Double labelling of subcellular structures with organelle-targeted GFP mutants in vivo

Double labelling of subcellular structures with organelle-targeted GFP mutants in vivo Background: The green fluorescent protein (GFP) of Aequorea victoria is emerging as a unique tool for monitoring complex phenomena such as gene expression and organelle structure and dynamics in living cells. The recent description of GFP mutants with modified spectral properties opens numerous new applications in cell biology. However, the expression and the characteristics of these GFP mutants in living eukaryotic cells have not been verified yet. Results Here, we demonstrate the usefulness of the GFP mutants for cell biology studies in vivo , by the use of wild-type GFP, a ‘bright’ GFP mutant (S65T) and a mutant with blue-shifted excitation and emission spectra (Y66H/Y145F). We have constructed two GFP chimeras targeted to mitochondria, mtGFP(S65T) and mtGFP(Y66H/Y145F), with the same strategy used previously for mtGFP. In addition, two GFP chimeras targeted to the nucleus, nuGFP and nuGFP(S65T), were constructed by fusing the wild-type GFP or the S65T mutant to the rat glucocorticoid receptor. By co-transfecting mtGFP(Y66H/Y145F) and nuGFP, the nucleus and the mitochondria were visualized simultaneously in living cells. Similarly, mtGFP and mtGFP(Y66H/Y145F) were transfected into different populations of cells, and the events of cellular fusion, and mitochondrial intermixing and/or fusion, were directly monitored. Conclusion The successful expression of organelle-targeted GFP mutants in live eukaryotes expands the uses of this fluorescent protein in cell biology, allowing direct access to key biological issues, such as the study of the interactions of different organelles in vivo . These results also open the way to other exciting applications, such as the direct study of protein redistribution and protein–protein interactions in living cells. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Current Biology Elsevier

Double labelling of subcellular structures with organelle-targeted GFP mutants in vivo

Loading next page...
 
/lp/elsevier/double-labelling-of-subcellular-structures-with-organelle-targeted-gfp-n5JB0iF0dp
Publisher
Elsevier
Copyright
Copyright © 1996 Elsevier Science Ltd
ISSN
0960-9822
D.O.I.
10.1016/S0960-9822(02)00451-7
Publisher site
See Article on Publisher Site

Abstract

Background: The green fluorescent protein (GFP) of Aequorea victoria is emerging as a unique tool for monitoring complex phenomena such as gene expression and organelle structure and dynamics in living cells. The recent description of GFP mutants with modified spectral properties opens numerous new applications in cell biology. However, the expression and the characteristics of these GFP mutants in living eukaryotic cells have not been verified yet. Results Here, we demonstrate the usefulness of the GFP mutants for cell biology studies in vivo , by the use of wild-type GFP, a ‘bright’ GFP mutant (S65T) and a mutant with blue-shifted excitation and emission spectra (Y66H/Y145F). We have constructed two GFP chimeras targeted to mitochondria, mtGFP(S65T) and mtGFP(Y66H/Y145F), with the same strategy used previously for mtGFP. In addition, two GFP chimeras targeted to the nucleus, nuGFP and nuGFP(S65T), were constructed by fusing the wild-type GFP or the S65T mutant to the rat glucocorticoid receptor. By co-transfecting mtGFP(Y66H/Y145F) and nuGFP, the nucleus and the mitochondria were visualized simultaneously in living cells. Similarly, mtGFP and mtGFP(Y66H/Y145F) were transfected into different populations of cells, and the events of cellular fusion, and mitochondrial intermixing and/or fusion, were directly monitored. Conclusion The successful expression of organelle-targeted GFP mutants in live eukaryotes expands the uses of this fluorescent protein in cell biology, allowing direct access to key biological issues, such as the study of the interactions of different organelles in vivo . These results also open the way to other exciting applications, such as the direct study of protein redistribution and protein–protein interactions in living cells.

Journal

Current BiologyElsevier

Published: Feb 1, 1996

References

You’re reading a free preview. Subscribe to read the entire article.


DeepDyve is your
personal research library

It’s your single place to instantly
discover and read the research
that matters to you.

Enjoy affordable access to
over 18 million articles from more than
15,000 peer-reviewed journals.

All for just $49/month

Explore the DeepDyve Library

Search

Query the DeepDyve database, plus search all of PubMed and Google Scholar seamlessly

Organize

Save any article or search result from DeepDyve, PubMed, and Google Scholar... all in one place.

Access

Get unlimited, online access to over 18 million full-text articles from more than 15,000 scientific journals.

Your journals are on DeepDyve

Read from thousands of the leading scholarly journals from SpringerNature, Elsevier, Wiley-Blackwell, Oxford University Press and more.

All the latest content is available, no embargo periods.

See the journals in your area

DeepDyve

Freelancer

DeepDyve

Pro

Price

FREE

$49/month
$360/year

Save searches from
Google Scholar,
PubMed

Create folders to
organize your research

Export folders, citations

Read DeepDyve articles

Abstract access only

Unlimited access to over
18 million full-text articles

Print

20 pages / month

PDF Discount

20% off