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. 2022 Feb 28:2022:3963883.
doi: 10.1155/2022/3963883. eCollection 2022.

LIMK1 Interacts with STK25 to Regulate EMT and Promote the Proliferation and Metastasis of Colorectal Cancer

Xuecheng Sun  1 , Shaotang Li  2 , Han Lin  3
Affiliations

LIMK1 Interacts with STK25 to Regulate EMT and Promote the Proliferation and Metastasis of Colorectal Cancer

Xuecheng Sun et al. J Oncol. .

Abstract

Objective: To investigate the interaction between LIMK1 and STK25 and its expression in colon cancer and its effect on the malignant evolution of colon cancer.

Methods: Fluorescence quantitative PCR and immunohistochemistry were used to detect the expression of the LIMK1 gene in cancer and adjacent tissues of 20 clinical colon cancer samples. The overexpression plasmids of LIMK1 and STK25 were constructed. An shRNA specific to LIMK1 was synthesized and transfected into colon cancer cell lines. The expression levels of EMT-related markers in cell lines were detected by real-time PCR. The effects of LIMK1 and STK25 on the proliferation and invasion of colon cancer cell lines were detected by CCK-8 assay, Transwell, and clonogenesis.

Results: LIMK1 interacted with STK25 and was highly expressed in colon cancer. High expression of LIMK1 and STK25 is associated with poor prognosis in colon cancer patients. LIMK silencing inhibits proliferation, invasion, and EMT of colon cancer. Cotransfection of LIMK1 and STK25 promotes the malignant progression and EMT of colon cancer.

Conclusion: Protein interaction between LIMK1 and STK25 occurs. Overexpression of LIMK1 and STK25 plays a role in promoting cell proliferation and invasion in colon cancer tissues and cells. They also play a role in promoting the occurrence and development of colon cancer.

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Conflict of interest statement

The authors declare that they have no conflicts of interest.

Figures

Figure 1
Figure 1
LIMK1 is upregulated in colon cancer and is associated with poor prognosis of colon cancer. (a) Survival analysis of colon cancer patients with different expression levels of LIMK1. (b) Immunohistochemical test results of LIMK1 in normal colon tissue and colon cancer tissue. The immunohistochemical images were obtained from The Human Protein Atlas website. (c) Immunohistochemical statistical results of LIMK1 in normal colon tissue and colon cancer tissue. (d) UALCAN website analyzes the relationship between LIMK1 expression and colorectal cancer staging and metastasis. (e) The expression level of LIMK1 in normal intestinal epithelial cells and colorectal cancer cells. ∗p < 0.05; ∗∗p < 0.01.
Figure 2
Figure 2
LIMK1 regulates the expression of EMT markers. (a) The expression level of LIMK1 is positively correlated with the coexpression of Vimentin. (b) The expression level of LIMK1 is positively correlated with the coexpression of Twist1. (c) The expression level of LIMK1 is positively correlated with the coexpression of Snail. (d) The expression level of LIMK1 is negatively correlated with the coexpression of Slug. (e) The efficiency of knockdown of LIMK1 in SW620 and HCT-8 cells was verified. (f) After knocking down LIMK1 in SW620 and HCT-8 cells, the expression of E-cadherin was detected. (g) After knocking down LIMK1 in SW620 and HCT-8 cells, the expression of Vimentin was detected. (h) After knocking down LIMK1 in SW620 and HCT-8 cells, the expression of Twist1 was detected. (i) Detection of Snail1 expression level after knocking down LIMK1 in SW620 and HCT-8 cells. (j) After knocking down LIMK1 in SW620 and HCT-8 cells, Slug expression was detected. ∗∗p < 0.01.
Figure 3
Figure 3
Knockdown of LIMK1 inhibits colon cancer proliferation, invasion, and colonization. (a) Knockdown of LIMK1 inhibits cell viability detection of colon cancer cells SW620 and HCT-8. (b) Knockdown of LIMK1 inhibits the invasion of colon cancer cells SW620 and HCT-8 (magnification: 100x). (c) Knockdown of LIMK1 inhibits the clonogenic ability of colon cancer cells SW620 and HCT-8. ∗∗p < 0.01.
Figure 4
Figure 4
LIMK1 gene knockdown inhibits the growth of colon cancer tumors. (a) Representative pictures of tumors in the control group and LIMK1 knockdown group. (b) Detection of the expression of LIMK1 in the tumor tissue after SW620 cells transfected with sh-LIMK1 in tumor-bearing mice. (c) Detection of the tumor volume of SW620 cells transfected with sh-LIMK1. (d) Detection of the tumor weight of SW620 cells transfected with sh-LIMK1. (e) Detection of the expression of E-cadherin in tumor tissue after SW620 cells transfected with sh-LIMK1 in tumor-bearing mice. (f) Detection of the expression of Vimentin in tumor tissue after SW620 cells transfected with sh-LIMK1 in tumor-bearing mice. (g) Detection of the expression of Twist1 in the tumor tissue after SW620 cells transfected with sh-LIMK1 in tumor-bearing mice. (h) Detection of the expression of Snail1 in the tumor tissue after SW620 cells transfected with sh-LIMK1 in tumor-bearing mice. ∗∗p < 0.01, ∗∗∗p < 0.001.
Figure 5
Figure 5
Protein-protein interaction between LIMK1 and STK25. (a) LIMK1 interaction protein prediction. (b) Immunofluorescence colocalization of LIMK1 and STK25 in colon cancer cells (scale bar: 10 μm). (c) Co-IP experiment verifies the interaction between LIMK1 and STK25. (d) Co-expression of LIMK1 and STK25 in colon cancer tissue is positively correlated.
Figure 6
Figure 6
STK25 is upregulated in colon cancer and is associated with poor prognosis of colon cancer. (a) Survival analysis of colon cancer patients with different STK25 expression levels. (b) Immunohistochemical test results of STK25 in normal colon tissue and colon cancer tissue. The experimental results were obtained from The Human Protein Atlas website. (c) The statistical results of immunohistochemical detection of STK25 in normal colon tissue and colon cancer tissue. (d) RT-qPCR analysis of STK25 expression in CRC tissues. (e) Detection of transfection efficiency of overexpression of STK25 in SW620 and HCT-8 cells. (f) Overexpression of STK25 promotes the detection of cell viability of colon cancer cells SW620 and HCT-8 (magnification: 100x). (g) Overexpression of STK25 promotes the invasion of colon cancer cells SW620 and HCT-8. ∗∗p < 0.01.
Figure 7
Figure 7
Overexpression of STK25 increases the tumor-promoting effect of LIMK1. (a) Detection of LIMK1 expression in SW620 cells. (b) SW620 cell proliferation rate detection. (c) SW620 cells are tested by Transwell (magnification: 100x). (d) Detection of E-cadherin expression in SW620 cells. (e) Detection of Vimentin expression in SW620 cells. (f) Detection of Twist1 expression in SW620 cells. (g) Detection of Snail1 expression in SW620 cells.
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