小柯机器人

美国芝加哥大学何川研究组的研究利用原位反转录测序分析RNA结合蛋白（RBPs）的结合位点。相关论文于2024年1月10日发表于国际学术期刊《自然-方法学》杂志上。

研究人员研发了一种基于反转录的RBP结合位点测序（ARTR-seq）方法，它通过抗体介导的RBP结合RNA原位反转录来识别RBP结合位点。ARTR-seq避免了紫外线交联和免疫沉淀，可从低至20个细胞或组织切片中高效、特异地鉴定RBP结合位点。

利用甲醛快速固定的优势，ARTR-seq能够捕捉RBPs在短时间内与RNA的动态结合，在短至10分钟的应激颗粒组装过程中G3BP1的动态RNA结合分析很好的印证了这一点。

据了解，RNA结合蛋白通过与RNA靶点的动态相互作用调节多种细胞过程。然而，目前仍缺乏有效的方法来捕捉RBPs与其RNA靶点之间稳定和瞬时的相互作用，尤其是当这种相互作用是动态的或样本数量有限时。

附：英文原文

Title: Profiling of RNA-binding protein binding sites by in situ reverse transcription-based sequencing

Author: Xiao, Yu, Chen, Yan-Ming, Zou, Zhongyu, Ye, Chang, Dou, Xiaoyang, Wu, Jinjun, Liu, Chang, Liu, Shun, Yan, Hao, Wang, Pingluan, Zeng, Tie-Bo, Liu, Qinzhe, Fei, Jingyi, Tang, Weixin, He, Chuan

Issue&Volume: 2024-01-10

Abstract: RNA-binding proteins (RBPs) regulate diverse cellular processes by dynamically interacting with RNA targets. However, effective methods to capture both stable and transient interactions between RBPs and their RNA targets are still lacking, especially when the interaction is dynamic or samples are limited. Here we present an assay of reverse transcription-based RBP binding site sequencing (ARTR-seq), which relies on in situ reverse transcription of RBP-bound RNAs guided by antibodies to identify RBP binding sites. ARTR-seq avoids ultraviolet crosslinking and immunoprecipitation, allowing for efficient and specific identification of RBP binding sites from as few as 20 cells or a tissue section. Taking advantage of rapid formaldehyde fixation, ARTR-seq enables capturing the dynamic RNA binding by RBPs over a short period of time, as demonstrated by the profiling of dynamic RNA binding of G3BP1 during stress granule assembly on a timescale as short as 10minutes.

DOI: 10.1038/s41592-023-02146-w

Source: https://www.nature.com/articles/s41592-023-02146-w

Nature Methods：《自然—方法学》，创刊于2004年。隶属于施普林格·自然出版集团，最新IF：47.99
官方网址：https://www.nature.com/nmeth/
投稿链接：https://mts-nmeth.nature.com/cgi-bin/main.plex