In Saccharomyces cerevisiae, adhesive growth on solid surfaces is mediated by the flocculin Flo11 to confer biofilm and filament formation. Expression of FLO11 is governed by a complex regulatory network that includes, e.g., the protein kinase A (PKA) signaling pathway. In addition, numerous regulatory genes, which have not been integrated into regulatory networks, affect adhesive growth, including WHI3 encoding an RNA-binding protein and YAK1 coding for a dual-specificity tyrosine-regulated protein kinase. In this study, we present evidence that Whi3 and Yak1 form part of a signaling pathway that regulates FLO11-mediated surface adhesion and is involved in stress resistance. Our study further suggests that Whi3 controls YAK1 expression at the post-transcriptional level and that Yak1 targets the transcriptional regulators Sok2 and Phd1 to control FLO11. We also discovered that Yak1 regulates acidic stress resistance and adhesion via the transcription factor Haa1. Finally, we provide evidence that the catalytic PKA subunit Tpk1 inhibits Yak1 by targeting specific serine residues to suppress FLO11. In summary, our data suggest that Yak1 is at the center of a regulatory cascade for adhesive growth and stress resistance, which is under dual control of Whi3 and the PKA subunit Tpk1.

23The ubiquitin-proteasome system (UPS) controls cellular functions by maintenance of a 24 functional proteome and degradation of key regulatory proteins. Central to the UPS is the 25 proteasome that adjusts the abundance of numerous proteins, thereby safeguarding their 26 activity or initiating regulatory events. Here, we demonstrate that the essential 27Saccharomyces cerevisiae protein Yjr141w/Ipa1 (Important for cleavage and 28 PolyAdenylation) belongs to the HECT_2 (homologous to E6-AP carboxyl terminus_2) 29 family. We found that five cysteine residues within the HECT_2 family signature and the C-30 terminus are essential for Ipa1 activity. Furthermore, Ipa1 interacts with several ubiquitin-31 conjugating enzymes in vivo and localizes to the cytosol and nucleus. Importantly, Ipa1 32 has impact on proteasome activity, which is indicated by the activation of the Rpn4 regulon 33 as well as by decreased turnover of destabilized proteasome substrates in an IPA1 mutant. 34These changes in proteasome activity might be connected to reduced maturation or 35 modification of proteasomal core particle proteins. Our results highlight the influence of 36Ipa1 on the UPS. The conservation within the HECT_2 family and the connection of the 37 human HECT_2 family member to an age-related degeneration disease might suggest that 38

In Saccharomyces cerevisiae, the RNA-binding protein Whi3 controls cell cycle progression, biofilm formation, and stress response by post-transcriptional regulation of the Cdc28-Cln3 cyclin-dependent protein kinase and the dual-specificity protein kinase Yak1. Previous work has indicated that Whi3 might govern these processes by additional, yet unknown mechanisms. In this study, we have identified additional effectors of Whi3 that include the G 1 cyclins Cln1/Cln2 and two known regulators of biofilm formation, the catalytic PKA subunit Tpk1 and the transcriptional activator Tec1. We also provide evidence that Whi3 regulates production of these factors by post-transcriptional control and might exert this function by affecting translational elongation. Unexpectedly, we also discovered that Whi3 is a key regulator of cellular ploidy, because haploid whi3Δ mutant strains exhibit a significant increase-in-ploidy phenotype that depends on environmental conditions. Our data further suggest that Whi3 might control stability of ploidy by affecting the expression of many key genes involved in sister chromatid cohesion and of NIP100 that encodes a component of the yeast dynactin complex for chromosome distribution. Finally, we show that absence of Whi3 induces a transcriptional stress response in haploid cells that is relieved by whole-genome duplication. In summary, our study suggests that the RNA-binding protein Whi3 acts as a central regulator of cell division and development by post-transcriptional control of key genes involved in chromosome distribution and cell signaling. E UKARYOTIC organisms have evolved highly complex signaling networks to control cell division and development. The functionality of signal transduction pathways crucially depends on the timely availability of individual components at sufficient levels to ensure adequate signaling capacity. A number of studies in different eukaryotes have shown that the production of individual signaling proteins is under the control not only of transcriptional regulators, but also of RNA-binding proteins (Lasko 2003;Sugiura et al. 2003;Prinz et al. 2007;Stewart et al. 2007;Malcher et al. 2011). This suggests that RNA-binding proteins could regulate the performance of whole signaling networks, but a precise view of key interconnections is often lacking.In the budding yeast Saccharomyces cerevisiae, the Pumilio family (PUF) protein Mpt5/Puf5 is a good example of an RNA-binding protein that controls cellular development by targeting signaling components. Mpt5 has been found to negatively affect the performance of at least two different mitogen-activated protein kinase (MAPK) signaling pathways that regulate adhesion/biofilm formation and cell wall integrity (Prinz et al. 2007;Stewart et al. 2007). In general, PUF proteins are known to bind to the 39-UTR of target mRNAs and reduce their stability or translatability (Quenault et al. 2011). This is also true for S. cerevisiae Mpt5, which represses the protein levels of key components of the MAPK cascade that regu...