miR-205 promotes proliferation and invasion of laryngeal squamous cell carcinoma by suppressing CDK2AP1 expression

Abstract

Background: The aberrant expression of microRNAs (miRNAs) has been found in various types of cancer. miR-205 was reported to be upregulated in laryngeal squamous cell carcinoma (LSCC) tissues, however, the mechanisms by which miR-205 functions as a regulator of LSCC are largely unknown.

Results: In this study, Real-time qPCR and Western blot assay showed that expression of miR-205 was upregulated and expression of cyclin-dependent kinase 2-associated protein 1 (CDK2AP1) was downregulated in LSCC tissues. The expression levels of miR-205 were negatively related to those of CDK2AP1 in LSCC tissues and cell lines. Moreover, we found that miR-205 was the upstream regulator of CDK2AP1 and could suppress the CDK2AP1 expression in LSCC cells. 3-(4,5-dimethylthiazal-2-yl)-2,5-diphenyl-tetrazolium bromide assays and transwell invasion assay were performed to test the proliferation and invasion of LSCC cells. Gelatin zymography was used to detect the activity of MMP2 and MMP9. CDK2AP1, c-Myc and CyclinD1 expression in cells was assessed with Western blotting. We found that miR-205 was the upstream regulator of CDK2AP1 and could suppress the expression of CDK2AP1 in LSCC cells. In addition, miR-205 significantly induced cell proliferation and invasion by suppressing CDK2AP1 expression. Consistent with miR-205 inhibitors, overexpressed CDK2AP1 suppressed the activity of MMP2 and MMP9 and c-Myc and CyclinD1 expression in LSCC cells.

Conclusion: These findings help us to better elucidate the molecular mechanisms of LSCC progression and provide a new theoretical basis to further investigate miR-205 as a potential biomarker and a promising approach for LSCC treatment.

Fig. 1

The expression of miR-205 and CDK2AP1 in NPC tissue. a miR-205 expression levels was examined by qRT-PCR in 10 cases of clinical LSCC tissues and paired non-tumorous tissues. b CDK2AP1 expression levels was examined by qRT-PCR in 10 cases of clinical LSCC tissues and paired non-tumorous tissues. c CDK2AP1 expression levels was examined by western blot in 10 cases of clinical LSCC tissues and paired non-tumorous tissues. d Correlation of miR-205 levels with CDK2AP1 levels was examined by qRT-PCR in 10 cases of clinical LSCC tissues (Pearson’s correlation coefficient, r = −0.7604)

Fig. 2

miR-205 was the upstream regulator of CDK2AP1. a miR-205 expression in LSCC cell lines. qRT-PCR analysis revealed miR-205 expression in TU212, TU686, M2e and Hep-2 cell lines. Error bars represent ± S.E. and ^ap < 0.01 versus TU212 cells. ^bp < 0.01 versus M2e cells. ^cp < 0.01 versus TU686 cells. b qRT-PCR analysis revealed CDK2AP1 expression in miR-205 expression in TU212, TU686, M2e and Hep-2 cell lines. c Western blot analysis CDK2AP1 expression in miR-205 expression in TU212, TU686, M2e and Hep-2 cell lines.Error bars represent ± SD and ^ap < 0.01 versus TU212 cells. ^bp < 0.01 versus M2e cells. ^cp < 0.01 versus TU686 cells. d qRT-PCR analysis revealed the effect of miR-205 and CDK2AP1 on miR-205 expression. e Western blot analysis revealed the effect of miR-205 and CDK2AP1 on CDK2AP1 expression. Hep-2 cells were transfected with miR-205 (miR-205 group) or miR-205 negtive control (miR-205 NC group) or CDK2AP1 (CDK2AP1 group). Error bars represent ± SD and ^*p < 0.01 versus control group

Fig. 3

miR-205 promotes cell proliferation and invasion by suppressing CDK2AP1 expression in LSCC. a Transwell invasion assay revealed the effects of miR-205 and CDK2AP1 on LSCC invasion. b MTT assay revealed the effects of miR-205 and CDK2AP1 on LSCC proliferation. c Gelatin zymography assay revealed effects of miR-205 and CDK2AP1 on activity of MMP2 and MMP9 in LSCC

Fig. 4

miR-205 induces the expression of c-Myc and CyclinD1 by targeting CDK2AP1. Hep-2 cells were transfected with miR-205 mimics (miR-205 group) or miR-205 negative control (miR-205 NC group) or CDK2AP1 (CDK2AP1 group). Forty-eight hours later, proteins were harvested for Western blotting; GAPDH served as an internal control. Error bars represent ± SD *p < 0.01 versus control and NC