Tracking Expression and Subcellular Localization of RNA and Protein Species Using High-throughput Single Cell Imaging Flow Cytometry

Overview

Journal RNA

Specialty Molecular Biology

Date 2012 Jun 30

PMID 22745225

Citations 12

Authors

Sumit Borah

Lisa A Nichols

Lynn M Hassman

Dean H Kedes

Joan A Steitz

Affiliations

Soon will be listed here.

Abstract

We report a high-throughput application of multispectral imaging flow cytometry (MIFC) for analyzing the expression and localization of both RNA and protein molecules in a heterogeneous population of cells. The approach was developed using polyadenylated nuclear (PAN) RNA, an abundant, noncoding RNA expressed by Kaposi's sarcoma-associated herpesvirus (KSHV) during the lytic phase of infection. High levels of PAN RNA are, in part, dependent on its interaction with poly(A)-binding protein C1 (PABPC1), which relocalizes from the cytoplasm to the nucleus of lytically infected cells. We quantitatively tracked the cytoplasmic to nuclear translocation of PABPC1 and examined how this translocation relates to the expression and localization of viral RNA and protein molecules in KSHV-infected cells. This high-throughput approach will be useful for other systems in which changes in subcellular localization of RNA and protein molecules need to be monitored simultaneously.

Citing Articles

PABPC1--mRNA stability, protein translation and tumorigenesis.

Qi Y, Wang M, Jiang Q Front Oncol. 2022; 12:1025291.

PMID: 36531055 PMC: 9753129. DOI: 10.3389/fonc.2022.1025291.

Flow-FISH as a Tool for Studying Bacteria, Fungi and Viruses.

Freen-van Heeren J BioTech (Basel). 2022; 10(4).

PMID: 35822795 PMC: 9245478. DOI: 10.3390/biotech10040021.

The mA landscape of polyadenylated nuclear (PAN) RNA and its related methylome in the context of KSHV replication.

Martin S, Gan H, Toomer G, Sridhar N, Sztuba-Solinska J RNA. 2021; 27(9):1102-1125.

PMID: 34187903 PMC: 8370742. DOI: 10.1261/rna.078777.121.

Viral non-coding RNAs: Stealth strategies in the tug-of-war between humans and herpesviruses.

Tagawa T, Serquina A, Kook I, Ziegelbauer J Semin Cell Dev Biol. 2020; 111:135-147.

PMID: 32631785 PMC: 7779692. DOI: 10.1016/j.semcdb.2020.06.015.

Quantitative Fluorescence In Situ Hybridization (FISH) and Immunofluorescence (IF) of Specific Gene Products in KSHV-Infected Cells.

Vallery T, Steitz J J Vis Exp. 2019; (150).

PMID: 31524859 PMC: 6750728. DOI: 10.3791/59697.

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