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Published on: March 5, 2018
Sf-Caspase-1-repressed stable cells: resistance to apoptosis and augmentation of recombinant protein production
Chih-Chien Lin1, John T-A Hsu, Kai-Ling Huang
1Institute of Biotechnology and Department of Life Science, National Tsing Hua University, Hsinchu, Taiwan, Republic of China.
Abstract:
Sf-Caspase-1 [Spodoptera frugiperda (fall armyworm) caspase-1] is the most studied effector caspase of Lepidoptera and its activation may lead cells to apoptosis (programmed cell death) when under UV irradiation or baculovirus infection. In the present study, we repressed the expression of Sf-caspase-1 in Sf9 (S. frugiperda 9) cells using constitutive RNA interference, and evaluated the effects of stress responses and the production of proteins in recombinant baculovirus-infected cells. The Sf-caspase-1-repressed stable cells, Sf9/pIBdsCasp-1 and Sf9/pIBdsCasp-2, showed a significant increase in resistance to UV- and baculovirus-induced apoptosis. These cells produced higher levels of both intracellular (luciferase) and extracellular [SEAP (secreted alkaline phosphatase)] recombinant proteins than the parental cells when infected with recombinant baculovirus. Thus Sf-caspase-1-repressed stable cells have a greater ability to adapt to various culture conditions, and also to provide the benefits of high-level protein production.
Insights
Repressing Sf-Caspase-1 in fall armyworm cells enhances resistance to UV and baculovirus-induced apoptosis. This leads to increased production of intracellular and extracellular recombinant proteins, improving cell adaptability and yield.
Area of Science:
- Molecular Biology
- Insect Cell Culture
- Biotechnology
Background:
- Sf-Caspase-1 is a key effector caspase in Lepidoptera, regulating apoptosis in response to stressors like UV irradiation and baculovirus infection.
- Apoptosis can limit cell viability and recombinant protein production in insect cell culture systems.
Purpose of the Study:
- To investigate the effect of repressing Sf-Caspase-1 expression on stress resistance and recombinant protein production in Sf9 cells.
- To determine if Sf-caspase-1 repression enhances cellular adaptation and protein yield in response to baculovirus infection.
Main Methods:
- Constitutive RNA interference was used to stably repress Sf-caspase-1 expression in Sf9 cells.
- Sf-caspase-1-repressed cells (Sf9/pIBdsCasp-1, Sf9/pIBdsCasp-2) were challenged with UV irradiation and baculovirus infection.
- Recombinant protein production (luciferase intracellularly, SEAP extracellularly) was quantified in infected cells.
Main Results:
- Sf-caspase-1-repressed cells exhibited significantly increased resistance to UV- and baculovirus-induced apoptosis.
- These modified cells demonstrated higher production levels of both intracellular (luciferase) and extracellular (SEAP) recombinant proteins compared to parental Sf9 cells.
- The repressed Sf-caspase-1 cells showed improved adaptability to various culture conditions.
Conclusions:
- Repression of Sf-caspase-1 enhances cellular resistance to apoptosis, offering a protective mechanism against common stressors in insect cell culture.
- Sf-caspase-1-repressed cells are a valuable tool for improving recombinant protein production yields in Lepidoptera cell lines.
- This strategy provides a method to enhance cell survival and productivity in biotechnological applications using insect cells.
