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Related Experiment Videos

Calbindin D28K expression in transfected mouse NIH3T3 cells

E M Muir1, D E Lawson, F V Sepúlveda

  • 1AFRC Institute of Animal Physiology and Genetics Research, Babraham, Cambridge, UK.

Cell Calcium
|April 1, 1993
PubMed
Summary
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Calbindin D28K protein, a calcium-binding protein, was permanently expressed in mouse cells. It appears to buffer extracellular calcium entry without affecting intracellular calcium levels during serum stimulation.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Calbindin D28K (calbindin) is a calcium-binding protein found in various tissues but its specific cellular function remains largely unknown.
  • Its ability to bind calcium suggests a role in calcium homeostasis or signaling pathways.

Purpose of the Study:

  • To investigate the cellular function of calbindin.
  • To establish a cell model for studying calbindin's role in intracellular calcium handling.

Main Methods:

  • Transfection of chick calbindin cDNA into mouse NIH3T3 fibroblasts to create a stably expressing cell line.
  • Immunofluorescence microscopy to determine calbindin localization and cycloheximide treatment to assess its half-life.
  • Intracellular calcium measurements using Fura-2 to analyze calcium dynamics in response to serum stimulation.

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Main Results:

  • Calbindin was successfully and permanently expressed in NIH3T3 fibroblasts, localized throughout the cytoplasm, and exhibited a long cellular half-life.
  • The presence of calbindin did not alter the magnitude or recovery rate of intracellular calcium increases induced by serum stimulation.
  • Calbindin may function as a buffer for extracellular calcium influx into the cell.

Conclusions:

  • Calbindin D28K expression in fibroblasts does not significantly impact serum-stimulated intracellular calcium dynamics.
  • Calbindin may play a role in modulating the entry of extracellular calcium, acting as a buffer.
  • This study provides a novel cell line for further investigation into calbindin's precise physiological functions.