(a) The upper part of 18S rRNA helix h44 is in an immature position in structural state A. The cryo-EM density map corresponding to this helix has been segmented (beige density); the atomic model of rRNA h44 in structural state A is represented in golden; superimposed 18S rRNA h44 as found in structural state B and in the mature 40S subunit (PDB 6EK0 Natchiar et al., 2018) are in green and blue, respectively. (b) Close-up on the platform domain of structural state A. Segmented cryo-EM density corresponding to 18S-E pre-rRNA is shown as a grey mesh. The 3′-end of the mature 18S rRNA (nucleotides 1865–1869) is shown in black, while A1870 in the ITS1 is in red. The 18S rRNA is otherwise shown as a gray ribbon. DIM2 is in orange; RPS3a is in turquoise. NOB1 was removed from this representation for the sake of clarity. (c) The catalytic pocket of RIO1 is in an ‘active’ state within structural state B pre-40S particles, and carries an ADP and a phospho-aspartate (pD341). The cryo-EM density corresponding to ADP, p-Asp, and magnesium is shown as a green mesh. RIO1 is shown in green; ADP in dark gray, Mg2+ in dark violet, and 18S rRNA G1639 closing RIO1 catalytic domain by a pi-stacking interaction (yellow dashed line) with RIO1 Phe 328 in light gray. (d) Close-up on the platform domain of structural state B. Segmented cryo-EM density corresponding to 18S rRNA 3′-end is shown as a light gray mesh. 18S rRNA 3′-end (nucleotides 1865–1869) is shown in black, while otherwise as gray ribbon. RPS26 is in magenta and RPS3a in turquoise.