Types and Functions of Mitoribosome-Specific Ribosomal Proteins across Eukaryotes

Abstract

Mitochondria are key organelles that combine features inherited from their bacterial endosymbiotic ancestor with traits that arose during eukaryote evolution. These energy producing organelles have retained a genome and fully functional gene expression machineries including specific ribosomes. Recent advances in cryo-electron microscopy have enabled the characterization of a fast-growing number of the low abundant membrane-bound mitochondrial ribosomes. Surprisingly, mitoribosomes were found to be extremely diverse both in terms of structure and composition. Still, all of them drastically increased their number of ribosomal proteins. Interestingly, among the more than 130 novel ribosomal proteins identified to date in mitochondria, most of them are composed of a-helices. Many of them belong to the nuclear encoded super family of helical repeat proteins. Here we review the diversity of functions and the mode of action held by the novel mitoribosome proteins and discuss why these proteins that share similar helical folds were independently recruited by mitoribosomes during evolution in independent eukaryote clades.

Keywords: helical repeat proteins; mitochondrial gene expression; pentatricopeptide repeat proteins; ribosomes; single particle cryo-EM; translation.

Figure 1

Structural comparison between available mitoribosome structures. To highlight the divergence of mitoribosomes as compared to bacteria and their diversity in terms of size and shape across eukaryotes, high resolution structures of mitoribosomes are presented and overlaid on a schematic evolutionary tree of eukaryotes. From left to right, mitoribosomes structures are from: ciliate (T. thermophilus), green alga (C. reinhardtii), flowering plant (A. thaliana), kinetoplasts (T. brucei), fungi (N. crassa) and mammals (H. sapiens). They are compared with E. coli ribosome structure, representing the ancestral form of mitoribosomes. Large subunits components are shown in blue shades and small subunit components in yellow shades. Mitochondria-specific ribosomal proteins (which includes shared and species-specific proteins) are highlighted in red. No structures have been solved yet for the supergroup Amoebozoa (hence not shown on the figure). SAR corresponds to the supergroup which includes the Stramenopiles, Alveolates, and Rhizaria subgroups.

Figure 2

Graphical summary of the functions of mitoribosome-specific ribosomal proteins. The recently identified proteins specifically occurring in mitochondrial ribosomes are shown in red. They can be involved in rRNA stabilization, in the recruitment of mRNA for translation initiation, in the assembly and maturation of mitoribosomes, and in its attachment to the mitochondrial inner membrane through the binding with the insertase Oxa1. The exemplary shown mS39 and mL45 specifically occur in mammalian mitoribosomes.

Figure 3

Examples of RNA-binding proteins recruited to mitoribosomes. (A,B) present close-up views of large alpha-helical proteins which were largely recruited to mitoribosomes. (A) The OPR mL115 involved in the stabilization of fragment L1 of the fragmented rRNAs of C. reinhardtii is shown. The OPR enlaces the 3′ single-stranded extremity of the L1 fragment which is stabilized via charge interaction. It also interacts with H19 and H4. The electrostatic potential of mL115 is shown with blue surfaces showing positive charges and red surfaces showing negative charges. (B) The PPR mL104 of the flowering plant mitoribosome is shown. It englobes helix H10, interacting with the rRNA backbone which is a different mode of action compared to non-ribosomal PPR proteins. It also interacts with H-p59, a plant specific RNA helix. As in (A), the electrostatic potential of mL115 is shown, revealing that the rRNA interaction is mostly mediated by positively charged residues. (C) The pseudo-dimer composed of mL65 and mL37, probably repurposed endonucleases, are shown as an example of non-alpha-helical mitoribosome-specific r-proteins. The r-proteins interact with the reduced domain III of the large subunit of the mammalian mitoribosome. (D) Structural comparison of the foot of the small subunit. This comparison highlights the conserved helical proteins involved in the stabilization of the foot of the small subunit—where the tip of h44 and h6 are either extended, reduced, or deleted—most likely due to the loss of the bacterial r-protein bS20 in mitoribosomes. All mitoribosomes share a PPR protein at this position (mS27, mS90, rPPR* and mS106), which may be the product of a single ancestral PPR protein. In trypanosoma, PPR protein mS63 is also present at this position but no longer interacts via RNA. The different parts of the small subunit are indicated on the bacterial model; h: head, b: body, f: foot, p: platform.

Figure 4

Specific proteins involved in the translation initiation process in mitochondria. Proposed mechanisms of mitochondria-specific translation initiation events involving mitochondria-specific r-proteins are shown. (A) The proposed positions and orientations of the mRNA on the small subunits of mammalian, plant and bacteria during translation initiation are indicated on the figure. The mRNA is shown in blue, the initiation codon (AUG) is in purple and the Shine-Dalgarno (SD) consensus, only present in bacteria, is in green. (B,C) present close-up views of mammalian and plant mRNA binders. (B) The mammalian mRNA stabilization by the PPR protein mS39 and probably the LRPPRC-SLIRP complex is shown. mS39 and LRPPRC-SLIRP interact with the ORF, downstream of the AUG. (C) In flowering plants, a proposed mechanism of mRNA stabilization would involve the PPR protein mS83, which would bind a consensus sequence located in the 5′ UTR of the mitochondrial mRNAs. A similar process was also proposed in yeast, involving specific proteins for each yeast mRNA.