FISH-Flow to quantify nascent and mature ribosomal RNA in mouse and human cells

Antony, Charles; Somers, Patrick; Gray, Erin M.; Pimkin, Maxim; Paralkar, Vikram R.

doi:10.1016/j.xpro.2023.102463

Cited by 2 publications

(1 citation statement)

References 4 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Ultimately, we anticipate that PERFF-seq will be particularly advantageous in settings where antibodies against marker proteins are either not available, not applicable, or poorly defined for a population of interest. In particular, as FlowFISH technologies have been established for viral gene expression 41 , ribosomal RNA content 42 , or long non-coding RNAs (Fig. 2b) 43 , our workflow provides unique access to the understudied populations defined by these markers.…”

Section: Discussionmentioning

confidence: 99%

Transcript-specific enrichment enables profiling rare cell states via scRNA-seq

Abay,

Stickels,

Takizawa

et al. 2024

Preprint

View full text Add to dashboard Cite

Single-cell genomics technologies have accelerated our understanding of cell-state heterogeneity in diverse contexts. Although single-cell RNA sequencing (scRNA-seq) identifies many rare populations of interest that express specific marker transcript combinations, traditional flow sorting limits our ability to enrich these populations for further profiling, including requiring cell surface markers with high-fidelity antibodies. Additionally, many single-cell studies require the isolation of nuclei from tissue, eliminating the ability to enrich learned rare cell states based on extranuclear protein markers. To address these limitations, we describe Programmable Enrichment via RNA Flow-FISH by sequencing (PERFF-seq), a scalable assay that enables scRNA-seq profiling of subpopulations from complex cellular mixtures defined by the presence or absence of specific RNA transcripts. Across immune populations (n = 141,227 cells) and fresh-frozen and formalin-fixed paraffin-embedded brain tissue (n = 29,522 nuclei), we demonstrate the sorting logic that can be used to enrich for cell populations via RNA-based cytometry followed by high-throughput scRNA-seq. Our approach provides a rational, programmable method for studying rare populations identified by one or more marker transcripts.

show abstract

Section: Discussionmentioning

confidence: 99%

Transcript-specific enrichment enables profiling rare cell states via scRNA-seq

Abay,

Stickels,

Takizawa

et al. 2024

Preprint

View full text Add to dashboard Cite

show abstract

Transcription factor regulation of ribosomal RNA in hematopoiesis

Paralkar

2024

Current Opinion in Hematology

Self Cite

View full text Add to dashboard Cite

Purpose of review Ribosomal RNAs (rRNAs) are transcribed within nucleoli from rDNA repeats by RNA Polymerase I (Pol I). There is variation in rRNA transcription rates across the hematopoietic tree, and leukemic blast cells have prominent nucleoli, indicating abundant ribosome biogenesis. The mechanisms underlying these variations are poorly understood. The purpose of this review is to summarize findings of rDNA binding and Pol I regulation by hematopoietic transcription factors. Recent findings Our group recently used custom genome assemblies optimized for human and mouse rDNA mapping to map nearly 2200 ChIP-Seq datasets for nearly 250 factors to rDNA, allowing us to identify conserved occupancy patterns for multiple transcription factors. We confirmed known rDNA occupancy of MYC and RUNX factors, and identified new binding sites for CEBP factors, IRF factors, and SPI1 at canonical motif sequences. We also showed that CEBPA degradation rapidly leads to reduced Pol I occupancy and nascent rRNA in mouse myeloid cells. Summary We propose that a number of hematopoietic transcription factors bind rDNA and potentially regulate rRNA transcription. Our model has implications for normal and malignant hematopoiesis. This review summarizes the literature, and outlines experimental considerations to bear in mind while dissecting transcription factor roles on rDNA.

show abstract

FISH-Flow to quantify nascent and mature ribosomal RNA in mouse and human cells

Cited by 2 publications

References 4 publications

Transcript-specific enrichment enables profiling rare cell states via scRNA-seq

Transcript-specific enrichment enables profiling rare cell states via scRNA-seq

Transcription factor regulation of ribosomal RNA in hematopoiesis

Contact Info

Product

Resources

About