title: Tracking expression and subcellular localization of RNA and protein species using high-throughput single cell imaging flow cytometry.
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.
authors: Department of Molecular Biophysics and Biochemistry, Howard Hughes Medical Institute, Yale University, New Haven, Connecticut 06536, USA.
authors: Sumit
authors: S
authors: Borah
authors: Lisa A
authors: LA
authors: Nichols
authors: Lynn M
authors: LM
authors: Hassman
authors: Dean H
authors: DH
authors: Kedes
authors: Joan A
authors: JA
authors: Steitz
mesh_terms: D002467:Cell Nucleus / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006566:Herpesviridae Infections / Q000235:genetics / Q000378:metabolism / Q000821:virology; D019288:Herpesvirus 8, Human / Q000235:genetics; D006801:Humans; D017403:In Situ Hybridization; D039122:Poly(A)-Binding Protein I / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D015138:RNA, Nuclear / Q000235:genetics / Q000378:metabolism*; D022661:RNA, Untranslated; D012367:RNA, Viral / Q000378:metabolism*; D013347:Subcellular Fractions; D014764:Viral Proteins / Q000378:metabolism*; D014779:Virus Replication
chemical_list: D039122:Poly(A)-Binding Protein I; D012333:RNA, Messenger; D015138:RNA, Nuclear; D022661:RNA, Untranslated; D012367:RNA, Viral; D014764:Viral Proteins