Rapid, diffusional shuttling of poly(A) RNA between nuclear speckles and the nucleoplasm

Joan C.Ritland Politz, Richard A. Tuft, Kannanganattu V. Prasanth, Nina Baudendistel, Kevin E. Fogarty, Larry M. Lifshitz, Jörg Langowski, David L. Spector, Thoru Pederson

Research output: Contribution to journalArticle

Abstract

Speckles are nuclear bodies that contain pre-mRNA splicing factors and polyadenylated RNA. Because nuclear poly(A) RNA consists of both mRNA transcripts and nucleus-restricted RNAs, we tested whether poly(A) RNA in speckles is dynamic or rather an immobile, perhaps structural, component. Fluorescein-labeled oligo(dT) was introduced into HeLa cells stably expressing a red fluorescent protein chimera of the splicing factor SC35 and allowed to hybridize. Fluorescence correlation spectroscopy (FCS) showed that the mobility of the tagged poly(A) RNA was virtually identical in both speckles and at random nucleoplasmic sites. This same result was observed in photoactivation-tracking studies in which caged fluorescein-labeled oligo(dT) was used as hybridization probe, and the rate of movement away from either a speckle or nucleoplasmic site was monitored using digital imaging microscopy after photoactivation. Furthermore, the tagged poly(A) RNA was observed to rapidly distribute throughout the entire nucleoplasm and other speckles, regardless of whether the tracking observations were initiated in a speckle or the nucleoplasm. Finally, in both FCS and photoactivation-tracking studies, a temperature reduction from 37 to 22°C had no discernible effect on the behavior of poly(A) RNA in either speckles or the nucleoplasm, strongly suggesting that its movement in and out of speckles does not require metabolic energy.

Original languageEnglish (US)
Pages (from-to)1239-1249
Number of pages11
JournalMolecular biology of the cell
Volume17
Issue number3
DOIs
StatePublished - Mar 1 2006
Externally publishedYes

Fingerprint

Messenger RNA
Fluorescence Spectrometry
Fluorescein
Protein Splicing
RNA Precursors
HeLa Cells
Microscopy
RNA
Temperature
oligo (dT)
RNA Splicing Factors

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

Cite this

Politz, J. C. R., Tuft, R. A., Prasanth, K. V., Baudendistel, N., Fogarty, K. E., Lifshitz, L. M., ... Pederson, T. (2006). Rapid, diffusional shuttling of poly(A) RNA between nuclear speckles and the nucleoplasm. Molecular biology of the cell, 17(3), 1239-1249. https://doi.org/10.1091/mbc.E05-10-0952

Rapid, diffusional shuttling of poly(A) RNA between nuclear speckles and the nucleoplasm. / Politz, Joan C.Ritland; Tuft, Richard A.; Prasanth, Kannanganattu V.; Baudendistel, Nina; Fogarty, Kevin E.; Lifshitz, Larry M.; Langowski, Jörg; Spector, David L.; Pederson, Thoru.

In: Molecular biology of the cell, Vol. 17, No. 3, 01.03.2006, p. 1239-1249.

Research output: Contribution to journalArticle

Politz, JCR, Tuft, RA, Prasanth, KV, Baudendistel, N, Fogarty, KE, Lifshitz, LM, Langowski, J, Spector, DL & Pederson, T 2006, 'Rapid, diffusional shuttling of poly(A) RNA between nuclear speckles and the nucleoplasm', Molecular biology of the cell, vol. 17, no. 3, pp. 1239-1249. https://doi.org/10.1091/mbc.E05-10-0952
Politz, Joan C.Ritland ; Tuft, Richard A. ; Prasanth, Kannanganattu V. ; Baudendistel, Nina ; Fogarty, Kevin E. ; Lifshitz, Larry M. ; Langowski, Jörg ; Spector, David L. ; Pederson, Thoru. / Rapid, diffusional shuttling of poly(A) RNA between nuclear speckles and the nucleoplasm. In: Molecular biology of the cell. 2006 ; Vol. 17, No. 3. pp. 1239-1249.
@article{faecd36cbf5a47449d26eb3360e7fc75,
title = "Rapid, diffusional shuttling of poly(A) RNA between nuclear speckles and the nucleoplasm",
abstract = "Speckles are nuclear bodies that contain pre-mRNA splicing factors and polyadenylated RNA. Because nuclear poly(A) RNA consists of both mRNA transcripts and nucleus-restricted RNAs, we tested whether poly(A) RNA in speckles is dynamic or rather an immobile, perhaps structural, component. Fluorescein-labeled oligo(dT) was introduced into HeLa cells stably expressing a red fluorescent protein chimera of the splicing factor SC35 and allowed to hybridize. Fluorescence correlation spectroscopy (FCS) showed that the mobility of the tagged poly(A) RNA was virtually identical in both speckles and at random nucleoplasmic sites. This same result was observed in photoactivation-tracking studies in which caged fluorescein-labeled oligo(dT) was used as hybridization probe, and the rate of movement away from either a speckle or nucleoplasmic site was monitored using digital imaging microscopy after photoactivation. Furthermore, the tagged poly(A) RNA was observed to rapidly distribute throughout the entire nucleoplasm and other speckles, regardless of whether the tracking observations were initiated in a speckle or the nucleoplasm. Finally, in both FCS and photoactivation-tracking studies, a temperature reduction from 37 to 22°C had no discernible effect on the behavior of poly(A) RNA in either speckles or the nucleoplasm, strongly suggesting that its movement in and out of speckles does not require metabolic energy.",
author = "Politz, {Joan C.Ritland} and Tuft, {Richard A.} and Prasanth, {Kannanganattu V.} and Nina Baudendistel and Fogarty, {Kevin E.} and Lifshitz, {Larry M.} and J{\"o}rg Langowski and Spector, {David L.} and Thoru Pederson",
year = "2006",
month = "3",
day = "1",
doi = "10.1091/mbc.E05-10-0952",
language = "English (US)",
volume = "17",
pages = "1239--1249",
journal = "Molecular Biology of the Cell",
issn = "1059-1524",
publisher = "American Society for Cell Biology",
number = "3",

}

TY - JOUR

T1 - Rapid, diffusional shuttling of poly(A) RNA between nuclear speckles and the nucleoplasm

AU - Politz, Joan C.Ritland

AU - Tuft, Richard A.

AU - Prasanth, Kannanganattu V.

AU - Baudendistel, Nina

AU - Fogarty, Kevin E.

AU - Lifshitz, Larry M.

AU - Langowski, Jörg

AU - Spector, David L.

AU - Pederson, Thoru

PY - 2006/3/1

Y1 - 2006/3/1

N2 - Speckles are nuclear bodies that contain pre-mRNA splicing factors and polyadenylated RNA. Because nuclear poly(A) RNA consists of both mRNA transcripts and nucleus-restricted RNAs, we tested whether poly(A) RNA in speckles is dynamic or rather an immobile, perhaps structural, component. Fluorescein-labeled oligo(dT) was introduced into HeLa cells stably expressing a red fluorescent protein chimera of the splicing factor SC35 and allowed to hybridize. Fluorescence correlation spectroscopy (FCS) showed that the mobility of the tagged poly(A) RNA was virtually identical in both speckles and at random nucleoplasmic sites. This same result was observed in photoactivation-tracking studies in which caged fluorescein-labeled oligo(dT) was used as hybridization probe, and the rate of movement away from either a speckle or nucleoplasmic site was monitored using digital imaging microscopy after photoactivation. Furthermore, the tagged poly(A) RNA was observed to rapidly distribute throughout the entire nucleoplasm and other speckles, regardless of whether the tracking observations were initiated in a speckle or the nucleoplasm. Finally, in both FCS and photoactivation-tracking studies, a temperature reduction from 37 to 22°C had no discernible effect on the behavior of poly(A) RNA in either speckles or the nucleoplasm, strongly suggesting that its movement in and out of speckles does not require metabolic energy.

AB - Speckles are nuclear bodies that contain pre-mRNA splicing factors and polyadenylated RNA. Because nuclear poly(A) RNA consists of both mRNA transcripts and nucleus-restricted RNAs, we tested whether poly(A) RNA in speckles is dynamic or rather an immobile, perhaps structural, component. Fluorescein-labeled oligo(dT) was introduced into HeLa cells stably expressing a red fluorescent protein chimera of the splicing factor SC35 and allowed to hybridize. Fluorescence correlation spectroscopy (FCS) showed that the mobility of the tagged poly(A) RNA was virtually identical in both speckles and at random nucleoplasmic sites. This same result was observed in photoactivation-tracking studies in which caged fluorescein-labeled oligo(dT) was used as hybridization probe, and the rate of movement away from either a speckle or nucleoplasmic site was monitored using digital imaging microscopy after photoactivation. Furthermore, the tagged poly(A) RNA was observed to rapidly distribute throughout the entire nucleoplasm and other speckles, regardless of whether the tracking observations were initiated in a speckle or the nucleoplasm. Finally, in both FCS and photoactivation-tracking studies, a temperature reduction from 37 to 22°C had no discernible effect on the behavior of poly(A) RNA in either speckles or the nucleoplasm, strongly suggesting that its movement in and out of speckles does not require metabolic energy.

UR - http://www.scopus.com/inward/record.url?scp=33644850343&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33644850343&partnerID=8YFLogxK

U2 - 10.1091/mbc.E05-10-0952

DO - 10.1091/mbc.E05-10-0952

M3 - Article

C2 - 16371503

AN - SCOPUS:33644850343

VL - 17

SP - 1239

EP - 1249

JO - Molecular Biology of the Cell

JF - Molecular Biology of the Cell

SN - 1059-1524

IS - 3

ER -