Function follows (iso)form: Exploring functional diversity of isoform expression in single cells

单细胞基因表达分析具有解决构成复杂生物系统的多种细胞类型的能力。大多数基因都有可能被选择性地拼接成一个以上的同种异构体，同一基因的不同同种异构体编码具有不同功能的不同蛋白质。我们假设这种异构体表达的多样性，在大多数单细胞分析中是看不见的，是单个细胞变得功能异质的一种方式。该项目将探索小鼠造血系统中异构体表达的多样性，目的是识别功能异构体及其在单个细胞中的表达模式。利用一系列技术，包括FACS，单细胞基因组学，下一代短读和长读测序和生物信息学分析，该项目将专注于识别单个血液干细胞和祖细胞中功能相关的异构体表达。然后，通过基因组编辑去除外显子并迫使细胞表达特定的同种异构体，研究同种异构体表达的功能影响，然后通过基因表达、体外和体内试验监测细胞的反应。

Single-cell gene expression analysis has the power to resolve the multitude of cell types that comprise a complex biological system. The majority of genes have the potential to be alternatively spliced into more than one isoform, with different isoforms of the same gene encoding distinct proteins with distinct functions. We hypothesise that this diversity in isoform expression, unseen in most single-cell analyses, is one way that individual cells can become functionally heterogeneous.This project will explore the diversity of isoform expression in the mouse haematopoietic system, with the aim of identifying functionally heterogeneous isoforms and their pattern of expression within individual cells. Using a range of techniques, including FACS, single-cell genomics, next generation short- and long- read sequencing and bioinformatic analysis, the project will focus on the identification of functionally consequential isoform expression within individual blood stem and progenitor cells. The functional effect of the isoform expression will then be studied through genome editing to remove exons and force cells to express specific sets of isoforms, then monitoring the cell's response through gene expression, in vitro and in vivo assays.