Capturig the Asc1p/RACK1 microenviroment at the head region of the 40S ribosome with quantitative BioID

The Asc1 protein of Saccharomyces cerevisiae is a Gβ-like scaffold protein considered to organize the local microenvironment at the head region of ribosomes in order to link mRNA translation with cellular signaling. In this study we used the in vivo protein labeling technique proximity-dependent Biotin IDentification (BioID) in combination with stable isotope labeling with amino acids in cell culture (SILAC) in S. cerevisiae to quantitatively monitor the proteinaceous Asc1p neighborhood in exponentially growing cells, but also to record alterations in response to glucose deprivation and heat stress. Beyond expected known interaction partners several proteins not known to co-localize with Asc1p were identified including mRNA-binding proteins, translational regulators and even transcription-related proteins. Additionally, we observed major alterations in response to glucose withdrawal. In contrast, heat stress caused only minor effects on the Asc1p neighborhood. Furthermore, we analyzed the molecular microenvironment of the “ribosome-binding deficient” Asc1DE protein, which revealed ribosome-association of the major Asc1DEp population in vivo, but suggests a distorted position of the variant at the ribosome.