Figure 1

Effect of hCycT1 and hCRM1 expression in rat T cell lines (part 1). (A) FPM1 cells were electroporated with 2 μg pΔpol, 1 μg pMax-GFP, and 1 μg pCXN2, pCXN2hCycT1-HA, pβhCycT1, or pCXN2rCycT1-HA. After 2 days, p24 levels in the medium were measured by ELISA. The percentage of living cells was approximately 18% and approximately 95% of the living cells were GFP⁺ based on FACS analysis. The ratio of p24 in the CycT1 containing samples relative to mock treated samples was calculated. The total amount of p24 in the hCycT-HA containing sample was 119 pg. Values are means of duplicate samples. rCycT1 and hCycT1 were detected by Western blotting using anti-HA. (B) FPM1 cells were electroporated with 2 μg pΔpol, 1 μg pMax-GFP, and 0.5 μg pSRα296, pSRα hCRM1-HA, pSRαrCRM1-HA, or pSRαhCRM1. The percentage of living cells was approximately 4%, and 60% of the living cells were GFP⁺. The total amount of p24 in the sample containing hCRM1 was 146 pg. In the right panel, 1 μg pCNXhCycT1 was included. Values are means of duplicate samples. The total amount of p24 in the sample containing hCRM1 was 15.7 ng. (C) pSRα296, pSRαhCRM1-HA, or pSRαrCRM1-HA (0.5 μg) were electroporated into FPM1 and Molt4 cells, and 50 μg/ml cycloheximide was added after 24 h. The cells were then collected at 0, 6, and 12 h after the drug addition, and analyzed by Western blotting. Various amounts of the cell lysates were used for blotting (25 μg of hCRM1-HA containing FPM1, 5 μg rCRM1-HA containing FPM1, and 25 μg of hCRM1-HA or 10 μg of rCRM1-HA containing Molt4, respectively).