associated with tumor progression.Our results demonstrate a possible role of STRN4 in tumor progression.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology and Hepatology, Nagoya University Graduate School of Medicine, Nagoya, Japan.

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Related in: MedlinePlus

Silencing of STRN4 inhibits cell migration and invasion. (a) Monolayers of siRNA-transfected cells were scratched, and the distance between the leading edges was measured 24 h later. Representative images of the cells are shown. The graph shows the average distance of wound healing. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (b) Cells were transfected with siRNA, and 72 h later, cell migration was examined. The graph indicates the average number of migrated cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (c) siRNA-transfected cells were subjected to invasion assay. The graph indicates the average number of invaded cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (d) KP4 cells that constitutively expressed Flag tag (Flag-KP4) or STRN4 (STRN4-KP4) were established by retroviral infection. Cells were transfected with siRNA, and 72 h later, expression of STRN4 was examined by immunoblot. (e) Cells were transfected with siRNA, and 72 h later, the cells were subjected to migration and invasion assay. The graph indicates the average number of migrated or invaded cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05; NS, not significant).
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fig02: Silencing of STRN4 inhibits cell migration and invasion. (a) Monolayers of siRNA-transfected cells were scratched, and the distance between the leading edges was measured 24 h later. Representative images of the cells are shown. The graph shows the average distance of wound healing. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (b) Cells were transfected with siRNA, and 72 h later, cell migration was examined. The graph indicates the average number of migrated cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (c) siRNA-transfected cells were subjected to invasion assay. The graph indicates the average number of invaded cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (d) KP4 cells that constitutively expressed Flag tag (Flag-KP4) or STRN4 (STRN4-KP4) were established by retroviral infection. Cells were transfected with siRNA, and 72 h later, expression of STRN4 was examined by immunoblot. (e) Cells were transfected with siRNA, and 72 h later, the cells were subjected to migration and invasion assay. The graph indicates the average number of migrated or invaded cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05; NS, not significant).

Mentions: We next investigated whether STRN4 knockdown affects cell migration and invasion. To assess changes in cell migration, we first performed a wound healing assay. Confluent monolayers of KP4, HCT116, TE1 and SKOV3 cells transfected with siRNA were scratched, and the migration of the cells into the free space was observed 24 h later. The migration of STRN4-depleted cells was clearly delayed compared with that of the control siRNA-transfected cells (Fig. 2a). To further confirm this result, we used a modified Boyden chamber assay. Cells transfected with siRNA were placed on the upper surface of the filter and allowed to migrate to the bottom surface, which was coated with fibronectin. Cells that migrated to the bottom surface were quantified. The migration of STRN4-depleted cells was suppressed compared with that of the control siRNA-transfected cells (Figs 2b, S2a). To investigate the effect that STRN4 suppression had on cell invasion, we used a Matrigel-coated Boyden chamber. The invasion of cancer cells was significantly reduced by STRN4 knockdown (Figs 2c, S2b). To exclude the possibility that the suppression of cell migration and invasion was an off-target effect of siRNA, we performed a rescue experiment using KP4 cells. We first established KP4 cell lines that constitutively expressed a Flag tag (Flag-KP4) or STRN4 (STRN4-KP4) by retroviral infection. We then used siRNA that targeted the 3′ UTR of STRN4 (siSTRN4-3). Transfection of siSTRN4-3 efficiently suppressed the endogenous STRN4 in Flag-KP4 cells but not the exogenously expressed STRN4 in STRN4-KP4 cells (Fig. 2d). The expression of exogenous STRN4 clearly rescued the suppression of migration and the invasion induced by siSTRN4-3 transfection (Fig. 2e). These results indicate that STRN4 depletion suppresses cell migration and invasion.


Silencing of STRN4 suppresses the malignant characteristics of cancer cells.

Wong M, Hyodo T, Asano E, Funasaka K, Miyahara R, Hirooka Y, Goto H, Hamaguchi M, Senga T - Cancer Sci. (2014)

Silencing of STRN4 inhibits cell migration and invasion. (a) Monolayers of siRNA-transfected cells were scratched, and the distance between the leading edges was measured 24 h later. Representative images of the cells are shown. The graph shows the average distance of wound healing. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (b) Cells were transfected with siRNA, and 72 h later, cell migration was examined. The graph indicates the average number of migrated cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (c) siRNA-transfected cells were subjected to invasion assay. The graph indicates the average number of invaded cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (d) KP4 cells that constitutively expressed Flag tag (Flag-KP4) or STRN4 (STRN4-KP4) were established by retroviral infection. Cells were transfected with siRNA, and 72 h later, expression of STRN4 was examined by immunoblot. (e) Cells were transfected with siRNA, and 72 h later, the cells were subjected to migration and invasion assay. The graph indicates the average number of migrated or invaded cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05; NS, not significant).
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Related In: Results  -  Collection

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fig02: Silencing of STRN4 inhibits cell migration and invasion. (a) Monolayers of siRNA-transfected cells were scratched, and the distance between the leading edges was measured 24 h later. Representative images of the cells are shown. The graph shows the average distance of wound healing. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (b) Cells were transfected with siRNA, and 72 h later, cell migration was examined. The graph indicates the average number of migrated cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (c) siRNA-transfected cells were subjected to invasion assay. The graph indicates the average number of invaded cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05, compared with Ctrl siRNA-transfected cells). (d) KP4 cells that constitutively expressed Flag tag (Flag-KP4) or STRN4 (STRN4-KP4) were established by retroviral infection. Cells were transfected with siRNA, and 72 h later, expression of STRN4 was examined by immunoblot. (e) Cells were transfected with siRNA, and 72 h later, the cells were subjected to migration and invasion assay. The graph indicates the average number of migrated or invaded cells per field. Three independent experiments were performed, and the data are shown as the mean ± SD (*P < 0.05; NS, not significant).
Mentions: We next investigated whether STRN4 knockdown affects cell migration and invasion. To assess changes in cell migration, we first performed a wound healing assay. Confluent monolayers of KP4, HCT116, TE1 and SKOV3 cells transfected with siRNA were scratched, and the migration of the cells into the free space was observed 24 h later. The migration of STRN4-depleted cells was clearly delayed compared with that of the control siRNA-transfected cells (Fig. 2a). To further confirm this result, we used a modified Boyden chamber assay. Cells transfected with siRNA were placed on the upper surface of the filter and allowed to migrate to the bottom surface, which was coated with fibronectin. Cells that migrated to the bottom surface were quantified. The migration of STRN4-depleted cells was suppressed compared with that of the control siRNA-transfected cells (Figs 2b, S2a). To investigate the effect that STRN4 suppression had on cell invasion, we used a Matrigel-coated Boyden chamber. The invasion of cancer cells was significantly reduced by STRN4 knockdown (Figs 2c, S2b). To exclude the possibility that the suppression of cell migration and invasion was an off-target effect of siRNA, we performed a rescue experiment using KP4 cells. We first established KP4 cell lines that constitutively expressed a Flag tag (Flag-KP4) or STRN4 (STRN4-KP4) by retroviral infection. We then used siRNA that targeted the 3′ UTR of STRN4 (siSTRN4-3). Transfection of siSTRN4-3 efficiently suppressed the endogenous STRN4 in Flag-KP4 cells but not the exogenously expressed STRN4 in STRN4-KP4 cells (Fig. 2d). The expression of exogenous STRN4 clearly rescued the suppression of migration and the invasion induced by siSTRN4-3 transfection (Fig. 2e). These results indicate that STRN4 depletion suppresses cell migration and invasion.

Bottom Line: We previously reported that STRN4 directly associated with protein kinases, such as MINK1, TNIK and MAP4K4, which are associated with tumor suppression or tumor progression.However, it remains unclear whether STRN4 is associated with tumor progression.Our results demonstrate a possible role of STRN4 in tumor progression.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology and Hepatology, Nagoya University Graduate School of Medicine, Nagoya, Japan.

Show MeSH
Related in: MedlinePlus