Department of Clinical Laboratory, The Second Hospital of Shandong University, Jinan, 250033, Shandong, China.
Pathology Tissue Bank, Qilu Hospital of Shandong University, Jinan, 250012, Shandong, China.
Biol Direct. 2023 Aug 18;18(1):48. doi: 10.1186/s13062-023-00397-3.
Circular RNAs (circRNAs) have been shown to play diverse biological functions in the progression of multiple diseases. However, the impacts of circRNAs on breast cancer (BC) progression remains unclear. Therefore, the objective of this paper is to investigate the role and mechanisms of a functional circRNA in BC metastasis and immune escape.
This study used a circRNA microarray and identified a novel circRNA hsa_circ_0067842. The validation and characteristics of hsa_circ_0067842 were investigated using qRT-PCR, sanger sequencing, RNase R treatment, actinomycin D treatment and fluorescence in situ hybridization (FISH). Gain- and loss-of-function assays were performed to evaluate the biological function of hsa_circ_0067842 in BC progression and immune escape. Mechanistically, the interaction between hsa_circ_0067842 and HuR was explored by RNA pull down, mass spectrometry (MS), subcellular component protein extraction and immunofluorescence (IF). The regulatory mechanisms of hsa_circ_0067842/HuR/CMTM6/PD-L1 axis were investigated by qRT-PCR, western blot, FISH, immunoprecipitation and rescue assays.
The expression of hsa_circ_0067842 was upregulated in BC tissues and cells, which was found to be significantly associated with poor prognosis, regardless of other clinical covariates. Function assays showed that hsa_circ_0067842 promoted the migration and invasion capacities of BC cells. Moreover, co-culture experiment with peripheral blood mononuclear cells (PBMCs) showed that hsa_circ_0067842 played a role in the immune escape of BC cells. Mechanistically, our study showed that hsa_circ_0067842 interacted with HuR, affecting its nuclear translocation, thus enhancing the stability of CMTM6. CMTM6 not only enhances the migration and invasion ability of BC cells, but also affects the ubiquitination of PD-L1 and inhibits its degradation.
Collectively, our results demonstrated that hsa_circ_0067842 promoted BC progression through the HuR/CMTM6/PD-L1 axis, providing new insight and a potential target for BC prognosis and therapy.
环状 RNA（circRNAs）在多种疾病的进展中表现出多种生物学功能。然而，circRNAs 对乳腺癌（BC）进展的影响尚不清楚。因此，本文旨在研究一种功能性 circRNA 在 BC 转移和免疫逃逸中的作用和机制。
本研究使用 circRNA 微阵列鉴定了一种新型 circRNA hsa_circ_0067842。通过 qRT-PCR、Sanger 测序、RNase R 处理、放线菌素 D 处理和荧光原位杂交（FISH）验证和鉴定 hsa_circ_0067842 的特征。进行 gain- 和 loss-of-function 测定以评估 hsa_circ_0067842 在 BC 进展和免疫逃逸中的生物学功能。通过 RNA 下拉、质谱（MS）、亚细胞成分蛋白提取和免疫荧光（IF）研究 hsa_circ_0067842 与 HuR 之间的相互作用。通过 qRT-PCR、western blot、FISH、免疫沉淀和挽救测定研究 hsa_circ_0067842/HuR/CMTM6/PD-L1 轴的调节机制。
hsa_circ_0067842 在 BC 组织和细胞中的表达上调，与预后不良显著相关，与其他临床协变量无关。功能测定表明 hsa_circ_0067842 促进了 BC 细胞的迁移和侵袭能力。此外，与外周血单核细胞（PBMCs）共培养实验表明 hsa_circ_0067842 在 BC 细胞的免疫逃逸中发挥作用。机制上，我们的研究表明 hsa_circ_0067842 与 HuR 相互作用，影响其核转位，从而增强 CMTM6 的稳定性。CMTM6 不仅增强了 BC 细胞的迁移和侵袭能力，还影响 PD-L1 的泛素化并抑制其降解。
综上所述，我们的研究结果表明，hsa_circ_0067842 通过 HuR/CMTM6/PD-L1 轴促进 BC 进展，为 BC 的预后和治疗提供了新的见解和潜在靶点。
