Teaching Resource

Monitoring Signaling Processes in Living Cells Using Biosensors

See allHide authors and affiliations

Science's STKE  21 Oct 2003:
Vol. 2003, Issue 205, pp. tr5
DOI: 10.1126/stke.2003.205.tr5

Additional Files

  • Monitoring Signaling Processes in Living Cells Using Biosensors

    Klaus Hahn*

    1Department of Cell Biology, Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA


    *Corresponding author. E-mail, khahn{at}scripps.edu


    This collection of nine animations shows how different types of biosensors report changes in cellular processes through the production of a visually detectable signal. Biosensors can be created by attaching one or more fluorescent proteins (such as green fluorescent protein) to a target protein or peptide or by attaching a fluorescent dye that is sensitive to its environment to a protein or peptide. Conformational changes in proteins in response to ligand binding, changes in the concentration of cellular metabolites or signaling messengers, changes in protein localization, and changes in protein activity or covalent modification can all be detected with biosensors. These animations can be used separately or together to illustrate how molecular biology, chemistry, and microscopy have converged to allow cellular processes to be visualized in living cells. Several of the animations describe the production of a fluorescent resonance energy transfer (FRET) signal.

    Please use the "Play" button to start the animations. Animations were created by Cameron Slayden, with scientific oversight by Klaus Hahn (Department of Cell Biology, Scripps Research Institute, CA, USA).

    Animation A. Fluorescence resonance energy transfer (FRET) reports the level and localization of a protein ligand. Two variants of green fluorescent protein (GFP) are attached to a protein or peptide that undergoes a change in conformation upon ligand binding. This conformational change alters intramolecular FRET. [Access Resource]

    Animation B. Intermolecular FRET reports protein activation. Variants of GFP are attached both to the target protein and a peptide or protein that can bind to only the activated state of the target protein. When the activated state occurs, the two fluorophores are brought close enough to undergo FRET. [Access Resource]

    Animation C. Biosensors report the accumulation of a signal in a specific cellular region. When a signal is localized in a particular subcellular compartment, a fluorescent molecule that binds specifically to that signal accumulates in the compartment, producing a local increase in fluorescence intensity. In the animation, the increase in a lipid second messenger recruits a biodetector to a region of the plasma membrane. [Access Resource]

    Animation D. Single-chain biosensors report protein activation through intramolecular FRET. Fusion of the detector (the protein or domain that binds to the activated state of the target protein), the target protein, and the fluorophores produces a single-chain biosensor. The detector binds to the activated target protein bringing the two fluorophores close enough to undergo intramolecular FRET. [Access Resource]

    Animation E. Detecting protein conformation changes with an attached fluorescent dye. Conformational changes in the protein can result in a change in either the wavelength or intensity of a dye covalently attached to the protein. These changes in fluorescence are often associated with a transition from a hydrophilic to a hydrophobic environment for the fluorophore when the attached protein changes conformation. [Access Resource]

    Animation F. FRET or a dye reports activation of an endogenous, untagged protein. Fusion of two GFP variants to the detector protein or peptide can report activation of endogenous, untagged proteins through intramolecular FRET. Alternatively, a single environmentally sensitive fluorescent dye can undergo a change in fluorescence in response to interaction with the activated form of an endogenous, untagged protein. [Access Resource]

    Animation G. Enzymatic substrate sensors report covalent peptide modifications by intramolecular FRET. The animation shows phosphorylation producing a change in the conformation of a peptide attached to two variants of GFP. This leads to intramolecular FRET. [Access Resource]

    Animation H. Permutation: Conformational changes affect the fluorescence of a GFP mutant fused to the target protein. In the animation, the protein that changes conformation is fused in frame with a GFP mutant and produces a change in the fluorescence of the GFP mutant in response to ligand binding. [Access Resource]

    Animation I. Bimolecular fluorescence complementation (BiFC) reports protein interactions. GFP can be divided into two nonfluorescent halves and each half attached to a protein domain. When the two domains interact, the GFP halves are united to produce fluorescence. [Access Resource]

    Educational Details

    Learning Resource Type: Animation

    Context: Undergraduate upper division, graduate, professional (degree program)

    Intended Users: Teacher, learner

    Intended Educational Use: Teach, learn

    Discipline: Cell biology, biochemistry, molecular biology, structural biology

    Keywords: Signal transduction, fluorescence resonance energy transfer (FRET), GFP, bimolecular fluorescence complementation (BiFC)

    Technical Details

    Format: Shockwave Flash Objects (swf file)

    Size: 16 kb to 20 kb per animation

    Requirements: Macromedia Flash 5 (http://www.macromedia.com/downloads/)

    Related Resources

    Perspective: F. Gaits, K. Hahn, Shedding light on cell signaling: Interpretation of FRET biosensors. Sci. STKE 2003, pe3 (2003). [Summary] [Full Text]

    Limits for Use

    Cost: Free

    Rights: This material may be downloaded, printed, linked to, and/or redistributed without modification for noncommercial, course-teaching purposes only, provided credit to STKE is included by listing the citation for the teaching resource. To request copies of the resource for the indicated purposes, please send a message that includes the details about the resource requested (title and URL), the location and title of the course, and the expected number of students to http://stke.sciencemag.org/cgi/feedback.


    Citation: K. Hahn, Monitoring signaling processes in living cells using biosensors. Sci. STKE2003, tr5 (2003).

    © 2003 American Association for the Advancement of Science

Stay Connected to Science Signaling