MicroRNA PC-5p-1090抑制细胞增殖与调控鹿茸角茸性上皮生长发育分子机理研究

Deer antlers are the only mammalian appendages capable of periodical epimorphic regeneration. During the rapid growth stage, along with antler ‘tumor-like’ overgrowth from the top growth zones, antler velvet covered outside shows high proliferation and no apoptosis; however it is very resistant to tumor formation actually. This hints that velvet regeneration is a highly-controlled and well-coordinated developmental process showing homeostasis status. The mechanisms remained unclear. Previously, we isolated a novel velvety-specific-expressed microRNA, named of PC-5p-1090, from red deer antler. Our recent studies showed that PC-5p-1090 was capable of inhibiting tumor/cancer-derived cell proliferation in vitro, and able to bind directly to the seed-sequence complimentary site located in target gene mRNA, a necessary structural character for microRNA fulfilling functions. Thus, we proposed a new hypothesis that, functioned as a tumor suppressor role, PC-5p-1090 could inhibit velvet epithelia proliferation, and further regulate velvet growth and development. We will develop experiments to find reliable evidence. PC-5p-1090 would open an important way for deep understanding novel negative regulation and homeostasis maintaining mechanisms in growing antler, further this project will provide sufficient scientific evidence for developing novel nucleic acid medicine precursor and mining new medicine targets of human tumor/cancer therapy.

鹿茸角是唯一具周期性表形态再生能力的哺乳动物器官。随着快速生长期鹿茸角“肿瘤样”生长，顶端增生区茸性上皮保持着高增殖无凋亡状态，却无癌变发生。这提示茸皮再生高度可控并受到精细调控，具有稳态特性，但其机制不清。我们近期发现了一个马鹿（Cervus elaphus）茸皮特异性表达的新microRNA PC-5p-1090。预研实验显示，离体条件下PC-5p-1090有抑制肿瘤/癌症来源细胞系增殖的功能；且具miRNA行使功能时必需的结构特性，即直接结合靶基因mRNA上seed序列的互补序列。由此我们提出假说：PC-5p-1090具抑癌基因功能，可抑制茸皮细胞增殖，并参与调控茸皮生长发育。本项目拟通过实验获取可靠证据。PC-5p-1090将为我们深入理解再生鹿茸角器官/组织中细胞增殖负调控，以及整个内稳态保持的新机制提供一条重要途径，也为治疗肿瘤/癌症的潜在新药前体与靶标研究提供充分科学依据。

MicroRNA PC-5p-1090是通过高通量测序在鹿茸茸皮中发现的新microRNA，比对结果显示PC-5p-1090位于马鹿11号染色体SCAI宿主基因的内含子中，PC-5p-1090具有表达并发挥功能的结构基础，且可能参与鹿茸茸皮再生生长的调控。为阐明PC-5p-1090的功能及其机制，项目组对PC-5p-1090直接结合并调控的候选靶标基因开展研究；阐明PC-5p-1090调控的相关通路，及其对细胞增殖、迁移侵袭和凋亡的影响。结果显示：（1）筛选出35个PC-5p-1090调控的靶标基因，生物信息学分析显示靶标基因主要富集于组织器官发育、细胞分化、转录调控、细胞周期等相关通路。（2）基于PUM2、HNRNPU、MARCKS、SMARCAD1和SOX9等与细胞增殖相关的关键性靶标基因，构建了PC-5p-1090/靶标基因调控轴与通路；进一步揭示靶标基因间存在互作网络，其中PUM2能够调控HNRNPU的表达。PC-5p-1090抑制MARCKS表达时能够避免PI3K/AKT信号通路的激活。SMARCAD1能介导PC-5p-1090对Wnt/β-catenin通路的调控。（3）PC-5p-1090不仅能够抑制马鹿鹿茸软骨细胞的增殖和迁移，而且过表达PC-5p-1090也会抑制肝癌细胞的增殖、迁移侵袭和促进凋亡，MARCKS、PUM2和HNRNPU能介导PC-1090对肝癌细胞增殖、迁移、侵袭和凋亡的调控；SOX9和SMARCAD1仅介导PC-1090对细胞迁移、侵袭的调控。PC-5p-1090为我们深入理解再生鹿茸角器官/组织中细胞增殖负调控，以及整个内稳态保持的新机制提供一条重要途径，也为治疗肿瘤/癌症的潜在新药前体与靶标研究提供充分科学依据。