Part:BBa_K2889000:Experience

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Applications of BBa_K2889000

1.cell migration

1.1 Cell migration is a central process in the development and maintenance of tumor. We cloned the full length of IL7-AS-S2 into PCDNA3.1 and transfected the plasmid to 786-O cells.Through in vitro scratch wound healing assay, overexpression of IL7-AS-S2 promoted cell migration of 786-O cells.These results suggested that IL7-AS-S2 contain key structural domains(Fig. 1 and 2).

Overexpression of IL7-AS-S2 promotes 786-O cells migration. 786-O cells were transfected with pcDNA3.1-IL7-AS-S2 for 24 h. The cells growth areas were calculated with Image J at the time points of 0, 6, 12 and 24 h. Bar=100 μm. Data represents means ± SE from 3 independent experiments. *p<0.05 versus non-transfected cells.

1.2 In order to investigate whether cell migration induced by IL7-AS-S2 depends on the amount of IL7-AS-S2 transfection. We transfected the different concentration of IL7-AS-S2 into 786-0 cells.(Fig. 3 and 4)

2.IL7-AS-S2 contains a complicated secondary structure

In silico prediction of lncRNA secondary structure is another useful method to assign putative functions to non-coding transcripts, based upon the widely held assumption that highly folded structures impart functionality through binding interactions with proteins/nucleotides. Characterization of IL7-AS-S2 using RNAfold minimum free energy estimations predicted a highly folded secondary structure with several hairpin loops, which imply interact with protein (Fig. 5).

The optimal secondary structure with a minimum free energy. The secondary structures of IL7-AS-S2 were predicted by the RNAfold webserver (http://rna.tbi.univie.ac.at/cgi-bin/RNAfold.cgi). The structure is colored according to base-pairing probabilities. For unpaired regions, the color denotes the probability of being unpaired.

3.Bioinformatic prediction of IL7-AS-S2 function

LncRNAs perform the function through interacting with proteins. Considering IL7-AS-S2 can influence the cell migration, we want to predict proteins which can interact with IL7-AS-S2. Using the catRAPID omics algorithm (http://service.tartaglialab.com/page/catrapid_omics_group), we determined whether IL7-AS-S2 interacts with proteins. Results showed that the related RNA-binding proteins were involved in RNA-splicing/processing (CWC15, RU2A), and protein synthesis (EIF3G, SBDS) (table 1). Based on the proteins predicted, we estimated that IL7-AS-S2 may regulate RNA splicing or protein synthesis of some tumor related genes though interacting with several RNA-splicing or protein synthesis associated proteins.

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