Dissecting mechanisms of translational regulation by specialised ribosomes

Ribosomes are fundamental cellular machines responsible for protein synthesis. Recent evidence suggests that rather than being homogenous, the protein and RNA composition of ribosomes can vary, which consequently can lead to specialisation of function. Several examples have been characterised in which ribosomes of a particular composition preferentially translate a specific pool of mRNAs or specific type of open reading frame. Mutations in ribosomal components lead to a number of diseases, termed ribosomopathies, so it is important to understand the molecular basis of regulation by ribosomes.

We identified specialised ribosomes induced during Kaposi’s sarcoma-associated herpesvirus (KSHV) infection (Murphy et al Nature Coms, 2023) and during spermatogenesis in Drosophila (Hopes et al, NAR, 2022). During KSHV infection, host ribosomes are altered in composition which results in changes to their ability to translate small open reading frames , that regulate viral proteins. However, the precise mechanisms of how this translational regulation takes place is still missing. How do these compositional variations in the ribosome result in changes in mRNA or open reading frame selection?

Objectives & experimental approaches:

1) Determine features of mRNAs and open reading frames preferentially translated-Analyse previously generated ribosome profiling data for specialised ribosomes in KSHV and Drosophila.

2) Dissect structural implications of changes in ribosome composition-Biochemically isolate ribosome populations and determine structural differences by cryo-EM.

3) Dissect precise mechanism of how composition affects mRNA/ORF selectivity-Test hypotheses generated from previous aims in translation reporter systems.

Outcomes:

This project work will enable us to understand how changes in ribosomes composition, result in changes to ribosomal structure and how this impacts on the ribosomes ability to select mRNAs or open reading frame to translate. This will provide insight that could enable us to design new therapeutic targets for KSHV or manipulate the ribosome for biotechnological benefit.