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  • br IPA network analysis of important tumor oncogenic

    2020-08-12


    IPA network analysis of important tumor oncogenic or suppressor genes were then constructed as shown in Fig. 4A, which illustrated the proposed gene network regulated by SNRPA1. A total of 38 genes in-teracting with SNRPA1 were found, including 17 upregulated and 21 down-regulated genes. From this figure, we can also see that SNPRA1 
    According to this figure, knocking down of SNRPA1 may inhibit the cell proliferation through interaction with EGFR and HGF genes, thus affecting their regulated down-stream targets. Previous study showed that SNRPA1 can directly interact with epidermal growth factor re-ceptor (EGFR) [14]. EGFR is a member of the epidermal growth factor receptor family, located on the surface of the cell membrane and acti-vated by binding to ligands, including epidermal growth factor (EGF) and transforming growth factor-α (TGF-α). Studies have shown that epidermal growth factor, fibroblast growth factor (FGF), platelet-de-rived growth factor (PDGF) and other growth factors are highly ex-pressed in many tumor cells. These growth factors bind to their cognate receptors, causing two monomeric receptor molecules to form dimers on the membrane, activating their protein kinase activity, and autop-hosphorylation by receptor tyrosine kinases to provide additional sig-naling proteins in the cell. The binding site activates multiple signal transduction pathways within the cell. At present, receptor tyrosine kinase inhibitors are a hot spot in anti-tumor drug research. Activated receptor tyrosine kinase promotes tumor cell proliferation and metas-tasis by activating RAF/MEK/ERK, PI3K/AKT/mTOR and Src/JAK/
    STAT signaling pathways; the PI3K/AKT pathway is involved in this process [15,16]. Combined with the experimental data of this work, the deletion of SNRPA1 may also affect the downstream signaling pathway of EGFR by interacting with EGFR, thereby inhibiting the proliferation of tumor 480-49-9 and promoting tumor cell apoptosis.
    To validate the role of SNRPA1 in regulating those genes predicted from the above bioinformatic analysis, we analyzed the mRNA ex-pression of those genes by western blotting analysis assay. The assay results shown in Fig. 4B–C clearly showed that among 6 selected genes of interest there are 4 of them (EZH2, VEGFC, MKI67, CDK1) that were downregulated and 2 (NRP1, PIK3R1) were upregulated when com-paring their protein expression in the knocking down group with the control group. Those results were in consistence with the microarray analysis results. Taken them together, those findings suggest that SNRPA1 play important roles in the development and/or progress of CRC by regulating at least EZH2, VEGFC, MKI67, NRP1, PIK3R1 and CDK1 genes.
    EZH2 (Enhancer of zeste homolog 2) has been implicated in many studies that its expression is associated with poor outcome in CRC [17,18] or its depletion blocks the proliferation of CRC cells [19]. So, its downregulation benefits the elimination of CRC cells, consistent with those previous reports, we also found it was downregulated afte knocking down of SNRPA1 which led to the inhibition of cell pro-liferation in CRC cells. Earlier studies have shown that SNRPA1 can directly interact with SMN1/SMN2 (survival motor neuron) [20]. The SMN gene is located on chromosome 5q13, SMN1 is a telomere copy of the SMN gene, SMN2 is a centromere copy of the SMN gene, and the SMN1 and SMN2 genes encode the same protein. The SMN complex catalyzes the assembly of small nuclear ribosomal proteins (snRNPs), which affect the cleavage of intracellular mRNA precursors [21]. SMN can also affect the ERK signaling pathway, which affects neuronal cell division, proliferation and differentiation [22]. In addition, SMN1/ SMN2 can be associated with EZH2 [23]. Studies have also shown that EZH2 can affect the chromatin modification by participating in the 480-49-9 regulation of the pRB‐E2F pathway [24]; inhibition of EZH2 causes G2 arrest in medulloblastoma cells, leading to apoptosis [25]; high ex-pression of EZH2 gene It can promote the proliferation and invasion of prostate cancer, breast cancer and endometrial cancer tumor cells [25,26]. Therefore, SNRPA1 deletion may also affect EZH2 and its downstream related signaling pathways through interaction with SMN gene, thereby inhibiting tumor cell proliferation and promoting tumor cell apoptosis.
    VEGFC, one of angiogenic cytokines and a member of VEGF family which has been linked with metastatic spread of CRC [27]. Previous studies already showed that VEGF-C and VEGF-A were significantly raised in CRCs [27]. Though there was no association between VEGF-C or VEGF-D and lymphatic spread, High levels of VEGF-A and VEGF-C will increase their binding to the VEGF receptors, and produce the angiogenic switch required for tumor growth. On the contrary, de-creased expression of VEGF-C will aid in the control of CRCs. From this point of view, SNRPA1 downregulates the expression of VEGFC to in-hibit the tumor growth in CRCs.