Decoding CD4+ T cell activation by multi-modal phenotyping

The activation of CD4+ T cells is strongly implicated in the biology of immune-mediated diseases, yet the molecular and functional mechanisms governing this process remain incompletely understood. With the explosion of profiling and functional genomic methods in the last decade, it is now feasible to interrogate the more intricate phenotypes of activation and the increasingly long list of disease-associated genes. While transcriptomics has largely dominated these advancements and has significantly transformed the field by providing valuable insights, it is essential to also consider other modalities. This thesis aims to delineate the mechanisms that underlie T cell activation and their genetic regulation with a focus on two functional read-outs: transcriptomics and image-based profiling. In the first chapter, I optimise a protocol for pooled CRISPR-KO screens in primary CD4+ T cells with single-cell RNA sequencing as a read-out, and implement it to explore the role of 20 immune-disease genes on T cell activation. The results of this chapter will be instrumental for the design of larger CRISPR screens as well as provide a clue to the genetic regulation of T cell activation. In the second chapter, I outline the development of TGlow: a high-throughput high-content image-based method to profile morphology, metabolic, proteomic and functional features of T cell activation. In the third chapter, I implement TGlow to characterise CD4+ T cell activation with high temporal resolution in naïve and memory populations. The results of this chapter are the first of its kind morphological reference/map of T cell activation. The thesis concludes by reflecting on the challenges and future opportunities in functional genomics for advancing our understanding of CD4+ T cell activation, emphasizing the critical need to bridge genetics and disease biology.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2024

Author dc:creator

• Matte, Julie

Advisor dc:contributor.advisor

• Trynka, Gosia

• T cell activation
• high-content imaging
• immune-mediated diseases
• single-cell CRISPR screen
• variant to function

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/553c6eed-fdf7-4e7b-86b6-219ba5a1725a/download
• http://purl.org/NET/rdflicense/allrightsreserved

Language dc:language

eng

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.117304

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/382537