RACK1/Asc1p, a Ribosomal Node in Cellular Signaling

Abstract

§ RACK1/Asc1p and its essential orthologues in higher eukaryotes, such as RACK1 in metazoa, are involved in several distinct cellular signaling processes. The implications of a total deletion have never been assessed in a comprehensive manner. This study reveals the major cellular processes affected in a Saccharomyces cerevisiae ⌬asc1 deletion background via de novo proteome and transcriptome analysis, as well as subsequent phenotypical characterizations. The deletion of ASC1 reduces iron uptake and causes nitrosative stress, both known indicators for hypoxia, which manifests in a shift of energy metabolism from respiration to fermentation in the ⌬asc1 strain. Asc1p further impacts cellular metabolism through its regulative role in the MAP kinase signal transduction pathways of invasive/filamentous growth and cell wall integrity. In the ⌬asc1 mutant strain, aberrations from the expected cellular response, mediated by these pathways, can be observed and are linked to changes in protein abundances of pathway-targeted transcription factors. Evidence of the translational regulation of such transcription factors suggests that ribosomal Asc1p is involved in signal transduction pathways and controls the biosynthesis of the respective final transcriptional regulators. Molecular & Cellular

Asc1p/RACK1 Connects Ribosomes to Eukaryotic Phosphosignaling

“…In line with the results of Coyle et al (1), the asc1DE strain behaved mainly like the ASC1 wild-type strain except for an increased sensitivity to cycloheximide (described below). Furthermore, the protein abundances of the transcription factors Flo8p, Tec1p, and Rap1p, which are heavily affected by the absence of Asc1p (15), remained unchanged in the DE background ( Fig. 2A).…”

“…Arginine-and lysine-auxotrophic strains were generated by successive replacement of the ARG4 and LYS1 genes with the recyclable loxP::URA3::loxP marker cassette, amplified from plasmid pUG72, according to the method of Gueldener et al (52). An arginine-and lysine-auxotrophic asc1SNR24 strain was generated from the ASC1 wild-type Δarg4::loxP Δlys1::loxP strain as described by Rachfall et al (15). Generation of an asc1DE strain expressing C-terminally 3ϫmyc-tagged Flo8p was done according to the method of Janke et al (53).…”

“…Asc1p/RACK1 affects translation, depending on the 5= untranslated regions of mRNAs (15) and through its interaction with mRNA-binding proteins, such as Scp160p in S. cerevisiae (16) and ZBP1 in mammalian neuronal cells (17). Mammalian RACK1 also regulates protein expression at a posttranslational level by affecting the degradation of certain proteins, e.g., the ␣-subunit of the transcription factor hypoxia-inducible factor 1 (HIF1␣) (18).…”

Asc1, homolog of human RACK1, prevents frameshifting in yeast by ribosomes stalled at CGA codon repeats

“…Human RACK1, a member of the TrpAsp (WD) repeat family, integrates numerous signaling pathways with translation regulation (Adams et al 2011), for instance, by mediating phosphorylation of eIF6 by protein kinase C to promote joining of the ribosomal subunits (Ceci et al 2003). Yeast Asc1 likewise has pleiotropic effects on the cell (Rachfall et al 2013), at least some of which are caused by its interaction with the ribosome (Coyle et al 2009); asc1-Δ mutants de-repress the general amino acid control genes in rich media (Hoffmann et al 1999), alter expression of several signaling pathways (Rachfall et al 2013), and fail to recruit the mRNA binding protein Scp160 to the ribosome (Baum et al 2004). Moreover, Asc1 protein levels are modulated, decreasing substantially during hypoxic stress (Bruckmann et al 2009), perhaps indicating that the response to ribosome stalls is regulated.…”

Identification of growth stage molecular markers in Trichoderma sp. ‘atroviride type B’ and their potential application in monitoring fungal growth and development in soil

“…Two global regulators (cross-pathway control proteins Cpc2 and 1 Cpc1/Gcn4) linked to amino acid biosynthesis and developmental programmes (Nishikawa et al, 2008) were identified in the germination stage. Cpc2/RACK1 is a highly conserved WD-domain protein found in all eukaryotes; in mammalian cells RACK1 acts as a scaffold protein that integrates inputs from different signalling pathways and affects translation through association with ribosomes (Rachfall et al, 2013). Genes encoding enzymes involved in the synthesis of amino acids were identified during germination stages such as: (a) TA-255300, which encodes a putative argininosuccinate synthetase protein involved in the arginine biosynthetic pathway (Wagemaker et al, 2007); (b) TA-302651, which encodes a saccharopine dehydrogenase that acts in the a-aminoadipate pathway for lysine biosynthesis in fungi (Xu et al, 2006); and (c) cystathionine gamma-synthase (TA-137525), which is an essential enzyme for methionine biosynthesis in various organisms, including Ara.…”