Mutually Exclusive Expression of Virulence Genes by Malaria Parasites Is Regulated Independently of Antigen Production (original) (raw)
Figure 3
Analysis of Levels of Transcription of the Entire var Family
All values are presented as relative copy number to the housekeeping gene seryl-tRNA synthetase (PF07_0073).
Top panel: NF54 parasites were cloned by limiting dilution, and var gene expression was measured by Q-RT-PCR as soon as the culture reached the required parasitemia, approximately 6 wk after plating. The clone NF54/C3, which was used to generate all transgenic lines, was predominantly expressing var PFD1005c (located on Chromosome 4 internal cluster) while expression of the rest of the var family was virtually undetectable.
Second panel: The recombinant line B12E3 growing under blasticidin pressure only transcribed bsd (red), while transcription levels of the rest of the var family was close to zero (blue).
Central panel: Expression of additional var genes was easily observed after the parasites were grown for 10 wk without drug pressure. At this point the culture is transcriptionally heterogeneous and bsd is no longer the dominant gene. Five genes were expressed at levels equal to or greater than the control (copy number = 1).
Fourth panel: The transcription pattern of clone DC-J that was re-cloned from this culture represents a population that had switched away from bsd and now predominantly expresses var PFD1015c. Applying blasticidin to DC-J resulted in the selection of parasites that had switched back to exclusively expressing bsd (bottom panel).