Elsevier

Biochemical Pharmacology

Volume 58, Issue 5, 1 September 1999, Pages 749-757
Biochemical Pharmacology

Commentary
Reporter gene technology: the future looks bright

https://doi.org/10.1016/S0006-2952(99)00096-9Get rights and content

Abstract

Reporter gene technology is widely used to monitor the cellular events associated with signal transduction and gene expression. Based upon the splicing of transcriptional control elements to a variety of reporter genes (with easily measurable phenotypes), it “reports” the effects of a cascade of signalling events on gene expression inside cells. The principal advantage of these assays is their high sensitivity, reliability, convenience, and adaptability to large-scale measurements. This review summarises the current status of reporter gene technology including its role in monitoring gene transfer and expression and its development as a biological screen. With the advances in this technology and in detection methods, it is likely that luciferase and green fluorescent protein will become increasingly popular for the non-invasive monitoring of gene expression in living tissues and cells. Such techniques will be important in defining the molecular events associated with gene transcription, which has implications for our understanding of the molecular basis of disease and will influence our approach to gene therapy and drug development.

Section snippets

Reporter gene technology

The term reporter gene is used to define a gene with a readily measurable phenotype that can be distinguished easily over a background of endogenous proteins [6]. Generally, such reporters are selected on the basis of the sensitivity, dynamic range, convenience, and reliability of their assay 6, 7, 8, 9. Reporter gene technology involves controlling the activity of such genes by defined cis-regulatory sequences (response elements), which are responsive to alterations in gene regulation and

Reporter genes

Table 1summarises some of the advantages and disadvantages of the most commonly used reporter genes and further details about specific assays and detection methods have been the subject of reviews elsewhere 7, 8, 9, 13, 14. In general, reporter genes have the advantage of low background activity in cells but amplify the signal from the cell surface to produce a highly sensitive, often easily detectable, response. The choice of reporter, however, will depend on the cell line used (endogenous

Promoter analysis

Reporter gene technology was first used as a method for analysing the activity of cis-acting genetic elements such as enhancers and promoters in the upstream regions of genes. Indeed, there are many recent studies utilising the technology in this way. For example, transcription elements responsible for the basal and tissue-specific expression of receptor genes (e.g. β1-adrenergic 37, 38, m1 muscarinic [39], luteinising hormone [40], CC chemokine [41], angiotensin II type 1 [42], and

Future prospects

The challenge facing cell and molecular biologists is to decipher how cellular events occur and are regulated at the single cell and organismal level. This will require the development of simple, sensitive, noninvasive methods for the visualisation of cellular events, and the versatility of the reporter gene technology in this regard has been highlighted in the numerous, although not exhaustive, set of applications covered in this and other reviews 13, 19, 20. The advantages of these assays are

References (120)

  • D. Manen et al.

    A sensitive reporter gene system using bacterial luciferase based on a series of plasmid cloning vectors compatible with derivatives of pBR322

    Gene

    (1997)
  • A. Joyeux et al.

    Engineered cell lines as a tool for monitoring biological activity of hormone analogs

    Anal Biochem

    (1997)
  • S. Welsh et al.

    Reporter gene expression for monitoring gene transfer

    Curr Opin Biotechnol

    (1997)
  • S.E. George et al.

    Evaluation of a CRE-directed luciferase reporter gene assay as an alternative to measuring cAMP accumulation

    J Biomol Screen

    (1997)
  • S.E. George et al.

    Functional analysis of the D2L dopamine receptor expressed in a cAMP-responsive luciferase reporter cell line

    Biochem Pharmacol

    (1998)
  • M. Brini et al.

    Transfected aequorin in the measurement of cytosolic Ca2+ concentration ([Ca2+]c). A critical evaluation

    J Biol Chem

    (1995)
  • D.C. Prasher et al.

    Primary structure of the Aequorea victoria green-fluorescent protein

    Gene

    (1992)
  • J.F. Thompson et al.

    Mutation of a protease-sensitive region in firefly luciferase alters light emission properties

    J Biol Chem

    (1997)
  • K.R. Siemering et al.

    Mutations that suppress the thermosensitivity of green fluorescent protein

    Curr Biol

    (1996)
  • R. Heim et al.

    Engineering green fluorescent protein for improved brightness, longer wavelengths and fluorescence resonance energy transfer

    Curr Biol

    (1996)
  • S.W. Bahouth et al.

    Thyroid hormone induces β1-adrenergic receptor gene transcription through a direct repeat separated by five nucleotides

    J Mol Cell Cardiol

    (1997)
  • S. Pepitoni et al.

    Structure of the m1 muscarinic acetylcholine receptor gene and its promoter

    J Biol Chem

    (1997)
  • C.M. Bamberger et al.

    Regulation of the human interleukin-2 gene by the α and β isoforms of the glucocorticoid receptor

    Mol Cell Endocrinol

    (1997)
  • P. Luzi et al.

    Analysis of the 5′ flanking region of the human galactocerebrosidase (GALC) gene

    Biochem Mol Med

    (1997)
  • S.B. Zhang et al.

    Defining a functional androgen responsive element in the 5′ far upstream flanking region of the prostate-specific antigen gene

    Biochem Biophys Res Commun

    (1997)
  • A.J. Caplan et al.

    Hormone-dependent transactivation by the human androgen receptor is regulated by a dnaJ protein

    J Biol Chem

    (1995)
  • D.W. Galbraith et al.

    Flow cytometry analysis of trangene expression in higher plantsGreen fluorescent protein

    Methods Cell Biol

    (1995)
  • A. Chiocchetti et al.

    Green fluorescent protein as a reporter of gene expression in transgenic mice

    Biochim Biophys Acta

    (1997)
  • J.G. Quintana et al.

    Use of GFAP-lacZ transgenic mice to determine astrocyte fate in grafts of embryonic ventral midbrain

    Dev Brain Res

    (1998)
  • D.J. Fink et al.

    Development of an HSV-based vector for the treatment of Parkinson’s disease

    Exp Neurol

    (1997)
  • V.A. Romoser et al.

    Detection in living cells of Ca2+-dependent changes in fluorescence emission of an indicator composed of two green fluorescent protein variants linked by a calmodulin-binding sequence

    J Biol Chem

    (1997)
  • M. Kneen et al.

    Green fluorescent protein as a noninvasive intracellular pH indicator

    Biophys J

    (1998)
  • C. Brandes et al.

    Novel features of Drosophila period transcription revealed by real-time luciferase reporting

    Neuron

    (1996)
  • G.A. Rutter et al.

    Involvement of MAP kinase in insulin signalling revealed by non-invasive imaging of luciferase gene expression in single living cells

    Curr Biol

    (1995)
  • J.M. Stadel et al.

    Orphan G-protein-coupled receptorsA neglected opportunity for pioneer drug discovery

    Trends Pharmacol Sci

    (1997)
  • S.P. Manly

    In vitro biochemical screening

    J Biomol Screen

    (1997)
  • Z. Parandoosh

    Cell-based assays

    J Biomol Screen

    (1997)
  • J. Stables et al.

    A bioluminescent assay for agonist activity at potentially any G-protein-coupled receptor

    Anal Biochem

    (1997)
  • C. Stratowa et al.

    Use of a luciferase reporter system for characterizing G-protein-linked receptors

    Curr Opin Biotechnol

    (1995)
  • T. Beckers et al.

    Characterization of gonadotropin-releasing hormone analogs based on a sensitive cellular luciferase reporter gene assay

    Anal Biochem

    (1997)
  • M.J. Castañón et al.

    Functional coupling of human adenosine receptors to a ligand-dependent reporter gene system

    Biochem Biophys Res Commun

    (1994)
  • W. Chen et al.

    A colorimetric assay for measuring activation of Gs- and Gq-coupled signalling pathways

    Anal Biochem

    (1995)
  • H. Brauner-Osborne et al.

    Pharmacology of muscarinic acetylcholine receptor subtypes (m1–m5)High throughput assays in mammalian cells

    Eur J Pharmacol

    (1996)
  • A.J. Kolb et al.

    Beyond the 96-cell microplate, Instruments and assay methods for the 384-cell format

    J Biomol Screen

    (1997)
  • Hancock JT, Cell Signalling. Addison Wesley Longman, Essex, England,...
  • M. Montminy

    Transcription regulation by cyclic AMP

    Annu Rev Biochem

    (1997)
  • R.E. Jones et al.

    Mammalian cell lines engineered to identify inhibitors of specific signal transduction pathways

    Oncogene

    (1991)
  • M.J. Tsai et al.

    Molecular mechanisms of action of steroid/thyroid receptor superfamily members

    Annu Rev Biochem

    (1994)
  • T. Misteli et al.

    Application of the green fluorescent protein in cell biology and biotechnology

    Nat Biotechnol

    (1997)
  • Cited by (318)

    • Image-guided cancer immunotherapy

      2022, Engineering Technologies and Clinical Translation: Volume 3 of Delivery Strategies and Engineering Technologies in Cancer Immunotherapy
    View all citing articles on Scopus
    View full text