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Proceedings Paper

Dynamic nuclear protein interactions investigated using fluorescence lifetime and fluorescence fluctuation spectroscopy
Author(s): Amanda P. Siegel; Nicole M. Hays; Richard N. Day
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Paper Abstract

The discovery and engineering of novel fluorescent proteins (FPs) from diverse organisms is yielding fluorophores with exceptional characteristics for live-cell imaging. In particular, the development of FPs for Förster resonance energy transfer (FRET) microscopy and fluorescence fluctuation spectroscopy (FFS) provide important tools for monitoring dynamic protein interactions inside living cells. Fluorescence lifetime imaging microscopy (FLIM) quantitatively maps changes in the spatial distribution of donor FP lifetimes that result from FRET with acceptor FPs. FFS probes dynamic protein associations through its capacity to monitor localized protein diffusion. Here, we use FRET-FLIM combined with FFS in living cells to investigate changes in protein mobility due to protein-protein interactions involving transcription factors and chromatin modifying proteins that function in anterior pituitary gene regulation. The heterochromatin protein 1 alpha (HP1α) plays a key role in the establishment and maintenance of heterochromatin through its interactions with histone methyltransferases. Recent studies, however, also highlight the importance of HP1α as a positive regulator of active transcription in euchromatin. Intriguingly, we observed that the transcription factor CCAAT/enhancer-binding protein alpha (C/EBPα) interacts with HP1α in regions of pericentromeric heterochromatin in mouse pituitary cells. These observations prompted us to investigate the relationship between HP1α dynamic interactions in pituitary specific gene regulation.

Paper Details

Date Published: 9 February 2012
PDF: 9 pages
Proc. SPIE 8226, Multiphoton Microscopy in the Biomedical Sciences XII, 82260B (9 February 2012); doi: 10.1117/12.912883
Show Author Affiliations
Amanda P. Siegel, Indiana Univ. School of Medicine (United States)
Nicole M. Hays, Indiana Univ. School of Medicine (United States)
Richard N. Day, Indiana Univ. School of Medicine (United States)


Published in SPIE Proceedings Vol. 8226:
Multiphoton Microscopy in the Biomedical Sciences XII
Karsten König, Editor(s)

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