Loss of function mutations in GEMIN5 cause a neurodevelopmental disorder.

GEMIN5, an RNA-binding protein is essential for assembly of the survival motor neuron (SMN) protein complex and facilitates the formation of small nuclear ribonucleoproteins (snRNPs), the building blocks of spliceosomes. Here, we have identified 30 affected individuals from 22 unrelated families presenting with developmental delay, hypotonia, and cerebellar ataxia harboring biallelic variants in the GEMIN5 gene. Mutations in GEMIN5 perturb the subcellular distribution, stability, and expression of GEMIN5 protein and its interacting partners in patient iPSC-derived neurons, suggesting a potential loss-of-function mechanism. GEMIN5 mutations result in disruption of snRNP complex assembly formation in patient iPSC neurons. Furthermore, knock down of rigor mortis, the fly homolog of human GEMIN5, leads to developmental defects, motor dysfunction, and a reduced lifespan. Interestingly, we observed that GEMIN5 variants disrupt a distinct set of transcripts and pathways as compared to SMA patient neurons, suggesting different molecular pathomechanisms. These findings collectively provide evidence that pathogenic variants in GEMIN5 perturb physiological functions and result in a neurodevelopmental delay and ataxia syndrome

The invariant uridine of stop codons contacts the conserved NIKSR loop of human eRF1 in the ribosome

To unravel the region of human eukaryotic release factor 1 (eRF1) that is close to stop codons within the ribosome, we used mRNAs containing a single photoactivatable 4-thiouridine (s(4)U) residue in the first position of stop or control sense codons. Accurate phasing of these mRNAs onto the ribosome was achieved by the addition of tRNA(Asp). Under these conditions, eRF1 was shown to crosslink exclusively to mRNAs containing a stop or s(4)UGG codon. A procedure that yielded (32)P-labeled eRF1 deprived of the mRNA chain was developed; analysis of the labeled peptides generated after specific cleavage of both wild-type and mutant eRF1s maps the crosslink in the tripeptide KSR (positions 63–65 of human eRF1) and points to K63 located in the conserved NIKS loop as the main crosslinking site. These data directly show the interaction of the N-terminal (N) domain of eRF1 with stop codons within the 40S ribosomal subunit and provide strong support for the positioning of the eRF1 middle (M) domain on the 60S subunit. Thus, the N and M domains mimic the tRNA anticodon and acceptor arms, respectively

X-ray crystallography study on ribosome recycling: the mechanism of binding and action of RRF on the 50S ribosomal subunit

This study presents the crystal structure of domain I of the Escherichia coli ribosome recycling factor (RRF) bound to the Deinococcus radiodurans 50S subunit. The orientation of RRF is consistent with the position determined on a 70S-RRF complex by cryoelectron microscopy (cryo-EM). Alignment, however, requires a rotation of 7° and a shift of the cryo-EM RRF by a complete turn of an α-helix, redefining the contacts established with ribosomal components. At 3.3 Å resolution, RRF is seen to interact exclusively with ribosomal elements associated with tRNA binding and/or translocation. Furthermore, these results now provide a high-resolution structural description of the conformational changes that were suspected to occur on the 70S-RRF complex, which has implications for the synergistic action of RRF with elongation factor G (EF-G). Specifically, the tip of the universal bridge element H69 is shifted by 20 Å toward h44 of the 30S subunit, suggesting that RRF primes the intersubunit bridge B2a for the action of EF-G. Collectively, our data enable a model to be proposed for the dual action of EF-G and RRF during ribosome recycling