A genome‐wide siRNA screen for regulators of tumor suppressor p53 activity in human non‐small cell lung cancer cells identifies components of the RNA splicing machinery as targets for anticancer treatment

Abstract

Reinstating wild‐type tumor suppressor p53 activity could be a valuable option for the treatment of cancer. To contribute to development of new treatment options for non‐small cell lung cancer (NSCLC), we performed genome‐wide siRNA screens for determinants of p53 activity in NSCLC cells. We identified many genes not previously known to be involved in regulating p53 activity. Silencing p53 pathway inhibitor genes was associated with loss of cell viability. The largest functional gene cluster influencing p53 activity was mRNA splicing. Prominent p53 activation was observed upon silencing of specific spliceosome components, rather than by general inhibition of the spliceosome. Ten genes were validated as inhibitors of p53 activity in multiple NSCLC cell lines: genes encoding the Ras pathway activator SOS1, the zinc finger protein TSHZ3, the mitochondrial membrane protein COX16, and the spliceosome components SNRPD3, SF3A3, SF3B1, SF3B6, XAB2, CWC22, and HNRNPL. Silencing these genes generally increased p53 levels, with distinct effects on CDKN1A expression, induction of cell cycle arrest and cell death. Silencing spliceosome components was associated with alternative splicing of MDM4 mRNA, which could contribute to activation of p53. In addition, silencing splice factors was particularly effective in killing NSCLC cells, albeit in a p53‐independent manner. Interestingly, silencing SNRPD3 and SF3A3 exerted much stronger cytotoxicity to NSCLC cells than to lung fibroblasts, suggesting that these genes could represent useful therapeutic targets.

Survival-Associated Metabolic Genes in Human Papillomavirus-Positive Head and Neck Cancers

“…However, as both are part of mitochondrial respiratory complex IV, it is possible that the complex IV inhibitors ADDA 5 [30] and tetrathiomolybdate [31] could prove useful to phenocopy any metabolic effects associated with low gene expression, promoting enhanced survival in HPV+ HNSCC. It is also important to note that COX16 is an inhibitor of p53 activity, which means that non-mitochondrial functions of COX16 should not be discounted [32]. E6 has been shown to inhibit expression of COX16 [33], which may contribute to the lower levels observed in HPV+ versus HPV-HNSCC ( Figure 3D).…”

Label propagation defines signaling networks associated with recurrently mutated cancer genes

“…Contribution of p53 and its downstream signaling events, such as plasminogen activator signaling, to the maximal common subgraph of TSHZ3 and SETD2 networks offers further support to the hypothesis that TSHZ3's role in lung adenocarcinoma include regulation of p53 activity [44]. Overall, comparing and contrasting of dysregulated signaling networks of different mutations offered us a unique look into the intricate molecular changes tumors rely on to survive.…”

“…revealed a relationship between SETD2 and p53 [45], in which SETD2 can contribute to the regulation of p53 signaling by enhancing its transcriptional activities. Additionally, a recent study discovered a connection between TSHZ3 and p53, where TSHZ3 is identified as an inhibitor of p53 activity in lung cancer cell lines [44]. Another major contributor to this maximal common subgraph is "Urokinasetype plasminogen activator (uPA) and uPAR-mediated signaling", highlighted in purple in Figure 4c.…”

“…Following these examples, we turned our attention to pairs of mutations whose overlapping biological functions are less intuitive, in order to see whether we can gain more functional information about a given gene or the pathways that are regulated by it by studying its network and similarity patterns. One such example is TSHZ3, which encodes for a zinc finger transcription factor [44]. The network that is the most similar to TSHZ3's network belongs to SETD2, which encodes for a histone methyltransferase [45].…”

Detailed analysis of public RNAseq data and long non-coding RNA: a proposed enhancement to mesenchymal stem cell characterisation

“…Mlinc.28428.2 is down-regulated when JPX, SERTAD4-AS1, BOLA3-AS1, and SNRPD3 are KD, and over-expressed with the KD of PTCHD3P1, ERVK3.1, MEG3, among other lncRNAs without reported function ( Figure 5A). Interestingly, interactions between p53 pathway and JPX [51], SNRPD3 [52] and MEG3 [53,54]) respectively, have been previously reported. All these features converge on the hypothesis of a link between the function of Mlinc.28428, stress response, senescence and cellular maintenance.…”