La-motif–dependent mRNA association with Slf1 promotes copper detoxification in yeast

Abstract

The La-motif (LAM) is an ancient and ubiquitous RNA-binding domain defining a superfamily of proteins, which comprises the genuine La proteins and La-related proteins (LARPs). In contrast to La, which binds and stabilizes pre-tRNAs and other RNA polymerase III transcripts, data on function and RNA targets of the LARPs have remained scarce. We have undertaken a global approach to elucidate the previously suggested role of the yeast LARP Slf1p in copper homeostasis. By applying RNA-binding protein immunopurification-microarray (RIP-Chip) analysis, we show that Slf1p and its paralog Sro9p copurify with overlapping sets of hundreds of functionally related mRNAs, including many transcripts coding for ribosomal proteins and histones. Interestingly, among these potential RNA targets were also mRNAs coding for proteins critical for protection of cells against elevated copper concentrations. Mutations introduced in the conserved aromatic patch of the LAM in Slf1p drastically impaired both association with its targets and Slf1-mediated protection of cells against toxic copper concentrations. Furthermore, we show that Slf1p stabilizes copper-related mRNA targets in a LAM-dependent manner. These results provide the first evidence for post-transcriptional regulation of factors/pathways implicated in copper homeostasis by a cytoplasmic RBP.

RNA regulons and the RNA-protein interaction network

“…Taken together, these results strongly suggested roles for Puf3p in the biogenesis and function of mitochondrial proteins. Indeed, later studies confi rmed the roles of Puf3p in mitochondrial function and in the localization of mRNAs (70) . Over-expression of Slf1p renders cells resistant to elevated copper concentrations in the medium, and this effect is dependent on the interaction of Slf1 with target mRNAs.…”

“…Over-expression of Slf1p renders cells resistant to elevated copper concentrations in the medium, and this effect is dependent on the interaction of Slf1 with target mRNAs. From this, we deduced that Slf1p regulates a copper regulon, thus allowing cells to adapt to elevated copper concentrations (70) . Such a mode of regulation would be analogous to a previously identifi ed yeast RBP network triggering coordinated responses to iron defi ciency (71) .…”

Quantitative proteomic analysis reveals concurrent RNA–protein interactions and identifies new RNA-binding proteins in Saccharomyces cerevisiae

“…It is thought that the specific complement of RBPs bound to a given mRNA specifies its post-transcriptional fate, but nearly all existing data are limited to defining pairwise interactions between a single RBP and a single mRNA species. Previous work to identify the mRNA targets bound by individual RBPs has mostly relied on purification of the RBP from a whole-cell lysate followed by analysis of the copurifying mRNAs (Gerber et al 2004;Ule et al 2005;Keene 2007a,b;Hogan et al 2008;Bohnsack et al 2009;Granneman et al 2009Granneman et al , 2010Wolf et al 2010;Scherrer et al 2011;Schenk et al 2012). These approaches do not differentiate between two RBPs that bind simultaneously to their common mRNA targets and two RBPs that bind to a common set of mRNA targets but at different times or in different cellular locations.…”

CAN1 Arginine Permease Deficiency Extends Yeast Replicative Lifespan via Translational Activation of Stress Response Genes

“…Slf1 has been previously shown to be involved in mRNA-specific regulation of translation in response to oxidative stress and to facilitate the translation of stress-responsive mRNAs (Kershaw et al, 2015), whereas Sbp1 is an abundant RBP found in stress granules and P-bodies implicated in translational repression (Mitchell et al, 2013; Segal et al, 2006). It has been also shown that overexpression of Slf1 increases mitochondrial respiration and leads to yeast chronological lifespan extension (Chatenay-Lapointe and Shadel, 2011), and in addition protects cells from copper-induced oxidative stress (Schenk et al, 2012). Consistent with activation of Slf1 in the tor1Δ mutant, cells lacking TOR1 demonstrated increased copper resistance compared to wild-type cells (Figure 4D).…”