The pre-40S ribosome that is exported to the cytoplasm contains pre-18S rRNA that requires 3′-end maturation (termed 20S or 18S-E pre-rRNA in yeast or humans, respectively), most of the RPs found in the mature 40S, and is bound to seven AFs (Ltv1, Enp1, Rio2, Dim1, Tsr1, Nob1, and Pno1) that block 60S subunit joining, the binding sites for translation initiation factors and tRNA, and obstruct the mRNA channel (Strunk et al., 2011; Heuer et al., 2017; Scaiola et al., 2018; Johnson et al., 2017; Ameismeier et al., 2018; Ameismeier et al., 2020; McCaughan et al., 2016; Fig. 2). This is the most stable assembly intermediate in yeast and is committed to the cytoplasmic assembly cascade by Hrr25(CK1δ in humans)-dependent phosphorylation of Ltv1 (Schäfer et al., 2006; Ghalei et al., 2015; Mitterer et al., 2016) to stimulate the ordered release of Ltv1, Enp1 (BYSL in humans), and Rio2 (Ghalei et al., 2015; Huang et al., 2020). This finalizes 40S head assembly via rRNA folding, re-positioning of Rps3 (uS3), and incorporation of Rps10 (eS10) and Asc1 (RACK1 in humans; Huang et al., 2020; Zemp et al., 2014; Ghalei et al., 2015; Schäfer et al., 2006; Huang and Karbstein, 2021; Mitterer et al., 2016; Mitterer et al., 2019). The translation initiation factor eIF5B then binds the pre-40S subunit to promote 60S subunit joining, forming an 80S-like ribosome that is not bound to mRNA or tRNA and will not produce protein (Strunk et al., 2012; Lebaron et al., 2012; Rai et al., 2021). Prior to or during this transition to 80S-like ribosomes, Tsr1 and Dim1 reposition, priming Dim1 for release (Rai et al., 2021). The ATPase Fap7 binds 80S-like ribosomes (Strunk et al., 2012; Ghalei et al., 2017; Granneman et al., 2005), taking advantage of the unfolded and opened platform of 80S-like ribosomes compared with mature 80S ribosomes (Rai et al., 2021) to induce a structure resembling the rotated state, an essential intermediate during translocation, resulting in Dim1 release (Ghalei et al., 2017). The release of Dim1 repositions the 3′-end of the 18S rRNA (Johnson et al., 2017; Ameismeier et al., 2018; Ameismeier et al., 2020; Rai et al., 2021) to the Nob1 active site (Lamanna and Karbstein, 2011; Lamanna and Karbstein, 2009; Pertschy et al., 2009), thereby licensing rRNA processing. Around the time of Fap7-dependent release of Dim1, Tsr3 binds 80S-like ribosomes to add an amino-carboxypropyl (acp) group to 18S:U1191 (U1248 in humans; Hector et al., 2014; Meyer et al., 2016; Huang et al., 2022). Tsr3, whose binding site overlaps those of Rio2 and Rio1 immediately before and after, respectively, is released after modifying the rRNA, thus allowing the ATPase Rio1 to bind pre-40S (Huang et al., 2022) and remove Nob1 and Pno1 following 3′-end formation of 18S rRNA by Nob1 (Parker et al., 2019; Ameismeier et al., 2020; Plassart et al., 2021; Widmann et al., 2012; Belhabich-Baumas et al., 2017; Turowski et al., 2014). 80S-like ribosomes are then separated by the translation termination factors Dom34 and Rli1 (Strunk et al., 2012). Rps26 (eS26), whose binding was blocked by Pno1, is incorporated into the now mature, translationally competent 40S ribosome (Ameismeier et al., 2020; Strunk et al., 2011; Belhabich-Baumas et al., 2017; Heuer et al., 2017).