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cpp32 messenger RNA neosynthesis is induced by fatal axotomy and is not regulated by athanatal Bcl-2 over-expression
Abstract:
In vivo, neuronal over-expression of the anti-apoptotic protein Bcl-2 prevents axotomy-induced motoneuron death and prolongs life in a mouse model of familial amyotrophic lateral sclerosis. The mechanism of these protective effects is still unknown. We have examined, in situ, the influence of Bcl-2 over-expression on the messenger RNA level of two pro-apoptotic, bax and cpp32, and one anti-apoptotic, bcl-xl, regulators of neuronal death. In neonates wild-type mice, cpp32 mRNA was increased in axotomized, dying motoneurons. No changes in bax and bcl-xl messenger RNAs expression were detected. A similar course was observed in protected axotomized neonate motoneurons of transgenic mice over-expressing Bcl-2. In adult wild-type mice no motoneuron death was detected one week after axotomy: bax and cpp32 messenger RNAs were increased and bcl-xl messenger RNA was decreased. Four weeks after the lesion, 60% of the lesioned facial motoneurons had disappeared. In the remaining motoneurons only cpp32 messenger RNA expression was superior to control level. In Bcl-2 transgenic mice, no axotomy-induced facial motoneurons death was detected but the course of the neosynthesis of cell death genes messenger RNAs was similar to wild-type mice. Bax, Bcl-x and CPP32 immunoreactivity were increased in facial motoneurons after axotomy. Thus, fatal axotomy induces cell death genes bax and cpp32 messenger RNAs neosynthesis which is not prevented by athanatal Bcl-2 over-expression. This suggests that the protective effect of Bcl-2 results from interactions with Bax and CPP32 at the post-translation level without repercussion at the messenger RNA level. Axotomy induces cell death messenger RNA neosynthesis potentially harmful at long-term despite Bcl-2 over-expression.
Insights
Over-expressing the anti-apoptotic protein Bcl-2 protects neurons from death after injury. However, Bcl-2’s protective mechanism doesn't alter cell death gene messenger RNA levels, suggesting post-translational regulation.
Area of Science:
- Neuroscience
- Molecular Biology
- Genetics
Background:
- Neuronal cell death is a hallmark of neurodegenerative diseases like amyotrophic lateral sclerosis (ALS).
- The anti-apoptotic protein Bcl-2 is known to prevent neuronal death in vivo.
- The precise molecular mechanisms underlying Bcl-2's neuroprotective effects remain largely unknown.
Purpose of the Study:
- To investigate the influence of Bcl-2 over-expression on the messenger RNA (mRNA) levels of key apoptosis regulators (Bax, CPP32, Bcl-xL) in axotomized motoneurons.
- To elucidate the role of Bcl-2 in regulating the expression of pro-apoptotic and anti-apoptotic genes at the mRNA level following neuronal injury.
- To determine if Bcl-2's protective effects against axotomy-induced motoneuron death involve transcriptional changes in cell death pathway genes.
Main Methods:
- Utilized transgenic mice over-expressing Bcl-2 and wild-type littermates.
- Examined mRNA expression levels of Bax, CPP32, and Bcl-xL in facial motoneurons at various time points post-axotomy using in situ methods.
- Assessed neuronal survival and quantified changes in protein immunoreactivity for Bax, Bcl-x, and CPP32.
Main Results:
- In neonate wild-type mice, axotomy increased CPP32 mRNA, while Bax and Bcl-xL mRNA remained unchanged. Bcl-2 over-expression mirrored these findings.
- In adult wild-type mice, axotomy initially increased Bax and CPP32 mRNA, and decreased Bcl-xL mRNA. Significant motoneuron loss occurred by four weeks post-axotomy.
- Despite preventing motoneuron death, Bcl-2 over-expression did not alter the mRNA expression patterns of Bax, CPP32, or Bcl-xL following axotomy. Protein levels of Bax, Bcl-x, and CPP32 increased post-axotomy.
Conclusions:
- Fatal axotomy induces the synthesis of cell death gene mRNAs (Bax and CPP32), a process not prevented by Bcl-2 over-expression at the mRNA level.
- The neuroprotective effects of Bcl-2 likely stem from post-translational interactions with Bax and CPP32, rather than transcriptional regulation.
- Axotomy-induced mRNA neosynthesis of cell death genes may pose long-term risks, even in the presence of Bcl-2 over-expression.