Reduced mobility of the alternate splicing factor (ASF) through the nucleoplasm and steady state speckle compartments

Michael J. Kruhlak, Melody A. Lever, Wolfgang Fischle, Eric Verdin, David P. Bazett-Jones, Michael J. Hendzel*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

140 Scopus citations

Abstract

Compartmentalization of the nucleus is now recognized as an important level of regulation influencing specific nuclear processes. The mechanism of factor organization and the movement of factors in nuclear space have not been fully determined. Splicing factors, for example, have been shown to move in a directed manner as large intact structures from sites of concentration to sites of active transcription, but splicing factors are also thought to exist in a freely diffusible state. In this study, we examined the movement of a splicing factor, ASF green fluorescent fusion protein (ASF-GFP) using time-lapse microscopy and the technique fluorescence recovery after photobleaching (FRAP). We find that ASF-GFP moves at rates up to 100 times slower than free diffusion when it is associated with speckles and, surprisingly, also when it is dispersed in the nucleoplasm. The mobility of ASF is consistent with frequent but transient interactions with relatively immobile nuclear binding sites. This mobility is slightly increased in the presence of an RNA polymerase II transcription inhibitor and the ASF molecules further enrich in speckles. We propose that the nonrandom organization of splicing factors reflects spatial differences in the concentration of relatively immobile binding sites.

Original languageEnglish (US)
Pages (from-to)41-51
Number of pages11
JournalJournal of Cell Biology
Volume150
Issue number1
DOIs
StatePublished - Jul 10 2000

Keywords

  • ASF/SF2
  • Cell nucleus
  • FRAP
  • IGCs
  • Nuclear matrix

ASJC Scopus subject areas

  • Cell Biology

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