Figure 2.
MRJP-3 Is a Multivalent RNA-Binding Oligomer
(A) Taxonomy tree analysis suggests that the MRJP-3 tandem-repeats region evolved in the genus Apis and is associated with jelly secretion.
(B) Purified MRJP-3 binds dsRNA and ssRNA as demonstrated by EMSA. MRJP-3 was incubated with increasing concentrations of dsRNA or ssRNA. Additional controls: dsRNA and ssRNA only, MRJP-1 mixed with 43.1 nM dsRNA or 0.3 μM ssRNA, and MRJP-3 only. Protein (42.8 μM) was applied in all MRJP-3- or MRJP-1-containing treatments.
(C) MRJP-3 efficiently binds ssRNA that is 18 nt and longer. Binding activity was tested using ssRNA substrates with different lengths and analyzed by EMSA. ssRNA (19.4 pmol) and proteins (42.8 μM) were applied in all ssRNA- and/or protein-containing treatments.
(D) Binding curves of Alexa Fluor-488-labeled 22 nt ssRNA and dsRNA to MRJP-3 in RJ buffer conditions (left and right curves, respectively). Calculated and estimated equilibrium Kd values are shown as dashed lines.
(E) The TRR of MRJP-3 is predicted to be intrinsically disordered by the PONDR VSL2 and IUPred algorithms.
(F) The TRR of MRJP-3 is required for RNP formation. Proteins (13.6 μM) and ssRNA∗ (0.2 μM) were applied in all RNA- and/or protein-containing treatments. Scale bar represents 1 μm.
See also Figure S2.