UPF3A and UPF3B Shape the Transcriptome Cooperatively Yet Oppose Cell Function

The nonsense mediated mRNA decay (NMD) pathway is a major regulator of gene expression that is essential for normal development and physiology. NMD is orchestrated by the activities of three core NMD proteins UPF1, UPF2 and UPF3B. While complete loss of function of UPF1 or UPF2 is embryonic lethal, loss of function of UPF3B is viable. In the absence of UPF3B, NMD is thought to be rescued by the redundant functions of the UPF3B paralog UPF3A. However, full redundancy of these paralogs is challenged by the embryonic lethality of UPF3A loss of function, and neurodevelopmental phenotypes observed in humans and mice lacking UPF3B. To compare the functions of the UPF3 paralogs we generated L-cell lines with different relative UPF3A and UPF3B abundances via knockdown and/or overexpression. RNA sequencing of UPF3 manipulated L-cells highlighted a major overlap of transcriptome changes following loss of UPF3A or UPF3B, yet overexpression of UPF3A could not rescue changes caused by loss of UPF3B. This suggested both genes are required for NMD in L-cells in a non-redundant manner. However, despite the general similarity of transcriptome changes, loss of UPF3B caused hyper-proliferation, while loss of UPF3A caused a hypo-proliferation of L-cells, and was associated with discordant activation of mTOR signalling. We observe similar opposing impacts of UPF3 manipulation in the context of neural progenitor cell proliferation and differentiation, and neuronal axon growth. These data suggest that while UPF3A and UPF3B generally act cooperatively to tune the transcriptome, their private or discordant functions can drive major biological impact.