Structures and stabilization of kinetoplastid-specific split rRNAs revealed by comparing leishmanial and human ribosomes

Abstract

The recent success in ribosome structure determination by cryoEM has opened the door to defining structural differences between ribosomes of pathogenic organisms and humans and to understand ribosome-targeting antibiotics. Here, by direct electron-counting cryoEM, we have determined the structures of the Leishmania donovani and human ribosomes at 2.9 Å and 3.6 Å, respectively. Our structure of the leishmanial ribosome elucidates the organization of the six fragments of its large subunit rRNA (as opposed to a single 28S rRNA in most eukaryotes, including humans) and reveals atomic details of a unique 20 amino acid extension of the uL13 protein that pins down the ends of three of the rRNA fragments. The structure also fashions many large rRNA expansion segments. Direct comparison of our human and leishmanial ribosome structures at the decoding A-site sheds light on how the bacterial ribosome-targeting drug paromomycin selectively inhibits the eukaryotic L. donovani, but not human, ribosome.

Daptomycin, a last-resort antibiotic, binds ribosomal protein S19 in humans

“…Recently the high resolution total structure of the human ribosome has been published and shows RPS19 to be located on the head of the 40S subunit, extending well into the functional center of the 40S subunit [23]. The protein is exposed (Fig.…”

The cryo-EM Structure of a Novel 40S Kinetoplastid-Specific Ribosomal Protein

“…In the 40S subunits of kinetoplastids, ES3 S , ES6 S , ES7 S , and ES9 S are substantially larger when compared with other known eukaryotic counterparts (Hashem et al, 2013). The secondary structures of these divergent rRNA ESs reveal that they could enclose various thermodynamically unstable domains, such as single-stranded RNA and large internal loops, possibly illustrating the need for more protein elements to stabilize these different rRNA domains (Hashem et al, 2013;Melnikov et al, 2015;Zhang et al, 2016). Consequently, due either to gene evolutionary shuffling or gene expansions, several kinetoplastid r-proteins have evolved larger N-or C-terminal tails (Ayub et al, 2009;Hashem et al, 2013;Zhang et al, 2016).…”

“…Hence, the main challenge for the specific targeting of the function or stability of the kinetoplastid ribosome is to uncover its kinetoplastid-specific structural features. Recently, intermediate to high-resolution structures of several kinetoplastidian ribosomes (Hashem et al, 2013;Liu et al, 2016;Shalev-Benami et al, 2016;Zhang et al, 2016) were determined by cryoelectron microscopy (cryo-EM) and pinpointed unique structural features when compared with other known eukaryotic ribosomes, such as large rRNA expansion segments (ES), as well as r-protein extensions. These cryo-EM studies led to the structural description of nearly all the rRNA chains, including most of the kinetoplastid-specific ESs and domains, as well as the assignment of all known eukaryotic ribosomal proteins (r-proteins).…”

“…The secondary structures of these divergent rRNA ESs reveal that they could enclose various thermodynamically unstable domains, such as single-stranded RNA and large internal loops, possibly illustrating the need for more protein elements to stabilize these different rRNA domains (Hashem et al, 2013;Melnikov et al, 2015;Zhang et al, 2016). Consequently, due either to gene evolutionary shuffling or gene expansions, several kinetoplastid r-proteins have evolved larger N-or C-terminal tails (Ayub et al, 2009;Hashem et al, 2013;Zhang et al, 2016). This genetic divergence within the r-protein terminal regions has resulted in local structural rearrangements that guide and stabilize the conformation of some regions of these extended rRNA ESs (Hashem et al, 2013).…”

G protein‐dependent signaling triggers a β‐arrestin‐scaffolded p70S6K/ rpS6 module that controls 5'TOP mRNA translation

“…By quantitative confocal microscopy, we show that p70S6K and rpS6 colocalize to a lesser extent in MEFs from b-arrestin 1,2 knockout mice. The crystal structure of the human ribosome (5T2C in the Protein DataBase) (80) suggests that the S235/S236 residues are protruding and thus are accessible to extraribosomal kinase. In our model of the b-arrestin/p70S6K complex, the orientation of the p70S6K makes it largely accessible to substrates and compatible with phosphorylation of the carboxyterminal regulatory sites on rpS6 within the ribosome (data not shown).…”