Group II Intron RNPs and Reverse Transcriptases: From Retroelements to Research Tools

Group II introns, self-splicing retrotransposons, serve as both targets of investigation into their structure, splicing, and retromobility and a source of tools for genome editing and RNA analysis.Here, we describe the first cryo-electron microscopy (cryo-EM) structure determination, at 3.8-4.5Å, of a group II intron ribozyme complexed with its encoded protein, containing a reverse transcriptase (RT), required for RNA splicing and retromobility.We also describe a method called RIG-seq using a retrotransposon indicator gene for high-throughput integration profiling of group II introns and other retrotransposons.Targetrons, RNA-guided gene targeting agents widely used for bacterial genome engineering, are described next.Finally, we detail thermostable group II intron RTs, which synthesize cDNAs with high accuracy and processivity, for use in various RNA-seq applications and relate their properties to a 3.0-Å crystal structure of the protein poised for reverse transcription.Biological insights from these group II intron revelations are discussed.Outline 1 Introduction 2 Technique 1. Isolation of native RNPs for cryo-EM structure determination 3 Technique 2. Retromobility indicator genes and RIG-seq for high-throughput integration profiling 4 Technique 3. Targetrons for bacterial gene targeting applications 5 Technique 4. RTs with novel enzymatic properties as potent research tools