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High-throughput Gene Tagging in Trypanosoma brucei
Published on: August 12, 2016
Programmed cell death in procyclic Trypanosoma brucei rhodesiense is associated with differential expression of mRNAs
1International Livestock Research Institute, P.O. Box 30709, Nairobi, Kenya.
Abstract:
Procyclic Trypanosoma brucei rhodesiense have a cell death mechanism which can be activated by an external signal, the lectin ConA, in vitro. ConA has been shown to cause profound changes in cellular morphology and induce fragmentation of nuclear DNA in T.b. rhodesiense which are characteristic of apoptosis, a form of programmed cell death (PCD) in other eukaryotic cells. RNA analysis of trypanosomes induced to undergo PCD revealed that RNA remains intact up to 48 h into the process, a time when nuclear DNA fragmentation has already started. Using the randomly amplified differentially expressed sequences polymerase chain reaction method, ConA-induced cell death in T.b. rhodesiense is shown to be associated with differential expression of mRNAs, including up regulation of mRNAs late in the death process. The results demonstrate that trypanosomes actively participate in their own destruction through a PCD process and confirm that cell death in trypanosomes is associated with de novo gene expression.
Insights
Trypanosomes possess a programmed cell death (PCD) mechanism, activated by the lectin ConA. This process involves DNA fragmentation and differential gene expression, indicating active self-destruction in Trypanosoma brucei rhodesiense.
Area of Science:
- Molecular biology
- Parasitology
- Cell biology
Background:
- Procyclic Trypanosoma brucei rhodesiense exhibit a cell death mechanism.
- This mechanism can be triggered in vitro by the lectin Concanavalin A (ConA).
- ConA induces morphological changes and DNA fragmentation, resembling apoptosis in other eukaryotes.
Purpose of the Study:
- To investigate the molecular events during ConA-induced programmed cell death (PCD) in Trypanosoma brucei rhodesiense.
- To determine the role of gene expression in trypanosome cell death.
Main Methods:
- RNA analysis to assess RNA integrity during PCD.
- Randomly Amplified Differentially Expressed Sequences (RADES) PCR to identify differentially expressed mRNAs.
Main Results:
- RNA remained intact for up to 48 hours during PCD, even after DNA fragmentation began.
- ConA-induced cell death was associated with differential mRNA expression.
- Upregulation of specific mRNAs was observed late in the cell death process.
Conclusions:
- Trypanosomes actively participate in their own destruction via a PCD pathway.
- Cell death in trypanosomes is linked to de novo gene expression.
- This study confirms a genetically regulated cell death process in trypanosomes.
