University of Cambridge
Elucidating the functions of RNA helicases DDX3X and DDX3Y in normal and malignant B cells
Abstract
dc:description.abstractDDX3X is an RNA helicase involved in multiple stages of RNA biogenesis, the most well-studied being as a regulator for translation. Its Y chromosome paralogue, DDX3Y shares 92% amino acid conservation. DDX3X is ubiquitously expressed and considered pan essential. While DDX3Y is widely transcribed, its protein expression is predominantly limited to spermatogonia cells or at least restricted to very low level in other cell types. DDX3X is frequently mutated or deleted in Burkitt lymphoma and in other forms of aggressive, MYC-driven lymphoma, which arise from germinal centre (GC) B cells. DDX3X mutations are especially enriched in male patients. In contrast, DDX3Y is never mutated in lymphoma. Previous work in our group showed DDX3X mutant male tumour cells were addicted to ectopic expression of DDX3Y – a phenomenon we term ‘switched paralogue dependency’. Precisely why tumour cells strive to delete DDX3X, only to upregulate an almost identical paralogue from the Y chromosome is an intriguing question that has not been fully explained. Therefore, the aim of this PhD study was to investigate the exclusive and redundant molecular functions of DDX3X and DDX3Y in B cell development and MYC-driven lymphomagenesis. Using CRISPR-edited, isogenic lymphoma cell lines, I found that DDX3Y could compensate for loss of DDX3X to maintain translation and viability and that DDX3X deficient cells were entirely dependent upon DDX3Y for survival. By epitope-tagging the endogenous loci of DDX3X and DDX3Y I performed comparative RNA and protein interactome studies using iCLIP and mass spectrometry. This revealed subtle differences in the pattern of RNA binding and interaction with the translation initiation machinery. Specifically, DDX3X showed preferential interaction with eIF3 subunits, suggesting possible differences in how DDX3X and DDX3Y regulate translation. Proteomic analysis following DDX3X deletion or degron-tag-induced DDX3Y degradation was used to reveal translational targets of DDX3X and DDX3Y, and highlighted the importance of total DDX3 protein abundance in regulating target expression. In parallel, I used conditional mouse models to reveal that DDX3 is required for GC formation and B cell survival. DDX3Y could compensate for DDX3X loss in terms of GC formation. However, DDX3Y alone was unable to support normal levels of class switch recombination and B cell proliferation. This phenotype was potentially explained by differences in total DDX3 dosage. Deletion of Ddx3x but not Ddx3y accelerated MYC-induced lymphomagenesis. Whole proteome and transcriptome analysis of tumours samples revealed changes in ribosome biogenesis and mitochondrial function potentially associated with reduction of MYC-driven apoptosis in early tumour cells. Overall, this study makes a comprehensive comparison between the functions of DDX3X and DDX3Y and highlights the importance of the dosage of total DDX3 in normal B cell function and in MYC-driven lymphomagenesis.
Degree
thesis:*- Name dc:type.qualificationname
- Doctor of Philosophy (PhD)
- Level dc:type.qualificationlevel
- Doctoral
- Grantor dc:publisher.institution
- University of Cambridge
- Year dc:date.issued
- 2025
Author and committee
dc:creator, dc:contributor.*- Author dc:creator
-
- Helian, Kaiyue
- Advisors dc:contributor.advisor
-
- Hodson, Daniel
- Gong, Chun
Subjects
dc:subject × 6Rights
dc:rightsIdentifiers
dc:identifier.*- DOI dc:identifier.doi
- https://doi.org/10.17863/CAM.123591
- OAI identifier oai:identifier
- oai:www.repository.cam.ac.uk:1810/393137