Identifying novel SNORD116 targets and signaling pathways
- Supervisors: Prof. Mandy Peffers
SNORD116 is a cluster of small nucleolar RNAs absent in Prader-Willi syndrome (PWS), with undefined molecular actions and no cure. PWS affects the neurological, musculoskeletal and cardiac systems. This project builds on our data to identify and confirm direct SNORD116 targets, interactions, and signaling pathways.
Our pilot data exploited differences in species (human versus mouse), tissue (cardiomyocytes versus chondrocytes), and model systems (overexpression (OE) versus knockout (KO)), identifying commonly conserved signaling pathways; PI3K/mTOR, and potentially the first direct SNORD116 target; plakophilin 1-PKP1. This project will validate our data in musculoskeletal,cardiac and neuronal contexts. Defining these targets and pathways could elucidate the molecular role of SNORD116 leading to improved therapies.
We also identified SNORD116 signaling in other disease settings including elevated exosomal SNORD116 in human pluripotent stem (hiPSC) cell models of hypertrophic cardiomyopathy, and human/mouse osteoarthritis cartilage. These data indicate conserved developmental SNORD116 signaling, disrupted in PWS contributes to additional age-related diseases. We compared proteomic analysis of hiPSC-derived cardiomyocytes OE SNORD116, with chondrocytes from SNORD116 KO mice identifying desmosomal signalling proteins and PI3K/mTOR pathways as disrupted. Of the commonly affected proteins one followed the biological relevant trend of potential SNORD116 target (up KO/down OE); desmosomal signalling protein PKP1, which regulates the PI3K/mTOR pathways. This project will confirm this experimentally, in chondrocyte, cardiomyocytes and neuronal development.
Objectives and experimental approach
We hypothesise SNORD116 loss in PWS disrupts desmosomal signaling during development as PKP1 is a SNORD116 target. Aims;
- Define desmosomal protein expression profiles during chondrocyte/cardiomyocyte/neuronal development.
- Investigate SNORD116 role in regulating this expression profile.
mRNA (qRT-PCR) and protein (immunocytochemistry) will be determined at daily timepoints in hiPSCs differentiation to chondrocyte/cardiomyocyte/neural progenitors.
Wild-type and SNORD116 KO hiPSCs will be differentiated to key chondrocyte/cardiomyocyte/neuronal timepoints and protein profile changes determined with label-free mass spectrometry and bioinformatics.
- Determine if SNORD116 directly binds specific desmosomal proteins and if this facilitates RNA modifications.
Interaction between PKP1 and SNORD116 will be determined using SHAPE-MaP for in-cell probing of intermolecular RNA interactions.
Open to students worldwide
This PhD is for a self-funded PhD student only. The successful applicant must provide all funds for tuition, bench fees and living expenses.