The role of NSUN2 in tRNA derived small RNA biogenesis and nascent RNA silencing
Lead Research Organisation:
University of Oxford
Department Name: Sir William Dunn Sch of Pathology
Abstract
All organisms consist of cells that multiply through cell division. The genetic code, which defines each cell type, is stored
within DNA molecules. DNA contains coding regions called genes. Genes are transcribed into messenger RNA (mRNA)
molecules, which are in turn translated into proteins, a process called gene expression. It is important that each cell makes
the right levels of proteins, which is achieved by controlled gene expression. Uncontrolled gene expression can lead to
cancer or cell death. One of the most important mechanisms for the regulation of gene expression is gene silencing. Gene
silencing in human cells requires the presence of small RNA molecules (siRNA) that lead to recognition and destruction of
target mRNA in the cytoplasm. Another type of gene silencing is the establishment of specific modifications on DNA, which
leads to inhibition of gene transcription, also called transcriptional gene silencing (TGS).
Pathway that leads to gene silencing is called RNA interference (RNAi) and this great discovery was recognised by Nobel
prize in 2006. RNAi was widely explored not only as an important scientific tool, but also as novel therapeutic agent. With
very first FDA approved drug, Patisiran, sRNA-based therapeutics are now receiving more and more attention.
We have discovered novel gene silencing pathway that is very distinct from well-known PTGS or TGS. We found that tRNA
molecules that are essential for protein production, can fold into various alternative shapes. Such non-canonical structures
are recognised by Dicer, enzyme that can cleave them into tRNA-derived small RNA molecules, called tsRNAs. tsRNAs
are not just degradation products of misfolded tRNAs, but they are functional and suppress expression of many disease
driving genes. It is very important to understand how is the balance between proper tRNA structures (clover leaf) and
alternative structures regulated. We propose that modifying enzyme, NSUN2, can determine the folding
abilities of tRNAs and consequently their recognition and cleavage by Dicer. Furthermore, we hypothesise that
modifications on tsRNAs can have direct impact on efficiency of nascent RNA silencing. This proposal has direct
translational potential in identifying key features of novel tsRNA-based therapeutics.
within DNA molecules. DNA contains coding regions called genes. Genes are transcribed into messenger RNA (mRNA)
molecules, which are in turn translated into proteins, a process called gene expression. It is important that each cell makes
the right levels of proteins, which is achieved by controlled gene expression. Uncontrolled gene expression can lead to
cancer or cell death. One of the most important mechanisms for the regulation of gene expression is gene silencing. Gene
silencing in human cells requires the presence of small RNA molecules (siRNA) that lead to recognition and destruction of
target mRNA in the cytoplasm. Another type of gene silencing is the establishment of specific modifications on DNA, which
leads to inhibition of gene transcription, also called transcriptional gene silencing (TGS).
Pathway that leads to gene silencing is called RNA interference (RNAi) and this great discovery was recognised by Nobel
prize in 2006. RNAi was widely explored not only as an important scientific tool, but also as novel therapeutic agent. With
very first FDA approved drug, Patisiran, sRNA-based therapeutics are now receiving more and more attention.
We have discovered novel gene silencing pathway that is very distinct from well-known PTGS or TGS. We found that tRNA
molecules that are essential for protein production, can fold into various alternative shapes. Such non-canonical structures
are recognised by Dicer, enzyme that can cleave them into tRNA-derived small RNA molecules, called tsRNAs. tsRNAs
are not just degradation products of misfolded tRNAs, but they are functional and suppress expression of many disease
driving genes. It is very important to understand how is the balance between proper tRNA structures (clover leaf) and
alternative structures regulated. We propose that modifying enzyme, NSUN2, can determine the folding
abilities of tRNAs and consequently their recognition and cleavage by Dicer. Furthermore, we hypothesise that
modifications on tsRNAs can have direct impact on efficiency of nascent RNA silencing. This proposal has direct
translational potential in identifying key features of novel tsRNA-based therapeutics.
Technical Summary
Gene expression regulation is fundamental process, essential for life of every cell. Gene silencing is one of the key
regulatory pathways. In mammalian cells, small non-coding RNAs (sncRNAs) negatively regulate gene expression in a
pathway known as RNA interference (RNAi). RNAi can be categorised into post-transcriptional gene silencing (PTGS),
which involves the cleavage of target messenger RNA (mRNA) or inhibition of translation in the cytoplasm, and
transcriptional gene silencing (TGS), which is mediated by the establishment of repressive epigenetic marks at target loci.
Transfer RNAs (tRNAs), which are essential for translation, can be processed into small ncRNAs, termed tRNA-derived
small RNAs (tsRNAs). The biogenesis of tsRNAs and their role in gene expression regulation has not yet been fully
understood.
The acquisition of a set of chemical modifications is an essential for production of functional tRNA molecules. More than
hundreds of tRNA modifications, ranging from simple methylation to complex hyper-modifications, are dynamically deposited and have different roles in controlling stability, folding and decoding properties of tRNA molecules. Impaired
deposition of modifications onto tRNA molecules, hindering the correct cloverleaf folding and increasing the presence of
alternative structures, can lead to Dicer cleavage and tsRNAs production.
We have recently discovered novel tsRNA mediated gene silencing pathway that suppresses expression of many disease
driving genes. This discovery led to international patent application and further funding for in vivo validation.
Therefore, it is crucial that we understand molecular mechanisms underlying tsRNA biogenesis and their function in
nascent RNA silencing. In particular how tRNA modifying enzyme NSUN2 can affect their alternative structures and how this might
lead to tsRNA production. We are also interested in modifications on tsRNAs and how these contribute to tsRNAs mediated
gene silencing.
regulatory pathways. In mammalian cells, small non-coding RNAs (sncRNAs) negatively regulate gene expression in a
pathway known as RNA interference (RNAi). RNAi can be categorised into post-transcriptional gene silencing (PTGS),
which involves the cleavage of target messenger RNA (mRNA) or inhibition of translation in the cytoplasm, and
transcriptional gene silencing (TGS), which is mediated by the establishment of repressive epigenetic marks at target loci.
Transfer RNAs (tRNAs), which are essential for translation, can be processed into small ncRNAs, termed tRNA-derived
small RNAs (tsRNAs). The biogenesis of tsRNAs and their role in gene expression regulation has not yet been fully
understood.
The acquisition of a set of chemical modifications is an essential for production of functional tRNA molecules. More than
hundreds of tRNA modifications, ranging from simple methylation to complex hyper-modifications, are dynamically deposited and have different roles in controlling stability, folding and decoding properties of tRNA molecules. Impaired
deposition of modifications onto tRNA molecules, hindering the correct cloverleaf folding and increasing the presence of
alternative structures, can lead to Dicer cleavage and tsRNAs production.
We have recently discovered novel tsRNA mediated gene silencing pathway that suppresses expression of many disease
driving genes. This discovery led to international patent application and further funding for in vivo validation.
Therefore, it is crucial that we understand molecular mechanisms underlying tsRNA biogenesis and their function in
nascent RNA silencing. In particular how tRNA modifying enzyme NSUN2 can affect their alternative structures and how this might
lead to tsRNA production. We are also interested in modifications on tsRNAs and how these contribute to tsRNAs mediated
gene silencing.