Exploration Of The Potential Roles Of M~6A Modification In MRNA Transport And Localization To Neuronal Axons

m~6A modification is the most popular chemical modification in mRNAs.It plays an important role with its regulatory proteins: writers,erasers,and readers.Its dynamic regulation process can affect cells proliferation and differentiation,nervous system development,human diseases,local translation of mRNAs.Recently,m~6 A modification is being studied in various fields because of its widespread presence in mRNAs.Previous studies have shown that loss of function of FTO in axons can lead to the higher level of m~6 A on mRNAs,which leads to the translation of mRNAs in axons suppressed.Before local translation of mRNAs in axons,mRNAs should be transported into axons firstly.It is unclear whether m~6 A modifications can participate in regulating the transport and localization of mRNAs through their readers.Neuron,as a highly polarized structure,is an ideal model for studying the transport and localization of mRNAs in axons.Our study establishes an assay for collecting axonal RNAs,a system for knocking down m~6 A modification-related genes in vitro.We will use neuron isolation and culture technology,RNA collection and sequencing technology,fluorescence in situ hybridization(FISH),etc.We combine microfluidic technology and neuron culture technology to achieve the purpose of separating neuron cell bodies and axons.By improving the parameters of the microfluidic chips or using innovative microfluidic chip,we can improve the collection efficiency of axonal RNAs.We will knock down the expression of m~6A-related genes by transfecting neurons with lenti-virus in vitro,the somal and axonal RNAs of the control group and experimental group will be collected.Next,we will analyze those m RNA-seq data,and find some mRNAs localization regulated by m~6 A modification.Studying the regulation mechanism of m RNA m~6 A modification on m RNA transport and localization in axons plays an important role in understanding the function of m~6 A modification in nervous system development,which is also conducive to further research on the maturation and polarization process of neurons,especially in axonal subcellular structure.