Rhinovirus Inhibitors: Including a New Target, the Viral RNA

Figure 1. Anti-RV targets and corresponding inhibitors. Steps of the rhinovirus replication cycle targeted with antiviral compounds. Note that the publication on the pla2g16 inhibitor LEI110 does not mention anything related to RV (see text and Table 3). nt, inhibition of RVs not explicitly demonstrated but tentatively inferred from inhibition of other Enteroviruses.

Figure 2. RV-A2 RNA morphology revealed by rotary shadowing electron microscopy. Purified RV-A2 in 100 mM potassium phosphate buffer (pH 7.4) was incubated with proteinase K for 12 h at 4 °C and divided into two aliquots. One was kept at 4 °C (a); one was kept for 10 min at 90 °C followed by slow cooling (b). The samples were mixed 1:1 with 200 mM ammonium acetate, 60% (v/v) glycerol, sprayed onto freshly cleaved mica chips, and coated with 0.6 nm platinum at an angle of 7° under high vacuum. Replicas were floated onto carbon-coated grids and imaged under an FEI Morgagni transmission electron microscope.

Figure 3. Temperature dependence of the accessibility of RV-A2 RNA for the intercalating dye SYTO 82. Purified RV-A2 in 100 mM potassium phosphate buffer (pH 7.4) was incubated with proteinase K for 12 h at 4 °C, as in Figure 2a. Samples were either kept at 4 °C (native—blue) or kept for 10 min at 90 °C followed by slow cooling (refolded—red) and mixed with SYTO 82 in 100 mM potassium phosphate buffer. Three aliquots of each experimental condition were disposed into the wells of a thin-walled PCR plate, the temperature was ramped from 25–95 °C at 1.5 °C/min, and the SYTO 82 fluorescence signal was recorded.

Figure 4. Representation of two GQ stabilisers. The structural formulas of PDS and PhenDC3 were prepared using Avogrado 1.2.0 software; atom colour codes are depicted. Hydrogens of the rings were omitted.

Figure 5. PDS Fibrils. (a) Two examples of RV entrapped by PDS fibrils. RV-A2 was mixed with PDS at 20 µM in Tris buffer, transferred onto EM grids, stained with 2% phosphotungstic acid (pH 7.4), and viewed with an electron microscope. Note the pearls-on-a-string-like appearance of the filaments in the left panel; (b) solvent effect on PDS fibril formation. PDS (200 µM final concentration) dissolved in the solutions indicated was applied onto EM grids, stained, and viewed as in (a). The white arrowheads point to larger amorphous aggregates; the black arrows point to ‘protofibrils’. Both panels are reproduced from ref. [25] with permission.

Figure 6. GQRS mapper prediction of GQs in the wt poliovirus 1 P1 region (NC_002058.3; grey) vs. the recoded sequence (P1-Max = EU095952; red). The P1 region comprises nucleotides from 746–3385 of the poliovirus polyprotein.