§ 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

WD40 repeat proteins fold into characteristic ␤-propeller structures and control signaling circuits during cellular adaptation processes within eukaryotes. The RACK1 protein of Saccharomyces cerevisiae, Asc1p, consists exclusively of a single sevenbladed ␤-propeller that operates from the ribosomal base at the head region of the 40S subunit. Here we show that the R38D K40E ribosomal binding-compromised variant (Asc1DEp) is severely destabilized through mutation of phosphosite T143 to a dephosphorylation-mimicking alanine, probably through proteasomal degradation, leading to asc1 Ϫ phenotypes. Phosphosite Y250 contributes to resistance to translational inhibitors but does not influence Asc1DEp stability. Beyond its own phosphorylation at T143, Y250, and other sites, Asc1p heavily influences the phosphorylation of as many as 90 proteins at 120 sites. Many of these proteins are regulators of fundamental processes ranging from mRNA translation to protein transport and turnover, cytoskeleton organization, and cellular signaling. Our data expose Asc1p/ RACK1 as a key factor in phosphosignaling and manifest it as a control point at the head of the ribosomal 40S subunit itself regulated through posttranslational modification.

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