|Budget Amount *help
¥43,100,000 (Direct Cost : ¥43,100,000)
Fiscal Year 1996 : ¥16,600,000 (Direct Cost : ¥16,600,000)
Fiscal Year 1995 : ¥26,500,000 (Direct Cost : ¥26,500,000)
We have developed a cultured B cell line, named CH12F3, which can be induced class switch recombination by the combinational addition of TGF-b, IL-4 and CD40L.As much as 50% of cells switch from lgM to lgA after 3 days of incubation.
We have sequenced more than 200 of the DNA fragment which contain the S-S switch joint. Surprisingly the sites of the joint is not only in the S-region but distributed around the peripheral regions. However, we rarely found the joint in l-region. These result suggests that the Sregion works only as the innitiation site of S-S recombination. The place for the whole SS recombination may not be restriced in S sequence.
We have constructed a plasmid which contains both Sm and Sa sequence, and introduced it into the CH12F3 cells. By inducing the recombination to the cell, we identified that the SS recombination occurs, at least in low frequency, in the episomal DNA.Presently, we are reconstructing the plasmid to improve the frequency of SS recombination.
For isolating a new gene which may control the SS recombination event, we have invented new method of cDNA subtraction, and identified 2 genes which is specifically induced by the stimulation of cytokines (TGF-b and CD40L). Identification of the rolls of these genes are on going.
We performed the physiological analysis of the innate lymphnode and germinal center deficient mouse (aly/aly) and found that the class switch recombination rarely occurs' in this mouse. Therefore, for the SS recombination, the cooperative control by B and T cell are required in germinal center.