Disease-linked mutation in RNase MRP inhibits ribosome synthesis
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Date
07/12/2021Author
Robertson, Nic
Metadata
Abstract
RMRP is a nuclear gene that encodes the RNA component of the endoribonuclease complex
RNase MRP. Mutations in human RMRP cause a spectrum of disorders characterised by
skeletal defects, hair anomalies and immunodeficiency. RNase MRP has an evolutionarily
conserved role cleaving pre-ribosomal RNA (rRNA), and several other functions have also
been proposed. However, why RMRP mutations cause disease is unknown. To look for
potential novel targets of RNase MRP, RNAs interacting with protein components of the
complex were mapped by the technique of cross-linking and analysis of cDNAs (CRAC).
This revealed interactions near the known cleavage site in yeast pre-rRNA. The same experiment in human cells unexpectedly only detected interactions with RNA components of the
complex, rather than potential targets. It has previously been shown that CRISPR-mediated
disruption of RMRP in immortalised cell lines causes a pre-rRNA processing defect. Here,
this result was confirmed in primary mouse T cells, which additionally showed impaired
proliferation when RMRP was non-specifically mutated. To answer whether disease-causing
mutations also cause this phenotype, a human cell line carrying the most common patient
mutation was generated with CRISPR. This showed a growth defect accompanied by an
accumulation of 41S precursor rRNA and reduced mature rRNA per cell. A similar rRNA
processing phenotype was also found in CHH patient fibroblasts. Finally, the RNA-bound
proteome of RMRP-mutant cells was mapped by the recently developed technique of Total
RNA-bound Protein Purification (TRAPP). This confirmed that mutant cells have reduced
cytosolic ribosomes relative to their mitochondrial ribosomal pool, and reduced intact RNase
MRP complexes. In summary, these results support the model that mutations in RMRP
cause disease by inhibiting ribosome synthesis.