High-throughput screen reveals sRNAs regulating crRNA biogenesis by targeting CRISPR leader to repress Rho termination

• Published in: Nature communications Vol. 10; no. 1; pp. 3728 - 12
• Main Authors: Lin, Ping, Pu, Qinqin, Wu, Qun, Zhou, Chuanmin, Wang, Biao, Schettler, Jacob, Wang, Zhihan, Qin, Shugang, Gao, Pan, Li, Rongpeng, Li, Guoping, Cheng, Zhenyu, Lan, Lefu, Jiang, Jianxin, Wu, Min
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 19.08.2019; Nature Publishing Group; Nature Portfolio
• Subjects: 14/35; 38/35; 38/39; 38/44; 38/47; 38/71; 38/77; 38/88; 38/90; 631/326/1321; 631/326/41; 631/326/41/2532; 631/326/41/2533; 631/326/41/2534; 82/29; Adaptive immunity; Bacterial physiology; Bacteriophages - genetics; Clustered Regularly Interspaced Short Palindromic Repeats - genetics; CRISPR; CRISPR-Cas Systems - genetics; Escherichia coli - genetics; High-Throughput Screening Assays; Humanities and Social Sciences; Immunity; Loci; Microbiology; multidisciplinary; Priming; Pseudomonas aeruginosa - genetics; Regulators; Regulatory mechanisms (biology); Rho Factor - antagonists & inhibitors; RNA ligase; RNA, Bacterial - biosynthesis; RNA, Bacterial - genetics; RNA, Small Untranslated - genetics; Science; Science (multidisciplinary); Transcription termination; Transcription Termination, Genetic - physiology
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-019-11695-8

Discovery of CRISPR-Cas systems is one of paramount importance in the field of microbiology. Currently, how CRISPR-Cas systems are finely regulated remains to be defined. Here we use small regulatory RNA (sRNA) library to screen sRNAs targeting type I-F CRISPR-Cas system through proximity ligation by T4 RNA ligase and find 34 sRNAs linking to CRISPR loci. Among 34 sRNAs for potential regulators of CRISPR, sRNA pant463 and PhrS enhance CRISPR loci transcription, while pant391 represses their transcription. We identify PhrS as a regulator of CRISPR-Cas by binding CRISPR leaders to suppress Rho-dependent transcription termination. PhrS-mediated anti-termination facilitates CRISPR locus transcription to generate CRISPR RNA (crRNA) and subsequently promotes CRISPR-Cas adaptive immunity against bacteriophage invasion. Furthermore, this also exists in type I-C/-E CRISPR-Cas, suggesting general regulatory mechanisms in bacteria kingdom. Our findings identify sRNAs as important regulators of CRISPR-Cas, extending roles of sRNAs in controlling bacterial physiology by promoting CRISPR-Cas adaptation priming. Small non-coding RNAs (sRNA) regulate bacterial functions by finding nucleic acids and proteins. Here the authors identify PhrS sRNA in Pseudomonas as a positive regulator of CRISPR, and show PhrS acts by binding to CRISPR leader, thereby preventing Rho-mediated transcription termination and promoting anti-bacteriophage immunity.