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When, where and how the bridge breaks: anaphase bridge breakage plays a crucial role in gene amplification and HSR generation
- Source :
- Experimental Cell Research. 302:233-243
- Publication Year :
- 2005
- Publisher :
- Elsevier BV, 2005.
-
Abstract
- Amplified genes are frequently localized on extrachromosomal double minutes (DMs) or in chromosomal homogeneously staining regions (HSRs). We previously showed that a plasmid bearing a mammalian replication initiation region could efficiently generate DMs and HSRs after transfection into human tumor cell lines. The Breakage-Fusion-Bridge (BFB) cycle model, a classical model that explains how HSRs form, could also be used to explain how the transfected plasmids generate HSRs. The BFB cycle model involves anaphase bridge formation due to the presence of dicentric chromosomes, followed by the breakage of the bridge. In this study, we used our plasmid-based model system to analyze how anaphase bridges break during mitosis. Dual-color fluorescence in situ hybridization analyses revealed that anaphase bridges were most frequently severed in their middle irrespective of their lengths, which suggests that a structurally fragile site exists in the middle of the anaphase bridge. Breakage of the chromosomal bridges occurred prior to nuclear membrane reformation and the completion of cytokinesis, which indicates that mechanical tension rather than cytokinesis is primarily responsible for severing anaphase bridges. Time-lapse observation of living cells revealed that the bridges rapidly shrink after being severed. If HSR length was extended too far, the bridge could no longer be resolved and became tangled depending on the tension. The unbroken bridge appeared to inhibit the completion of cytokinesis. These observations strongly suggest that anaphase bridges are highly elastic and that the length of the spindle axis determines the maximal HSR length.
- Subjects :
- Recombinant Fusion Proteins
Green Fluorescent Proteins
Replication Origin
Biology
Transfection
Chromatin bridge
Cell Line, Tumor
medicine
Chromosomes, Human
Humans
Nuclear membrane
Mitosis
In Situ Hybridization, Fluorescence
Cell Line, Transformed
Cytokinesis
Anaphase
Microscopy, Video
Models, Genetic
Chromosome Fragility
Chromosomal fragile site
Gene Amplification
Chromosome Breakage
Cell Biology
Cell Transformation, Viral
Immunohistochemistry
Molecular biology
Clone Cells
Cell biology
Retroviridae
medicine.anatomical_structure
Chromosome breakage
Colorectal Neoplasms
Plasmids
Subjects
Details
- ISSN :
- 00144827
- Volume :
- 302
- Database :
- OpenAIRE
- Journal :
- Experimental Cell Research
- Accession number :
- edsair.doi.dedup.....a214d659fbc992c5259069f64bab5506