Shi Yixin, Li Dingru, Xu Yunchao, Guo Yijun, Mao Jun, Lu Ying
Liaoning Laboratory of Cancer Genomics, Department of Cell Biology, College of Basic Medical Sciences, Dalian Medical University, Dalian 116044, China.
School of Biology and Biological Engineering, South China University of Technology, Guangzhou 510641, China.
Int J Mol Sci. 2025 Mar 24;26(7):2937. doi: 10.3390/ijms26072937.
Despite advancements in diagnostic efficiency, colorectal cancer (CRC) remains a leading cause of cancer-related mortality, with increasing incidence rates. Circular RNA (circRNA) is a closed-loop, generally stable noncoding RNA that functions as a sponge for microRNAs in CRC. The purpose of this study was to investigate the function and underlying mechanism of circ_RUSC2, a new circRNA, in CRC. The expression levels of circ_RUSC2, miR-661, and TUSC2 were assessed using qRT-PCR, Western blot, and immunohistochemistry. Functional assays, including CCK-8, Transwell, and scratch wound healing, were performed to evaluate cell proliferation, migration, and invasion. RNA pull-down and actinomycin D assays were used to study RNA interactions and stability. In both CRC cells and tissues, miR-661 was markedly elevated, while circ_RUSC2 expression was considerably reduced. Poor differentiation, distant metastases, lymph node metastases, and an advanced stage were all strongly correlated with either miR-661 overexpression or circ_RUSC2 downregulation. circ_RUSC2 was more stable compared to its linear RUSC2 mRNA. CRC cell invasion, migration, and proliferation were suppressed by circ_RUSC2 ectopic expression; this inhibitory effect was restored by a miR-661 mimic. Circ_RUSC2 served as miR-661's sponge. TUSC2 counteracted the effects of miR-661, which stimulated CRC cell proliferation, migration, and invasion. At the post-transcriptional level, miR-661 controlled the expression of TUSC2 in CRC cells. In comparison to the negative control, circ_RUSC2 expression was markedly reduced, and its half-life was shortened by methyltransferase-like 3 (METTL3) knockdown. Circ_RUSC2 is a stable cytoplasmic circRNA. Circ_RUSC2 inhibits CRC cell malignant phenotypes via the miR-661/TUSC2 axis. The onset and progression of CRC are linked to the downregulation of Circ_RUSC2. circ_RUSC2 might become more stable through N6-methyladenosine (m6A) methylation regulated by METTL3. According to our research, circ_RUSC2 might be a new biomarker and treatment target for CRC.
尽管诊断效率有所提高，但结直肠癌（CRC）仍是癌症相关死亡的主要原因，且发病率不断上升。环状RNA（circRNA）是一种闭环、通常稳定的非编码RNA，在结直肠癌中充当微小RNA的海绵。本研究的目的是探讨一种新的环状RNA——circ_RUSC2在结直肠癌中的功能及其潜在机制。使用qRT-PCR、蛋白质印迹法和免疫组织化学评估circ_RUSC2、miR-661和TUSC2的表达水平。进行了包括CCK-8、Transwell和划痕伤口愈合实验在内的功能实验，以评估细胞增殖、迁移和侵袭能力。采用RNA下拉实验和放线菌素D实验研究RNA相互作用和稳定性。在结直肠癌细胞和组织中，miR-661均显著升高，而circ_RUSC2表达则明显降低。低分化、远处转移、淋巴结转移和晚期均与miR-661过表达或circ_RUSC2下调密切相关。与线性RUSC2 mRNA相比，circ_RUSC2更稳定。circ_RUSC2的异位表达抑制了结直肠癌细胞的侵袭、迁移和增殖；miR-661模拟物可恢复这种抑制作用。Circ_RUSC2充当miR-661的海绵。TUSC2抵消了miR-661刺激结直肠癌细胞增殖、迁移和侵袭的作用。在转录后水平，miR-661控制结直肠癌细胞中TUSC2的表达。与阴性对照相比，circ_RUSC2的表达明显降低，且敲低甲基转移酶样3（METTL3）可缩短其半衰期。Circ_RUSC2是一种稳定的细胞质环状RNA。Circ_RUSC2通过miR-661/TUSC2轴抑制结直肠癌细胞的恶性表型。结直肠癌的发生和进展与Circ_RUSC2的下调有关。circ_RUSC2可能通过METTL3调节的N6-甲基腺苷（m6A）甲基化而变得更稳定。根据我们的研究，circ_RUSC2可能是结直肠癌的一种新的生物标志物和治疗靶点。
