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Cellular expression of the beige mouse mutation and its correction in hybrids with control human fibroblasts
J B Gow1, S Lainwala, T A Lyerla
1Department of Biology, Clark University, Worcester, Massachusetts 01610.
Abstract:
Fibroblasts from a beige mouse (C57BL/6J; bgJ bgJ) have been established and maintained in culture for more than 3 yr. At early passages, the mutant cells were distinguishable from C57BL/6J control mouse fibroblasts at the ultrastructural level by the presence of enlarged cytoplasmic granules. After continuous passaging, this distinguishing feature was lost from the mutant cells, correlated with their increased growth rate. Clustered, perinuclear distribution of lysosomes was retained, however, and was quantitatively different at any passage number of the beige cell line from the dispersed distribution of these organelles in control mouse fibroblasts, as analyzed by computer-aided, video-enhanced light microscopy. In somatic cell hybrids between the established beige cell line and a control human diploid fibroblast cell strain, seven uncorrected hybrid lines retained a lysosomal dispersion pattern statistically indistinguishable from that of the beige mouse cell lines. Three corrected hybrid lines had lysosomal dispersion patterns that were significantly different from the beige parent line and indistinguishable from that of the control mouse fibroblast line. Thus, lysosomal dispersion can be used objectively and quantitatively to distinguish mutant beige and control mouse fibroblasts and corrected vs. uncorrected cell hybrids made from the beige/control human somatic cell crosses.
Insights
Mouse fibroblasts with beige mutations show altered lysosome distribution. This trait, unlike other cellular changes, persists and can distinguish beige cells from normal cells and their hybrids.
Area of Science:
- Cell Biology
- Genetics
- Biochemistry
Background:
- Beige mouse fibroblasts (C57BL/6J; bgJ bgJ) exhibit ultrastructural differences, including enlarged cytoplasmic granules, compared to control C57BL/6J fibroblasts.
- These distinguishing granule features are lost with continuous culturing, correlating with an increased growth rate in mutant cells.
Purpose of the Study:
- To investigate the persistent cellular differences in beige mouse fibroblasts.
- To determine if lysosomal distribution patterns can serve as a stable marker for distinguishing beige fibroblasts and their cellular interactions.
Main Methods:
- Establishment and long-term culturing of beige mouse fibroblasts.
- Ultrastructural analysis of fibroblast morphology.
- Computer-aided, video-enhanced light microscopy to quantify lysosomal distribution.
- Somatic cell hybridization between beige mouse fibroblasts and human diploid fibroblasts.
Main Results:
- Beige fibroblasts consistently displayed a clustered, perinuclear lysosomal distribution, distinct from the dispersed pattern in control fibroblasts, across all passage numbers.
- Somatic cell hybrids revealed that uncorrected lines retained the beige lysosomal pattern, while corrected lines showed a pattern similar to control fibroblasts.
- Lysosomal dispersion proved to be a stable and quantifiable characteristic differentiating beige from control fibroblasts and hybrid cell lines.
Conclusions:
- Lysosomal dispersion is a reliable and objective marker for identifying beige mouse fibroblasts.
- This lysosomal characteristic can be used to assess the correction status in somatic cell hybrids involving beige fibroblasts.
- The findings highlight the utility of lysosomal distribution analysis in cell biology research and genetic studies.