Functions of the Nonsense-Mediated mRNA Decay Pathway in Drosophila Development

Abstract

Nonsense-mediated mRNA decay (NMD) is a cellular surveillance mechanism that degrades transcripts containing premature translation termination codons, and it also influences expression of certain wild-type transcripts. Although the biochemical mechanisms of NMD have been studied intensively, its developmental functions and importance are less clear. Here, we describe the isolation and characterization of Drosophila “photoshop” mutations, which increase expression of green fluorescent protein and other transgenes. Mapping and molecular analyses show that photoshop mutations are loss-of-function mutations in the Drosophila homologs of NMD genes Upf1, Upf2, and Smg1. We find that Upf1 and Upf2 are broadly active during development, and they are required for NMD as well as for proper expression of dozens of wild-type genes during development and for larval viability. Genetic mosaic analysis shows that Upf1 and Upf2 are required for growth and/or survival of imaginal cell clones, but this defect can be overcome if surrounding wild-type cells are eliminated. By contrast, we find that the PI3K-related kinase Smg1 potentiates but is not required for NMD or for viability, implying that the Upf1 phosphorylation cycle that is required for mammalian and Caenorhabditis elegans NMD has a more limited role during Drosophila development. Finally, we show that the SV40 3′ UTR, present in many Drosophila transgenes, targets the transgenes for regulation by the NMD pathway. The results establish that the Drosophila NMD pathway is broadly active and essential for development, and one critical function of the pathway is to endow proliferating imaginal cells with a competitive growth advantage that prevents them from being overtaken by other proliferating cells.

Smg1 is required for embryogenesis and regulates diverse genes via alternative splicing coupled to nonsense-mediated mRNA decay

“…For example, in C. elegans, the Smg1 homologue is important for NMD but dispensable for survival (27). In D. melanogaster and D. rerio, the NMD factors Upf1 and Upf2, but not Smg1, are necessary for development (28,29). In mice, deficiency for Upf1 or Upf2 abolishes NMD and results in early embryonic lethality during the periimplantation period (30,31).…”

High-resolution profiling of NMD targets in yeast reveals translational fidelity as a basis for substrate selection

“…However, NMD also targets a significant fraction of apparently normal and physiologically functional wild-type mRNAs (Schweingruber et al 2013), indicating that it also serves as a fundamental posttranscriptional regulatory mechanism for eukaryotic gene expression. Consistent with these important roles, NMD function is linked to diverse cellular processes, including cell growth and proliferation (Weischenfeldt et al 2008;Avery et al 2011;Lou et al 2014), development and differentiation (Medghalchi et al 2001;Metzstein and Krasnow 2006;Gong et al 2009;Wittkopp et al 2009), innate immunity (Gloggnitzer et al 2014), antiviral or stress responses (Sakaki et al 2012;Balistreri et al 2014), and neuronal activity or behavior (Giorgi et al 2007;Colak et al 2013).…”

Rapid identification of mRNA processing defects with a novel single-cell yeast reporter

“…A similar approach was used with a lacZ construct that was used to assay the effects of cis-elements on transcript stability (Muhlrad et al 1995). There are also NMD reporters that use GFP (Paillusson et al 2005) and chemiluminescence (Boelz et al 2006) in mammalian cells, one for Giardia lamblia that uses chemiluminescence (Chen et al 2008), and one utilizing GFP and/or DsRed in Drosophila melanogaster (Metzstein and Krasnow 2006).…”