Abstract
miR-29b2 and miR-29c play a suppressive role in breast cancer progression. C1orf132 (also named MIR29B2CHG) is the host gene for generating both microRNAs. However, the region also expresses longer transcripts with unknown functions. We employed bioinformatics and experimental approaches to decipher C1orf132 expression and function in breast cancer tissues. We also used the CRISPR/Cas9 technique to excise a predicted C1orf132 distal promoter and followed the behavior of the edited cells by real-time PCR, flow cytometry, migration assay, and RNA-seq techniques. We observed that C1orf132 long transcript is significantly downregulated in triple-negative breast cancer. We also identified a promoter for the longer transcripts of C1orf132 whose functionality was demonstrated by transfecting MCF7 cells with a C1orf132 promoter-GFP construct. Knocking-out the promoter by means of CRISPR/Cas9 revealed no alterations in the expression of the neighboring genes CD46 and CD34, while the expression of miR-29c was reduced by half. Furthermore, the promoter knockout elevated the migration ability of the edited cells. RNA sequencing revealed many up- and downregulated genes involved in various cellular pathways, including epithelial to mesenchymal transition and mammary gland development pathways. Altogether, we are reporting here the existence of an additional/distal promoter with an enhancer effect on miR-29 generation and an inhibitory effect on cell migration.
Highlights
No ortholog gene for C1orf132 has been reported in mice, mostly because Long non-coding RNAs (lncRNAs) are not conserved during evolution
We searched for potential lncRNAs overlapping with miR-29b2 and miR-29c in mouse genome
We demonstrated that fibroblast growth factor 2 (FGF2) and FGFR1 were differentially overexpressed in edited MCF12 cells by almost four-fold
Summary
Licensee MDPI, Basel, Switzerland.Attribution (CC BY) license (https://creativecommons.org/licenses/by/Breast cancer (BC) is the leading cause of cancer-related death in women worldwide, with an estimated 2,100,000 new cases and 627,000 deaths in 2018 [1,2]. In Iran, the age distribution of BC is nearly 10 years lower than that of counterparts in many countries [3].Despite the increased incidence rate, BC is highly curable if diagnosed and appropriately treated at early stages. Clinically, classification of BC is based on histopathological findings of the patients’ tissue samples obtained through surgery or biopsy. This includes the assessment of the receptor status of tumor cells, namely estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) [4,5]. One important
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