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Dominant inhibitory Ras delays Sindbis virus-induced apoptosis in neuronal cells
A K Joe1, G Ferrari, H H Jiang
1Department of Medicine, Columbia University College of Physicians and Surgeons, New York, New York 10032, USA.
Abstract:
Mature neurons are more resistant than dividing cells or differentiating neurons to Sindbis virus-induced apoptotic death. Therefore, we hypothesized that mitogenic signal transduction pathways may influence susceptibility to Sindbis virus-induced apoptosis. Since Ras, a 21-kDa GTP-binding protein, plays an important role in cellular proliferation and neuronal differentiation, we investigated the effect of an inducible dominant inhibitory Ras on Sindbis virus-induced death of a rat pheochromocytoma cell line, PC12 cells. Dexamethasone induction of dominant inhibitory Ras (Ha Ras(Asn17)) expression in transfected PC12 cell lines (MMTV-M17-21 and GSrasDN6 cells) resulted in a marked delay in Sindbis virus-induced apoptosis, compared with infected, uninduced cells. The delay in death after Sindbis virus infection in induced versus uninduced PC12 cells was not associated with differences in viral titers or viral infectivity. No delay in Sindbis virus-induced apoptosis was observed in Ha Ras(Asn17)-transfected PC12 cells if dexamethasone induction was initiated less than 12 h before Sindbis virus infection or in wild-type PC12 cells infected with a chimeric Sindbis virus construct that expresses Ha Ras(Asn17). The delay in Sindbis virus-induced apoptosis in induced Ha Ras(Asn17)-transfected PC12 cells was associated with a decrease in cellular DNA synthesis as measured by 5'-bromo-2'-deoxyuridine incorporation. Thus, in PC12 cells, inducible dominant inhibitory Ras inhibits cellular proliferation and delays Sindbis virus-induced apoptosis. These findings suggest that a Ras-dependent signaling pathway is a determinant of neuronal susceptibility to Sindbis virus-induced apoptosis.
Insights
Inhibiting Ras signaling in PC12 cells delays Sindbis virus-induced apoptosis, suggesting Ras pathways influence neuronal death. This research highlights Ras-dependent signaling as a key factor in neuronal susceptibility to viral apoptosis.
Area of Science:
- Neuroscience
- Molecular Biology
- Virology
Background:
- Mature neurons exhibit greater resistance to Sindbis virus-induced apoptosis than dividing or differentiating cells.
- Mitogenic signal transduction pathways are hypothesized to influence susceptibility to Sindbis virus-induced apoptosis.
- Ras, a GTP-binding protein, is crucial for cellular proliferation and neuronal differentiation.
Purpose of the Study:
- To investigate the effect of inducible dominant-negative Ras on Sindbis virus-induced apoptosis in PC12 cells.
- To determine if Ras signaling pathways mediate neuronal susceptibility to Sindbis virus infection.
Main Methods:
- PC12 cell lines were transfected to express inducible dominant-inhibitory Ras (Ha-Ras(Asn17)).
- Dexamethasone was used to induce dominant-inhibitory Ras expression.
- Sindbis virus infection was performed, and apoptosis was assessed.
- Viral titers, infectivity, and cellular DNA synthesis (using 5'-bromo-2'-deoxyuridine incorporation) were measured.
Main Results:
- Inducible dominant-inhibitory Ras expression significantly delayed Sindbis virus-induced apoptosis in PC12 cells.
- The delay in apoptosis was not linked to altered viral titers or infectivity.
- Inhibition of Ras signaling was associated with decreased cellular DNA synthesis.
- No delay was observed if induction occurred shortly before infection or in wild-type cells infected with a chimeric virus expressing Ha-Ras(Asn17).
Conclusions:
- Inducible dominant-inhibitory Ras inhibits proliferation and delays Sindbis virus-induced apoptosis in PC12 cells.
- Ras-dependent signaling pathways are determinants of neuronal susceptibility to Sindbis virus-induced apoptosis.
- These findings provide insights into the molecular mechanisms underlying neuronal vulnerability to viral-induced cell death.