BARD1 Deletion Suppresses Progression of Colorectal Cancer through Regulation of SLIT3/Cyclin D3

Abstract

Background: Breast-cancer susceptibility gene 1 (BRCA1) associated RING domain 1 (BARD1) expression is upregulated in colorectal cancer (CRC), and its mutation forms are also related to clinical prognosis of the cancer. The primary focus of this study is to delineate the mechanism of BARD1 underlying the development and progression of CRC. Methods: BARD1 expression pattern in CRC was uncovered by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) with the aid of bioinformatics means. Following transfection of small interfering RNA targeting BARD1 (siBARD1) and short hairpin RNA against slit guidance ligand 3 (SLIT3; shSLIT3), CRC cell viability, proliferation, apoptosis, migration, and invasion were measured by cell counting kit-8 assay, colony formation assay, flow cytometry, wound healing assay, and Transwell assay. To verify how BARD1 impacts SLIT3 degradation, CRC cells were treated with cycloheximide (CHX) for different periods of time (0, 2, 4, 6, 8 h). After administration of siBARD1-transfected CRC cells or blank treatment into Balb/c nude mice, the tumor volume and weight of the animals were determined, followed by quantification of cyclin D3 (immunohistochemistry) and corresponding genes/proteins (qRT-PCR and western blotting). Results: BARD1 expression was upregulated in CRC cells (p < 0.001). SiBARD1 reduced cell viability, proliferation, migration and invasion; increased apoptosis; upregulated E-cadherin level; and downregulated N-cadherin, Snail, and cyclin D3 levels in CRC cells (p < 0.05). In contrast, shSLIT3 presented an opposite effect on these indexes in CRC cells (p < 0.05). SiBARD1 suppressed the degradation of SLIT3, hampered tumor growth, reduced cyclin D3 expression, and promoted SLIT3 expression in vivo (p < 0.001). Additionally, shSLIT3 was found to reverse the effects of siBARD1, and vice versa (p < 0.05). Conclusion: BARD1 deletion stifled the progression of CRC in vitro and in vivo by upregulating SLIT3 expression and inhibiting cyclin D3 expression, corroborating BARD1 as a potential biomarker in CRC carcinogenesis.