Fig. 2.
Ribosomal loading of Plasmodium falciparum mRNAs. (A) Comparison of ribosomal loading profile of several malaria transcripts. Malaria polysomes were harvested, profiled, and fractionated 32 hours post-infection, as described in Fig. 1. Total parasite RNA was isolated from each fraction using a mirVana Isolation Kit (Ambion). Transcript abundance was quantified in fractions across the gradient by SYBR Green qPCR for the following parasite proteins: knob-associated histidine rich protein 3 (KAHRP), ring-infected erythrocyte surface antigen (RESA), cyclic AMP-dependent protein kinase, regulatory subunit (PKA-R) and phosphoethanolamine N-methyltransferase (PEAMT) (primers indicated in Supplemental Table 1). Transcript abundance was normalized against total transcript level for each fraction within the gradient (n = 3; mean ± s.e.m.) (B) Lifecycle stage-dependent differences in ribosomal loading. At 8 hours (red) and 32 hours (blue) post-infection, malaria polysomes were harvested, profiled, and fractionated as above. RESA mRNA was quantified in fractions across the gradient by SYBR Green qPCR as described in (A) (n = 3; mean ± s.e.m.) Symbol legend: *40S ribosomal subunit, +60S ribosomal subunit, ‡80S monosome.