Acquisition of Drug Resistance and Dependence by Prions
Figure 5
Standard Scrapie Cell Assay of swa-resistant and swa-dependent RML prions transferred to PK1 cells and propagated in the presence or absence of swa.
AMO10 and 2E4 cells were infected with RML prions, cultured in the presence of swa, and prions secreted into the conditioned medium (CM) were concentrated (CCM) and used to infect fresh batches of cells in the constant presence of swa. This cycle was repeated once more, whereupon the prions were transferred to PK1 cells in the presence or absence of swa (Figure 2B, C). CCM was analyzed by the SSCA on PK1 cells in the absence (blue line) or presence (red, dashed line) of swa. A. Swa-resistant AMO10-derived prions developed swa dependence when propagated in PK1 cells in the presence of swa (<4>); when propagated in PK1 cells in the absence of swa, they reverted to swa sensitivity (<5>). In contrast, swa-dependent 2E4-derived prions remained swa dependent in PK1 cells, whether they were propagated in the presence (<7>) or absence (<8>) of swa. RIs are the reciprocals of the dilutions yielding 500 PrPres positive cells per 20000 cells. Qswa = RIcell/RIcell+swa reflects inhibition by swa and may be compared to the value for swa-sensitive, brain-derived RML prions. B. AMO10- or 2E4-derived prions that had acquired or retained swa dependence after propagation in PK1 cells in the presence of swa were either continuously propagated in the presence of swa (<10>, <14>) or cultured for eleven splits in the absence of swa (<11>, <15>). Once swa dependent, they remained dependent, whether propagated in the presence or absence of swa. RIs are the reciprocals of the dilutions required to yield 1000 PrPres positive cells per 20000 cells. Qswa for RMLbrain is unusually high in this assay and that of Figure 4C, leading to the low Qrel values shown in Table 1.