CRISPR Interference (CRISPRi) and CRISPR Activation (CRISPRa) to Explore the Oncogenic lncRNA Network

• DOI: 10.1007/978-1-0716-1581-2_13
• Publication Date: 2021-06-23T11:06:11Z
• Authors: Eugenio Morelli; Annamaria Gulla’; Nicola Amodio; Elisa Taiana; Antonino Neri; Mariateresa Fulci...; Nikhil C. Munshi
• Keywords: Gene Editing; Genetic Vectors; Oncogenes; RNA, Guide, CRISPR-Cas Systems; 3. Good health; Gene Expression Regulation, Neoplastic; Long noncoding RNA; lncRNA; CRISPR interference; CRISPRi; CRISPR activation; CRISPRa; Cancer; Multiple myeloma; Transduction, Genetic; Cell Line, Tumor; Mutation; Humans; Clustered Regularly Interspaced Short Palindromic Repeats; Gene Regulatory Networks; RNA, Long Noncoding; CRISPR-Cas Systems; Multiple Myeloma

The human genome contains thousands of long noncoding RNAs (lncRNAs), even outnumbering protein-coding genes. These molecules can play a pivotal role in the development and progression of human disease, including cancer, and are susceptible to therapeutic intervention. Evidence of biologic function, however, is still missing for the vast majority of them. Both loss-of-function (LOF) and gain-of-function (GOF) studies are therefore necessary to advance our understanding of lncRNA networks and programs driving tumorigenesis. Here, we describe a protocol to perform lncRNA's LOF or GOF studies in multiple myeloma (MM) cells, using CRISPR interference (CRISPRi) or CRISPR activation (CRISPRa) technologies, respectively. These approaches have many advantages, including applicability to large-scale genetic screens in mammalian cells and possible reversibility of modulating effects; moreover, CRISPRa offers the unique opportunity to enhance lncRNA expression at the site of transcription, with relevant biologic implications.