'Single cell quantification of ribosome occupancy in early mouse development'

'''Technological limitations have precluded transcriptome-wide analyses of translation at single cell resolution. Here, we developed a novel microfluidic isotachophoresis approach, named RIBOsome profiling via IsoTachoPhoresis (Ribo-ITP), to simultaneously purify and size-select ribosome footprinted RNAs to solve this challenge. We applied this technique to characterize translation dynamics of single oocytes and single embryos from different stages of mouse preimplantation development. We identified differential translation efficiency as a key shared regulatory mechanism of genes involved in centrosome organization and N6-methyladenosine modification of RNAs. We also discovered stage- and allelic-specific differential engagement of zygotic RNAs with ribosomes. Finally, by integrating our measurements with proteomics data, we discovered that ribosome occupancy in germinal vesicle stage oocytes is the predominant determinant of protein abundance in the zygote. Furthermore, we find that ribosome occupancy is dramatically remodeled at the 4 and 8-cell stage and strongly correlates with the proteome of the morula stage embryos. These findings resolve the long-standing paradox of low correlation between RNA expression and protein abundance in early embryonic development. The novel Ribo-ITP approach will enable numerous applications by providing high coverage and high resolution ribosome occupancy measurements from ultra-low input samples including single cells. To study translation in mouse development, we performed our novel low-input ribosome profiling method (named Ribo-ITP) and RNA sequencing (Smart-seq3) in mouse oocytes and early stage embryos. To validate our method, we performed Ribo-ITP and conventional ribosome profiling using human K562 cells.'''