RNPC3 knockdown in A549 lung adenocarcinoma cells

Purpose of the study was to investigate the impact of reducing the efficiency of minor (U12-type) splicing on the growth and gene expression of a human cancer cell line. We chose the A549 cell line, derived from a human lung adenocarcinoma, for this study. There are only approximately 700 genes (3.5%) in the human genome that contain a minor intron. Most transcripts (96.5%) require only major (U2-dependent) splicing for correct gene expression. This is carried out by the major spliceosome. A different spliceosome, known as the minor or U11/12-type spliceosome, is required to excise minor introns from pre-RNA transcripts that contain them. One of the unique protein components of the minor spliceosome is a 65kDa protein encoded by the RNPC3 gene. To reduce the efficiency of minor splicing, we used siRNA knockdown methodology to deplete RNPC3 transcripts in A549 cells by approximately 70%. Specifically, A549 cells in growth phase were transfected with siRNA targeted to RNPC3 transcripts for 72h. Control A549 cells were treated in the same experiment with non-targeted (NT) siRNA molecules instead of siRNA targeted to RNPC3. Overall design: RNA-seq was performed on 8 samples of the A549 human lung adenocarcinoma derived cell line. Of these 8 samples, 4 were treated with non-targeted (NT) siRNA and are the control group, The other 4 were treated with siRNA targeted to RNPC3 and are the test group.