CASC3 promotes transcriptome-wide activation of nonsense-mediated decay by the exon junction complex

Gerbracht, Jennifer V.; Boehm, Volker; Britto‐Borges, Thiago; Kallabis, Sebastian; Wiederstein, Janica L.; Ciriello, Simona; Aschemeier, Dominik; Krüger, Marcus; Frese, Christian K.; Altmüller, Janine; Dieterich, Christoph; Gehring, Niels H.

doi:10.1093/nar/gkaa564

Cited by 38 publications

(39 citation statements)

References 97 publications

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“…Only when we deplete in UPF3A or UPF3B KO cell lines the respective other protein by RNAi or genomic KO, NMD efficiency substantially decreases. It should be noted that the effects of a KO on NMD activity are typically stronger than those of a KD, which is consistent with our earlier observations (Boehm et al, 2021;Gerbracht et al, 2020 (Yi et al, 2021).…”

Section: Discussionsupporting

confidence: 92%

UPF3A and UPF3B are redundant and modular activators of nonsense-mediated mRNA decay in human cells

Wallmeroth

Boehm

Lackmann

et al. 2021

Preprint

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The paralogous human proteins UPF3A and UPF3B are involved in recognizing mRNAs targeted by nonsense-mediated mRNA decay (NMD). While UPF3B has been demonstrated to support NMD, contradicting reports describe UPF3A either as an NMD activator or inhibitor. Here, we present a comprehensive functional analysis of UPF3A and UPF3B in human cells using combinatory experimental approaches. Overexpression or knockout of UPF3A as well as knockout of UPF3B did not detectably change global NMD activity. In contrast, the co-depletion of UPF3A and UPF3B resulted in a marked NMD inhibition and a transcriptome-wide upregulation of NMD substrates, demonstrating a functional redundancy between both NMD factors. Although current models assume that UPF3 bridges NMD-activating exon-junction complexes (EJC) to the NMD factor UPF2, UPF3B exhibited normal NMD activity in rescue experiments when UPF2 or EJC binding was impaired. Further rescue experiments revealed partially redundant functions of UPF3B domains in supporting NMD, involving both UPF2 and EJC interaction sites and the central region of UPF3. Collectively, UPF3A and UPF3B serve as fault-tolerant NMD activators in human cells.

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Section: Discussionsupporting

confidence: 92%

UPF3A and UPF3B are redundant and modular activators of nonsense-mediated mRNA decay in human cells

Wallmeroth

Boehm

Lackmann

et al. 2021

Preprint

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“…Mass spectrometry of RNPS1 and CASC3 containing EJCs showed that CASC3 but not RNPS1 preferentially associates with UPF3B (Mabin et al, 2018). Consistent with this observation, a recent report found a much-reduced EJC-UPF3B association in CASC3 knockout HEK293 cells (Gerbracht et al, 2020). Together, these observations suggest a link between EJC composition and UPF3B-mediated NMD.…”

Section: Introductionsupporting

confidence: 75%

“…We next tested the contribution of other factors from the EJC-mediated NMD pathway to UPF3 function in this branch of NMD. We first focused on CASC3-containing EJC as previous observations from human embryonic kidney (HEK293) cells suggest that there exists a functional synergy between the CASC3-containing EJC and UPF3B (Gerbracht et al, 2020; Mabin et al, 2018). To test if CASC3-UPF3B preferential association is also present in other cell types, we overexpressed in HeLa cells either wild-type CASC3 or a mutant CASC3 (HDAA) that is unable to associate with the EJC core (Ballut et al, 2005).…”

Section: Resultsmentioning

confidence: 99%

Mammalian UPF3A and UPF3B activate NMD independently of their EJC binding

Yi¹,

Arvola²,

Myers

et al. 2021

Preprint

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Nonsense-mediated mRNA decay (NMD) is governed by the three conserved factors - UPF1, UPF2 and UPF3. While all three are required for NMD in yeast, UPF3B is dispensable for NMD in mammals, with its paralog UPF3A suggested to only weakly activate or even repress NMD due to its weaker binding to the exon junction complex (EJC). Here we characterize the UPF3B-dependent and -independent NMD in human cell lines knocked-out of one or both UPF3 paralogs. We show that in human colorectal cancer HCT116 cells, EJC-mediated NMD can operate in UPF3B-dependent and -independent manner. While UPF3A is almost completely dispensable for NMD in wild-type cells, it strongly activates EJC-mediated NMD in cells lacking UPF3B. Surprisingly, this major NMD branch can operate in UPF3-independent manner questioning the idea that UPF3 is needed to bridge UPF proteins to the EJC during NMD. Complementation studies in UPF3 knockout cells further show that EJC-binding domain of UPF3 paralogs is not essential for NMD. Instead, the conserved mid domain of UPF3B, previously shown to engage with ribosome release factors, is required for its full NMD activity. Altogether, UPF3 plays a more active role in NMD than simply being a bridge between the EJC and the UPF complex.

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“…A fourth core subunit, known as Barentsz (Btz), Cancer susceptibility candidate gene 3 (CASC3), or Metastatic lymph node 51 (MLN51), associates with the complex following the completion of splicing, and is required for the effects of the EJC on translation, NMD and mRNA localization (Chazal et al, 2013; Palacios et al, 2004; Shibuya et al, 2006; van Eeden et al, 2001). The transition between RNPS1 and CASC3-containing forms of the EJC alters the set of transcripts subject to NMD, although distinct effects have been reported in two studies (Gerbracht et al, 2020; Mabin et al, 2018). In the mouse brain, haploinsufficiency for Magoh, Rbm8a or Eif4a3 causes severe microcephaly, but complete loss of Casc3 has a much milder effect that can be attributed to developmental delay (Mao et al, 2017; Mao et al, 2016; Mao et al, 2015; Silver et al, 2010).…”

Section: Introductionmentioning

confidence: 94%

An exon junction complex-independent function of Barentsz in neuromuscular synapse growth

Roignant

Tang

et al. 2021

Preprint

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The exon junction complex controls the translation, degradation and localization of spliced mRNAs, and three of its four core subunits also play a role in splicing. Here we show that the fourth subunit, Barentsz, has distinct biological functions within and separate from the exon junction complex in neuromuscular development. Barentsz controls the distribution of mitochondria in larval muscles, a function that also depends on other subunits of the exon junction complex and that is not rescued by a transgene in which residues required for binding to the core subunit eIF4AIII are mutated. In contrast, interactions with the exon junction complex are not required for Barentsz to promote the growth of neuromuscular synapses. We found that the Activin ligand Dawdle shows reduced expression in barentsz mutants and acts downstream of Barentsz to control synapse growth. Both barentsz and dawdle are required in motor neurons, muscles and glia for normal synapse growth, and exogenous Dawdle can rescue synapse growth in the absence of barentsz. These results identify a biological function for Barentsz that is independent of the exon junction complex.

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CASC3 promotes transcriptome-wide activation of nonsense-mediated decay by the exon junction complex

Cited by 38 publications

References 97 publications

UPF3A and UPF3B are redundant and modular activators of nonsense-mediated mRNA decay in human cells

UPF3A and UPF3B are redundant and modular activators of nonsense-mediated mRNA decay in human cells

Mammalian UPF3A and UPF3B activate NMD independently of their EJC binding

An exon junction complex-independent function of Barentsz in neuromuscular synapse growth

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