转移RNA衍生片段(tRF)是一类新的非编码小RNA。最近的研究表明,tRF参与了一些病理过程。然而,tRFs在非小细胞肺癌(NSCLC)中的生物学功能和作用机制尚不清楚.
使用9对来自NSCLC患者的手术前和手术后血浆,通过tRF和tiRNA测序鉴定差异表达的tRF。定量实时PCR(qRT-PCR)和荧光原位杂交(FISH)用于确定组织中的tRF水平,等离子体,和细胞。实施了功能增益和功能丧失实验以研究tRF对NSCLC细胞的体外和体内致癌作用。染色质免疫沉淀(ChIP),荧光素酶报告基因,RNA下拉,质谱,RNA免疫沉淀(RIP),蛋白质印迹,免疫共沉淀(Co-IP)测定,并进行了抢救实验,以探讨tRF在NSCLC中的调控机制。
AS-tDR-007333是一种未表征的tRF,在NSCLC组织中显著上调,等离子体,和细胞。临床上,AS-tDR-007333过表达可以将NSCLC患者与健康对照区分开来,并与NSCLC患者预后较差相关。功能上,AS-tDR-007333的过表达增强了NSCLC细胞的增殖和迁移,而AS-tDR-007333的敲低导致相反的效果。机械上,AS-tDR-007333通过两种不同的机制激活MED29促进NSCLC细胞的恶性化。首先,AS-tDR-007333与HSPB1结合并相互作用,通过增强MED29启动子中的H3K4me1和H3K27ac激活MED29表达。第二,AS-tDR-007333刺激转录因子ELK4的表达,该转录因子与MED29启动子结合并增加其转录。治疗学上,AS-tDR-007333的抑制抑制了体内NSCLC细胞的生长。
我们的研究确定了一种新的致癌tRF,并揭示了AS-tDR-007333通过HSPB1-MED29和ELK4-MED29轴促进NSCLC恶性肿瘤的新机制。AS-tDR-007333是NSCLC的潜在诊断或预后标志物和治疗靶标。
Transfer RNA-derived fragments (tRFs) are a new class of small non-coding RNAs. Recent studies suggest that tRFs participate in some pathological processes. However, the biological functions and mechanisms of tRFs in non-small cell lung cancer (NSCLC) are largely unknown.
Differentially expressed tRFs were identified by tRF and tiRNA sequencing using 9 pairs of pre- and post-operation plasma from patients with NSCLC. Quantitative real-time PCR (qRT-PCR) and fluorescence in situ hybridization (FISH) were used to determine the levels of tRF in tissues, plasma, and cells. Gain- and loss-of-function experiments were implemented to investigate the oncogenic effects of tRF on NSCLC cells in vitro and in vivo. Chromatin immunoprecipitation (ChIP), luciferase reporter, RNA pulldown, mass spectrum, RNA immunoprecipitation (RIP), Western blot, co-immunoprecipitation (Co-IP) assays, and rescue experiments were performed to explore the regulatory mechanisms of tRF in NSCLC.
AS-tDR-007333 was an uncharacterized tRF and significantly up-regulated in NSCLC tissues, plasma, and cells. Clinically, AS-tDR-007333 overexpression could distinguish NSCLC patients from healthy controls and associated with poorer prognosis of NSCLC patients. Functionally, overexpression of AS-tDR-007333 enhanced proliferation and migration of NSCLC cells, whereas knockdown of AS-tDR-007333 resulted in opposite effects. Mechanistically, AS-tDR-007333 promoted the malignancy of NSCLC cells by activating MED29 through two distinct mechanisms. First, AS-tDR-007333 bound to and interacted with HSPB1, which activated MED29 expression by enhancing H3K4me1 and H3K27ac in MED29 promoter. Second, AS-tDR-007333 stimulated the expression of transcription factor
ELK4, which bound to MED29 promoter and increased its transcription. Therapeutically, inhibition of AS-tDR-007333 suppressed NSCLC cell growth in vivo.
Our study identifies a new oncogenic tRF and uncovers a novel mechanism that AS-tDR-007333 promotes NSCLC malignancy through the HSPB1-MED29 and
ELK4-MED29 axes. AS-tDR-007333 is a potential diagnostic or prognostic marker and therapeutic target for NSCLC.