lncRNA GK-IT1 influences the carcinogenesis of non-small cell lung cancer cells through regulating aldolase A

doi:10.3969/j.issn.1674-8115.2022.05.006

Abstract

Abstract: Objective

·To investigate the effects of long non-coding RNA (lncRNA) GK-IT1 on the carcinogenesis of non-small-cell lung cancer (NSCLC) A549 and H358 cells by regulating aldolase A (ALDOA).

Methods

·The expression of GK-IT1 in 42 pairs of NSCLC tissues and adjacent tissues, and NSCLC cell lines were tested by quantitative real-time PCR (qRT-PCR). The subcellular localization of GK-IT1 in A549 and H358 cells was tested by fluorescence in situ hybridization (FISH). RNA immunoprecipitation (RIP) was used to assess the interaction of GK-IT1 and ALDOA. A549 and H358 cells were transfected with GK-IT1 small interfering RNA sequences (Si-GK-IT1 #1 and Si-GK-IT1 #2) and negative control (Si-NC), and then the two cell lines were also cotransfected with Si-GK-IT1 #2 and ALDOA overexpression plasmid. The Si-NC, Si-GK-IT1 #1, Si-GK-IT1 #2 groups were set. CCK-8 and EdU assay were used to detect cell proliferation. Transwell invasion and scratch assay were used to observe cell invasion and migration. Western blotting was carried out to verify the expression of vimentin, E-cadherin, N-cadherin and ALDOA in each experimental group.

Results

·Compared with the normal tissues, the relative expression of GK-IT1 was significantly higher in NSCLC tissues. Compared with BEAS-2B cells, the expression of GK-IT1 in H358 and A549 cells was significantly higher (P<0.05). FISH assay indicated that GK-IT1 was mainly located in the cytoplasm. RIP and bioinformatics analysis suggested that GK-IT1 might interact with ALDOA. Compared with the Si-NC group, the results of CCK-8 assay showed that the cell proliferation of the Si-GK-IT1 #1 and Si-GK-IT1 #2 group was significantly inhibited (P<0.05). The results of EdU assay showed that the ratio of EdU positive cells in the Si-GK-IT1 #1 and Si-GK-IT1 #2 group was significantly lower than that in the Si-NC group (P<0.05). Compared with the Si-NC group, the results of Transwell invasion assay indicated that the invasion of the Si-GK-IT1 #1 and Si-GK-IT1 #2 group was significantly inhibited (all P=0.000). The results of cell scratch assay showed that the healed wound ratio in the Si-GK-IT1 #1 and Si-GK-IT1 #2 group was remarkably lower than that in Si-NC group (all P=0.000). GK-IT1 knockdown significantly decreased the expression of ALDOA and vimentin, while the expression of E-cadherin was increased. Overexpression of ALDOA reversed the effects of GK-IT1 silencing on cell proliferation, invasion, migration and protein expression of NSCLC cells (all P<0.05).

Conclusion

·GK-IT1 could promote the proliferation, invasion and migration of NSCLC cells via regulating ALDOA.

Key words: long non-coding RNA (lncRNA), GK-IT1, aldolase A (ALDOA), non-small cell lung cancer (NSCLC), cell invasion, cell migration

CLC Number:

R734.2

LIU Ziyang, WANG Xiaowen, CHEN Li. lncRNA GK-IT1 influences the carcinogenesis of non-small cell lung cancer cells through regulating aldolase A[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2022, 42(5): 591-601.

Figures/Tables 9

Fig 1 Expression of GK-IT1 and ALDOA in NSCLC tissues

Fig 2 GK-IT1 expression and knockdown effect in NSCLC cell lines

Fig 3 E?ects of GK-IT1 knockdown on the proliferation of NSCLC cells

Fig 4 Effects of GK-IT1 knockdown on the invasion and migration of NSCLC cells

Fig 5 Subcellular localization of GK-IT1

Fig 6 Effects of GK-IT1 knockdown on the expression of ALDOA and EMT-related proteins in NSCLC cells

Fig 7 Interaction between GK-IT1 and ALDOA

Fig 8 Effects of ALDOA overexpression on the proliferation, invasion and migration of GK-IT1-knockdown NSCLC cells

Fig 9 Effects of ALDOA overexpression on the expression of ALDOA and EMT-related proteins in GK-IT1-knockdown NSCLC cells

TrendMD

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