Investigation of the Role of Srsf1 in Class Switch Recombination
Date Issued
2014
Date
2014
Author(s)
Lin, Jia-Yu
Abstract
To effectively protect the host against different kinds of pathogens, B cells are capable to secret various isotypes of antibodies. Through class switch recombination (CSR), the immunoglobulin isotype of B cells switch from IgM to other isotypes. Srsf1 was known to maintain genome stability by preventing R-loops formation, and its isoform Srsf1-3 might be involved in somatic hypermutation (SHM). Since R-loop structures exist in the CSR process and SHM shares similar mechanisms with CSR, we speculated that Srsf1 might also participate in CSR.
The aim of the thesis is to investigate the role of Srsf1 in CSR from two aspects. First, the characteristics changes of Srsf1 during CSR were addressed, including the expression level and subcellular location of Srsf1. Second, the srsf1 gene was knocked out in CH12F3 cells, a murine B cell line as the CSR model. After CSR stimulation, the mRNA amounts of srsf1-1 remained unchanged in two-day-stimulated group, and decrease by 10% in three-day-stimulated group, and the protein amount of Srsf1-1 decreased slightly by about 20% after stimulation for two days and three days. The mRNA amounts of srsf1-3 decreased slightly by about 20% after stimulation for two days and three days. Next, Srsf1-1 was found mostly located in the nuclear fraction in both unstimulated and stimulated cells. Moreover, the Srsf1-1 amount in the cytoplasm fraction decreased apparently by about 40%. However, the change of the Srsf1 amount in the nuclear fraction was not determined yet due to the uncertainty of the suitable internal control. Finally, srsf1+/- cell clones were generated by knocking out the srsf1 gene in CH12F3 cells. Preliminary results of srsf1+/- cells showed no significant influence on CSR. Nevertheless, srsf1-/- cells failed to be obtained possibly due to the important role of Srsf1 in cell survival. In conclusion, the role of Srsf1 in CSR needs more experiments to be confirmed.
Subjects
抗體類型轉換重組
Srsf1
Srsf1異構型
核質分離
基因剔除
Type
thesis
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