Piwi like RNA-mediated gene silencing 1gene as a possible major player in gastric cancer

Abstract

AIMTo establish a permanent piwi like RNA-mediated gene silencing 1 (PIWIL1) gene knockout in AGP01 gastric cancer cell line using CRISPR-Cas9 system and analyze phenotypic modifications as well as gene expression alterations.METHODSCRISPR-Cas9 system used was purchased from Dharmacon GE Life Sciences (Lafayette, CO, United States) and permanent knockout was performed according to manufacturer’s recommendations. Wound-healing assay was performed to investigate the effect of PIWIL1 knockout on migration capability of cells and Boyden chamber invasion assay was performed to investigate the effect on invasion capability. For the gene expression analysis, a one-color microarray-based gene expression analysis kit (Agilent Technologies, Santa Clara, CA, United States) was used according to the protocol provided by the manufacturer.RESULTSPIWIL1 gene knockout caused a significant decrease in AGP01 migration capacity as well as a significant decrease in cell invasiveness. Moreover, functional analysis based on grouping of all differentially expressed mRNAs identified a total of 35 genes (5 up-regulated and 30 down-regulated) encoding proteins involved in cellular invasion and migration. According to current literature, 9 of these 35 genes (DOCK2, ZNF503, PDE4D, ABL1, ABL2, LPAR1, SMAD2, WASF3 and DACH1) are possibly related to the mechanisms used by PIWIL1 to promote carcinogenic effects related to migration and invasion, since their functions are consistent with the changes observed (being up- or down-regulated after knockout).CONCLUSIONTaken together, these data reinforce the idea that PIWIL1 plays a crucial role in the signaling pathway of gastric cancer, regulating several genes involved in migration and invasion processes; therefore, its use as a therapeutic target may generate promising results in the treatment of gastric cancer.

The emerging role of the piRNA/piwi complex in cancer

“…PIWIL1 gene knockout results in altered expression (up- or down-regulation) of numerous genes, such as DOCK2, ZNF503, PDE4D, ABL1, and ABL2, whose encoded proteins are involved in cellular invasion and migration. Consequently, PIWIL1 may play a crucial role in the GC signaling pathway, and may be useful as a therapeutic target of GC [106]. PIWIL1 mainly localizes in the cytoplasm of CRC tumor cells.…”

The Prognosis Value of PIWIL1 and PIWIL2 Expression in Pancreatic Cancer

“…In contrast, PIWIL1 expression downregulates MMP-2 and MMP-9 and inhibits the proliferation and migration ability of chronic myeloid leukemia cells [63]. In gastric cancer, PIWIL1 has been related to OCK2 , ZNF503 , PDE4D , ABL1 , ABL2 , LPAR1 , SMAD2 , WASF3 , and DACH1 genes [73], and it has exhibited a regulation activity of epithelial-to-mesenchymal transition in endometrial cancer [74]. PIWIL2 regulates BCL-XL and STAT3, and its downregulation both suppresses protein expression triggering apoptosis cascade [75], and enhances cisplatin sensitivity [76].…”

Human cathelicidin antimicrobial peptide suppresses proliferation, migration and invasion of oral carcinoma HSC-3 cells via a novel mechanism involving caspase-3 mediated apoptosis

“…The medium containing G418 was replaced every 48 h. cells were subsequently cultured with 200 ng/µl G418 in the medium to maintain cell resistance. Single positively transfected cell clones were screened using the filter paper method (27). The expression of either CDEL or CAMP was confirmed by western blot analysis and immunofluorescence (28)(29)(30).…”

“…no. sc-166770; Santa cruz Biotechnology, inc.) antibodies at room temperature for 1 h, followed by incubation with alexa Fluor 488-conjugated goat anti-rabbit igG or alexa Fluor 594-conjugated goat anti-mouse igG (ProteinTech Group, inc.) at room temperature for 1 h (27,28). nuclei were stained with 300 nM daPi at room temperature for 1 min, and the cells were then imaged using an epi fluor escence microscope (Nikon Eclipse Ti; Nikon Corporation; magnification, x400).…”