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Rapid apoptotic cell death of B-cell hybridomas in absence of gene expression
1Canadian Red Cross Blood Transfusion Service, Quebec City.
Abstract:
Apoptosis is an active form of cell death which plays an important role in different biological processes. The induction of apoptosis usually requires de novo gene expression but has also been observed in certain types of cells in absence of gene expression or following a block of gene expression. We show here that inhibition of macromolecular synthesis induced the rapid apoptotic death of most antibody-secreting B-cell hybridomas. The effect was observed in presence of both protein synthesis (cycloheximide [CHX]) and transcription (actinomycin D [Act D]) inhibitors and was characterized by extensive degradation of nucleic acids (DNA and RNA) within 2-3 h of treatment. The CHX treatment not only severely impaired the proliferation of the cells but also resulted in the loss of cell viability (MTT assay) without the need of de novo gene expression. The susceptibility to apoptosis varied among different B-cell hybridomas and was inherited from the SP2/0 myeloma cell fusion partner. These results indicate the constitutive activation of a death program in B-cell hybridomas and its inhibition at a late stage by the continuous expression of gene(s) coding for short-lived protein(s). The occurrence of this phenomenon may well be related to the abundant and deregulated (translocation) expression, in this type of cells, of the c-myc gene which has recently been shown to be a potent inducer of apoptosis in growth-arrested fibroblasts.
Insights
Inhibition of macromolecular synthesis triggers rapid apoptotic cell death in B-cell hybridomas, even without new gene expression. This suggests a constitutively active cell death program regulated by short-lived proteins.
Area of Science:
- Cell Biology
- Immunology
Background:
- Apoptosis, or programmed cell death, is crucial for biological processes.
- While often requiring new gene expression, apoptosis can occur without it in some cells.
Purpose of the Study:
- To investigate the role of macromolecular synthesis inhibition in B-cell hybridoma apoptosis.
- To determine if de novo gene expression is necessary for apoptosis induction in these cells.
Main Methods:
- Treatment of B-cell hybridomas with protein synthesis inhibitor (cycloheximide) and transcription inhibitor (actinomycin D).
- Assessment of cell viability using MTT assay.
- Analysis of nucleic acid degradation.
Main Results:
- Inhibition of macromolecular synthesis rapidly induced apoptosis in most B-cell hybridomas.
- Apoptotic cell death occurred without de novo gene expression, characterized by nucleic acid degradation.
- Susceptibility to apoptosis varied among hybridomas and was linked to the SP2/0 myeloma fusion partner.
Conclusions:
- B-cell hybridomas possess a constitutively active apoptotic program.
- This death program is inhibited by the expression of short-lived proteins.
- Deregulation of the c-myc gene may contribute to this phenomenon in B-cell hybridomas.