Exploration of the proxiOME of large subunit ribosomal proteins reveals Acl1 and Bcl1 as cooperating dedicated chaperones of Rpl1.

In eukaryotes, most newly synthesized ribosomal proteins (r-proteins) need to rapidly and safely get into the nucleus to reach their assembly site on pre-ribosomal particles. However, only for few r-proteins tailored support mechanisms involving so-called dedicated chaperones (DCs) could so far be revealed. Here, with the primary aim of identifying novel DCs, we performed TurboID-based proximity labelling with all 46 large subunit r-proteins of Saccharomyces cerevisiae, which unveiled the fungi-specific Acl1 and the conserved Bcl1 as candidate DCs of Rpl1. We show that the functionally cooperating Acl1 and Bcl1 both directly interact with Rpl1, form a trimeric Acl1-Rpl1-Bcl1 complex, and enable the nuclear import of Rpl1. Moreover, our crystal structure of the minimal Acl1-Rpl1 complex reveals how Acl1's ankyrin repeat domain shields a positively charged ribosomal RNA-binding surface of Rpl1. Our proximity labelling approach also permitted to establish novel interactions between four r-proteins and distinct importins and to illuminate r-protein neighbourhoods on successive pre-60S particles. Additionally, reciprocal proximity labelling with the known DCs indicates that almost all appear to be transiently associated with pre-ribosomal particles. Our study provides for the first time comprehensive insight into the physical proximities of large subunit r-proteins along their entire life cycle.