Detecting dynamics

A new live-cell-imaging approach that gives insights into the dynamic nature of protein-interaction networks in intact cell nuclei is now described by Day and colleagues in Nature Methods. This approach combines the use of photoactivated green fluorescent protein (PA-GFP) and fluorescence resonance energy transfer (FRET) microscopy, and has been named photoquenching FRET (PQ-FRET).

The transcription factor CCAAT/enhancer-binding protein-α (C/EBPα) surprisingly localizes to regions of heterochromatin that are usually associated with transcriptional silencing and are marked by heterochromatin protein-1α (HP1α). The authors therefore developed PQ-FRET to define the interaction between HP1α and C/EBPα. By photoactivating PA-GFP–HP1α, they could monitor the mobility of this protein. Furthermore, using PA-GFP–HP1α as a photoactivatable FRET acceptor and cyan fluorescent protein (CFP)–C/EBPα as a FRET donor, they could quantify the dynamic interaction between HP1α and C/EBPα. They showed, for the first time, that there is a dynamic association of HP1α and C/EBPα in regions of heterochromatin. They have also provided an assay that can directly measure a protein's mobility, its exchange within macromolecular complexes and its interactions with other proteins in living cells. REFERENCE Demarco, I. A. et al. Monitoring dynamic protein interactions with photoquenching FRET. Nature Methods 3, 519–524 (2006)

Seeing inside

The optical absorption of biological tissues is strongly associated with physiological status. Despite this, current high-resolution optical imaging techniques such as confocal microscopy do not detect optical absorption directly. Furthermore, optical scattering prevents these techniques from penetrating deeper than ∼1 mm beneath the surface of tissues. However, in Nature Biotechnology, Wang and co-workers now describe a functional photoacoustic microscopy (fPAM) system that measures optical absorption directly and has been shown to have an imaging depth of more than 3 mm in live animals.

The irradiation of biological tissues with a short-pulsed laser induces wideband ultrasonic waves that are known as photoacoustic waves. fPAM detects absorbed photons ultrasonically through the photoacoustic effect. The authors have shown that their fPAM system has several in vivo imaging applications. For example, they showed that it can be used to image angiogenesis, which is potentially useful for understanding tumour growth and metastasis, for diagnosing cancers and for evaluating therapies. They also showed that fPAM allows the vessel-by-vessel mapping of the oxygen saturation of haemoglobin, which permits the monitoring of, for example, oxygen consumption by tumours. This technique, which is safe for human subjects, is therefore likely to have applications in clinical and basic research. REFERENCEZhang, H. F. et al. Functional photoacoustic microscopy for high-resolution and noninvasive in vivo imaging. Nature Biotechnol. 25 June 2006 doi:10.1038/nbt1220