Principles of mitoribosomal small subunit assembly in eukaryotes

Abstract

Mitochondrial ribosomes (mitoribosomes) synthesize proteins encoded within the mitochondrial genome that are assembled into oxidative phosphorylation complexes. Thus, mitoribosome biogenesis is essential for ATP production and cellular metabolism1. Here we used cryo-electron microscopy to determine nine structures of native yeast and human mitoribosomal small subunit assembly intermediates, illuminating the mechanistic basis for how GTPases are used to control early steps of decoding centre formation, how initial rRNA folding and processing events are mediated, and how mitoribosomal proteins have active roles during assembly. Furthermore, this series of intermediates from two species with divergent mitoribosomal architecture uncovers both conserved principles and species-specific adaptations that govern the maturation of mitoribosomal small subunits in eukaryotes. By revealing the dynamic interplay between assembly factors, mitoribosomal proteins and rRNA that are required to generate functional subunits, our structural analysis provides a vignette for how molecular complexity and diversity can evolve in large ribonucleoprotein assemblies.

Let’s make it clear: Systematic exploration of mitochondrial DNA- and RNA-protein complexes by complexome profiling

“…Particularly, 12S mt-rRNA-binding assembly factors NOA1, ERAL1, TFB1M and MCAT, comigrated with SSU and did not change substantially their migration patterns and abundance in response to EtBr (67%, 55%, 62%, 67% of total abundance in control, respectively) (Figure 9C). These proteins are involved in maturation of the decoding center during early stages of SSU assembly together with METTL17 and RBFA (63). Conversely, these two proteins appeared more affected by EtBr (decreased by 5-and 4-fold, respectively) (Figure 9C).…”

“…The mitoribosome assembly is a complex process that consists of mt-rRNA synthesis and its processing, including tRNA cleavage, rRNA modification and folding; sequential recruitment of assembly factors and MRPs to mt-rRNAs; and joining of the subunits into the 55S monosome (76). The assembly of SSU and LSU has been extensively studied in recent years by analysis of cryo-EM structures at atomic resolution, which detailed multiple assembly intermediates (56- 58,63,77). In addition, the hierarchy of MRPs binding to mt-rRNAs was investigated in a pulse-chase stable isotope labeling by amino acids in cell culture (SILAC) study that described whether particular MRPs incorporated in the mitoribosomes at early or late stages of assembly (78).…”

Physiological variability in mitochondrial rRNA predisposes to metabolic syndrome

“…During the biogenesis of mtSSU, the variable part of h44 assumes its mature conformation only after dissociation of the assembly factor NOA1, which interferes with its position (Harper et al 2023). The polymorphisms might affect the process of h44 folding or play a role in an earlier, so far uncharacterized, phase of the assembly (Itoh et al 2022, Harper et al 2023). Together, these findings may indicate a possible interaction with yet unknown regulator, either during the translational cycle or during the mtSSU assembly.…”

“…The mt-Rnr1 polymorphisms identified in mtF344 are located in the variable part of the 3’ minor domain helix h44 (Figure 7C), a key decoding centre component. The maturation of h44 requires dissociation of the NOA1 assembly factor that otherwise hinders the interaction of h44 with the docking site (Harper et al 2023). Our data showing decreased levels of mtSSU proteins and attenuated mitochondrial protein synthesis are consistent with affected mtSSU biogenesis, likely resulting from the sequence variability in mt-Rnr1 .…”

“…Superposition of human, mouse and porcine mtSSU structures showed that the variable region (depicted in red in Figure 7D) adopts slightly different structures in the three species, and it is relatively loose and exposed to the mtSSU surface on the intersubunit side. During the biogenesis of mtSSU, the variable part of h44 assumes its mature conformation only after dissociation of the assembly factor NOA1, which interferes with its position (Harper et al 2023). The polymorphisms might affect the process of h44 folding or play a role in an earlier, so far uncharacterized, phase of the assembly (Itoh et al 2022, Harper et al 2023).…”