Development of New Proteomics Technology and its Application to Study Cellular Organization

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Deciphering the Relationship Between mRNA and Protein Dynamics in a Developing Proto-Vertebrate The broad goal of the parent award (Development of New Proteomics Technology and its Application to Study Cellular Organization) is to advance quantitative proteomics technology and apply it to obtain a systems level understanding of cellular organization. Recently, we have demonstrated that TMTproC significantly increases sensitivity and multiplexing capacity of the complement reporter ion approach while making these approaches compatible with a broad set of mass spectrometers (Johnson, 2021). We have also worked to improve access to high- quality proteomics data by integrating TMTproC with ion-trap only instruments which cost 80% less than ones typically used for multiplexed quantitative proteomics. We have successfully leveraged our proteomics technology development for multiple collaborations (Crapse, 2021; Hart, 2020; Cao, 2020) which has advanced our understanding of cellular organization. One focus of the lab is to better understand developmental progression (Nguyen, 2021; Keber, 2021; Nofal, 2021). Recently, we have published a preprint showing that, in frog embryos, nuclear proteins access the nucleus in a temporal order set via importin affinities during development, thereby acting as an elegant timing mechanism to control the onset of their downstream functions (Nguyen, 2021). Extending beyond our work in developmental biology, we have developed methods that allow us to parameterize all aspects of protein abundance control on a global scale through multi-omics integration of transcription, translation, and degradation. Key to these advances was developing new methods that allow measurement of perturbation-free protein turnover. So far, we have applied these methods to E. coli and yeast (Gupta, et al., in preparation), and Alex Frese is expanding these approaches to the developing Ciona embryo with the goal of generalizing these techniques to complex higher organisms such as Xenopus and Drosophila.