Analytical subcellular distribution of calmodulin and calmodulin-binding proteins in normal and virus-transformed...

We report a quantitative subcellular localization study of calmodulin and the subcellular distribution of calmodulin-binding proteins in normal and virus-transformed chicken embryo fibroblasts. We developed homogenization conditions for fibroblasts which give maximal cell disruption with minimal organelle destruction, prepared and characterized subcellular fractions, and measured the levels and distribution of calmodulin in the fractions. We found that the majority of calmodulin is present in the soluble fraction, but that a small, reproducible amount of calmodulin is present in each of the particulate fractions. The amount of calmodulin in all subcellular fractions of transformed fibroblasts is greater than or equal to the amount of calmodulin in the corresponding fractions of normal fibroblasts. In contrast to the mostly soluble distribution of calmodulin, many of the calmodulin-binding proteins in fibroblasts are associated with the particulate fractions. Although there are no apparent qualitative differences between normal and transformed fibroblasts with respect to the number, distribution, or apparent molecular weight of the calmodulin-binding proteins, there may be quantitative changes in the levels of some calmodulin-binding proteins after transformation. These data suggest that many of the calmodulin-binding proteins may be particulate proteins rather than soluble proteins, and that several calmodulin-binding proteins in fibroblasts may bind calmodulin in a calcium-independent manner.