Department of Pathology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, P.R. China.
Int J Oncol. 2018 Dec;53(6):2659-2670. doi: 10.3892/ijo.2018.4560. Epub 2018 Sep 13.
KRAS oncogene point mutations occur in >95% of patients with pancreatic cancer. The KRAS protein can activate various downstream effector molecules that affect proliferation and differentiation. MS2 binding sites (MS2bs) are RNAs of 19 bp in length that can bind MS2 coat proteins with their specific stem-loop structure. The MS2 binding site sequence of the 19-nucleotide stem-loop is ACATGAGGATCACCCATGT. We constructed an expression vector that expresses the KRAS non‑coding region coupled with 12 copies of MS2bs in series and established a high-throughput library for collecting microRNA (miRNA or miR)- and long non‑coding RNA (lncRNA)-omics that regulate KRAS. To the best of our knowledge, this is the first study to combine RNA-protein interactions with RNA sequencing to obtain KRAS-associated non‑coding RNAs. As a result, we identified several miRNA precursors that belong to the let-7 and miR-34, -30 and -143 families, as well as relevant lncRNAs and their families (MALAT1, MEG3_2 and TUG1_1-4). Our databank of non‑coding RNAs (mainly miRNAs and lncRNAs) that regulate KRAS is expected to greatly enhance our understanding of KRAS regulation-associated tumorigenesis and may aid in the development of gene therapies for pancreatic cancer.
KRAS 癌基因点突变发生在>95%的胰腺癌患者中。KRAS 蛋白可以激活各种下游效应分子，影响增殖和分化。MS2 结合位点（MS2bs）是长度为 19bp 的 RNA，可以通过其特定的茎环结构与 MS2 外壳蛋白结合。19 个核苷酸茎环的 MS2 结合位点序列为 ACATGAGGATCACCCATGT。我们构建了一个表达载体，该载体表达 KRAS 非编码区，与串联的 12 个 MS2bs 结合，并建立了一个高通量文库，用于收集调节 KRAS 的 microRNA（miRNA 或 miR）和长非编码 RNA（lncRNA）组学。据我们所知，这是首次将 RNA-蛋白质相互作用与 RNA 测序相结合，以获得与 KRAS 相关的非编码 RNA。结果，我们鉴定了几个属于 let-7 和 miR-34、-30 和 -143 家族的 miRNA 前体，以及相关的 lncRNA 及其家族（MALAT1、MEG3_2 和 TUG1_1-4）。我们的 KRAS 调节非编码 RNA（主要是 miRNA 和 lncRNA）数据库有望极大地增强我们对 KRAS 调节相关肿瘤发生的理解，并可能有助于开发胰腺癌的基因治疗方法。
