Efficacy of Semliki Forest virus transduction of bovine adrenal chromaffin cells: an analysis of heterologous protein targeting and distribution

Rory R Duncan, Jennifer Greaves, Sompol Tapechum, David K Apps, Michael J Shipston, Robert H Chow

Research output: Contribution to journalReview article

4 Citations (Scopus)

Abstract

In using chromaffin cells as a model for studying the mechanism of regulated exocytosis, there is a requirement for an efficient, safe, and robust system for the transduction and expression of heterologous cDNA in these cells. We have used Semliki Forest virus to transduce cDNAs encoding various proteins fused to enhanced green fluorescent protein (EGFP) into cultured bovine adrenal cells. Transduction is highly efficient but has no significant effect on the steady state levels of several endogenous proteins or of catecholamines in the transfected cells. Furthermore, the transfected cells show depolarization-induced calcium currents and nicotine-induced catecholamine release. We present data to show that virally transduced proteins are targeted to their intracellular locations correctly in chromaffin cells. The fusion protein pro-ANF-EGFP is specifically targeted to large dense-core vesicles as shown by its colocalization with acidophilic dyes and chromogranin A, making this a useful system for the study of secretory vesicle dynamics.

Original languageEnglish
Pages (from-to)641-646
Number of pages6
JournalAnnals of the New York Academy of Sciences
Volume971
Issue number1
DOIs
Publication statusPublished - Oct 2002

Fingerprint

Semliki forest virus
Chromaffin Cells
Protein Transport
Viruses
Secretory Vesicles
Catecholamines
Proteins
Complementary DNA
Chromogranin A
Exocytosis
Depolarization
Atrial Natriuretic Factor
Nicotine
Coloring Agents
Fusion reactions
Calcium
Cells
Transduction
Efficacy
Virus

Keywords

  • Animals
  • Cattle
  • Cells, Cultured
  • Chromaffin Cells
  • DNA, Complementary
  • Exocytosis
  • Gene Transfer Techniques
  • Green Fluorescent Proteins
  • Humans
  • Luminescent Proteins
  • Microscopy, Confocal
  • Recombinant Fusion Proteins
  • Semliki forest virus
  • Time Factors
  • Transduction, Genetic
  • Journal Article
  • Review

Cite this

Efficacy of Semliki Forest virus transduction of bovine adrenal chromaffin cells : an analysis of heterologous protein targeting and distribution. / Duncan, Rory R; Greaves, Jennifer; Tapechum, Sompol; Apps, David K; Shipston, Michael J; Chow, Robert H.

In: Annals of the New York Academy of Sciences, Vol. 971, No. 1, 10.2002, p. 641-646.

Research output: Contribution to journalReview article

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abstract = "In using chromaffin cells as a model for studying the mechanism of regulated exocytosis, there is a requirement for an efficient, safe, and robust system for the transduction and expression of heterologous cDNA in these cells. We have used Semliki Forest virus to transduce cDNAs encoding various proteins fused to enhanced green fluorescent protein (EGFP) into cultured bovine adrenal cells. Transduction is highly efficient but has no significant effect on the steady state levels of several endogenous proteins or of catecholamines in the transfected cells. Furthermore, the transfected cells show depolarization-induced calcium currents and nicotine-induced catecholamine release. We present data to show that virally transduced proteins are targeted to their intracellular locations correctly in chromaffin cells. The fusion protein pro-ANF-EGFP is specifically targeted to large dense-core vesicles as shown by its colocalization with acidophilic dyes and chromogranin A, making this a useful system for the study of secretory vesicle dynamics.",
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T2 - an analysis of heterologous protein targeting and distribution

AU - Duncan, Rory R

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AU - Apps, David K

AU - Shipston, Michael J

AU - Chow, Robert H

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KW - DNA, Complementary

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KW - Gene Transfer Techniques

KW - Green Fluorescent Proteins

KW - Humans

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KW - Microscopy, Confocal

KW - Recombinant Fusion Proteins

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KW - Time Factors

KW - Transduction, Genetic

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JO - Annals of the New York Academy of Sciences

JF - Annals of the New York Academy of Sciences

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