Analysis of subcellular transcriptomes by RNA proximity labeling with Halo-seq

Kl, Engel; Hg, Lo; Goering, Raeann; Y, Li; Spitale, Robert C.; Taliaferro, J. Matthew

doi:10.1101/2021.06.08.447604

Cited by 5 publications

(2 citation statements)

References 72 publications

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“…The H2A–H2B dimer was not previously predicted and was only identified based on the cryo-EM density. In support of our identification, recent RNA proximity labelling studies show that hTR is enriched in histone H2B pulldown [ 80 ]. The P6.1 stem of the CR4/5 is highly conserved among vertebrates and essential for telomerase catalytic activity [ 29 , 81 ].…”

Section: Progresssupporting

confidence: 88%

Structural biology of human telomerase: progress and prospects

Nguyen

2021

Biochemical Society Transactions

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Telomerase ribonucleoprotein was discovered over three decades ago as a specialized reverse transcriptase that adds telomeric repeats to the ends of linear eukaryotic chromosomes. Telomerase plays key roles in maintaining genome stability; and its dysfunction and misregulation have been linked to different types of cancers and a spectrum of human genetic disorders. Over the years, a wealth of genetic and biochemical studies of human telomerase have illuminated its numerous fascinating features. Yet, structural studies of human telomerase have lagged behind due to various challenges. Recent technical developments in cryo-electron microscopy have allowed for the first detailed visualization of the human telomerase holoenzyme, revealing unprecedented insights into its active site and assembly. This review summarizes the cumulative work leading to the recent structural advances, as well as highlights how the future structural work will further advance our understanding of this enzyme.

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Section: Progresssupporting

confidence: 88%

Structural biology of human telomerase: progress and prospects

Nguyen

2021

Biochemical Society Transactions

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“…The authors also showed that dynamic changes in RNA localization could be investigated using Halo-seq. [83] Figure 6. Dibromofluorescein (DBF)-dependent RNA and protein oxidation with subsequent propargyl amine labeling.…”

Section: Photoactivated Pl Strategiesmentioning

confidence: 99%

Proximity Labeling Techniques: A Multi‐Omics Toolbox

Shkel

Kharkivska

Kim

et al. 2021

Chemistry An Asian Journal

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Proximity labeling techniques are emerging highthroughput methods for studying protein-protein, protein-RNA, and protein-DNA interactions with temporal and spatial precision. Proximity labeling methods take advantage of enzymes that can covalently label biomolecules with reactive substrates. These labeled biomolecules can be identified using mass spectrometry or next-generation sequencing. The main advantage of these methods is their ability to capture weak or transient interactions between biomolecules. Prox-imity labeling is indispensable for studying organelle interactomes. Additionally, it can be used to resolve spatial composition of macromolecular complexes. Many of these methods have only recently been introduced; nonetheless, they have already provided new and deep insights into the biological processes at the cellular, organ, and organism levels. In this paper, we review a broad range of proximity labeling techniques, their development, drawbacks and advantages, and implementations in recent studies.

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RNA localization mechanisms transcend cell morphology

Goering

Arora

Taliaferro

2022

Preprint

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RNA molecules are localized to specific subcellular regions through interactions between RNA regulatory elements and RNA binding proteins (RBPs). Generally, our knowledge of the mechanistic details behind the localization of a given RNA is restricted to a particular cell type. Here, we show that RNA/RBP interactions that regulate RNA localization in one cell type predictably regulate localization in other cell types with vastly different morphologies. To determine transcriptome-wide RNA spatial distributions across the apicobasal axis of human intestinal epithelial cells, we used our recently developed RNA proximity labeling technique, Halo-seq. We found that mRNAs encoding ribosomal proteins (RP mRNAs) were strongly localized to the basal pole of these cells. Using reporter transcripts and single molecule RNA FISH, we found that pyrimidine-rich TOP motifs in the 5′ UTRs of RP mRNAs were sufficient to drive basal RNA localization. Interestingly, the same TOP motifs were also sufficient to drive RNA localization to the neurites of mouse neuronal cells. In both cell types, the regulatory activity of the TOP motif was dependent on it being at the extreme 5′ end of the transcript, was abolished upon perturbation of the TOP-binding protein LARP1, and was reduced upon inhibition of kinesins. To extend these findings, we compared subcellular RNAseq data from neuronal and epithelial cells. We found that the basal compartment of epithelial cells and the projections of neuronal cells were enriched for highly similar sets of RNAs, indicating that broadly similar mechanisms may be transporting RNAs in both cell types. These findings identify the first RNA element known to regulate RNA localization across the apicobasal axis of epithelial cells, establish LARP1 as an RNA localization regulator, and demonstrate that RNA localization mechanisms cut across cell morphologies.

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Analysis of subcellular transcriptomes by RNA proximity labeling with Halo-seq

Cited by 5 publications

References 72 publications

Structural biology of human telomerase: progress and prospects

Structural biology of human telomerase: progress and prospects

Proximity Labeling Techniques: A Multi‐Omics Toolbox

RNA localization mechanisms transcend cell morphology

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