Cell Morphogenesis Proteins Are Translationally Controlled through UTRs by the Ndr/LATS Target Ssd1

Abstract

Eukaryotic cells control their growth and morphogenesis to maintain integrity and viability. Free-living cells are further challenged by their direct interaction with the environment and in many cases maintain a resilient cell wall to stay alive under widely varying conditions. For these organisms, stringent and highly localized control of the cell wall's remodeling and expansion is crucial for cell growth and reproduction. In the budding yeast Saccharomyces cerevisiae the RNA binding protein Ssd1 helps control cell wall remodeling by repressing translation of proteins involved in wall expansion. Ssd1 is itself negatively regulated by the highly conserved Ndr/LATS protein kinase Cbk1. We sought to identify mRNA regions that confer Ssd1-mediated translational control. After validating a GFP reporter system as a readout of Ssd1 activity we found that 3′ untranslated regions of the known Ssd1 targets CTS1, SIM1 and UTH1 are sufficient for Cbk1-regulated translational control. The 5′ untranslated region of UTH1 also facilitated Ssd1-mediated translational control in a heterologous context. The CTS1 and SIM1 3′ untranslated regions confer Ssd1 binding, and the SIM1 3′ untranslated region improves Ssd1 immunoprecipitation of the endogenous SIM1 transcript. However, SIM1's 3′ untranslated region is not essential for Ssd1-regulated control of the message's translation. We propose that Ssd1 regulates translation of its target message primarily through UTRs and the SIM1 message through multiple potential points of interaction, permitting fine translational control in various contexts.

The NDR Kinase Cbk1 Downregulates the Transcriptional Repressor Nrg1 through the mRNA-Binding Protein Ssd1 in Candida albicans

“…The finding that SSD1 deletion conferred the ability to completely eliminate Nrg1 during hyphal initiation in the cbk1⌬/⌬ mutant indicates that Ssd1 function is linked with Nrg1 downregulation. Because Ssd1 is an mRNA-binding protein that controls the translation of associated mRNAs (30,31,33), one or more specific mRNA(s) bound to Ssd1 might be responsible for Nrg1-related Ssd1 function (Fig. 8).…”

“…Although Ssd1 was reported to be genetically linked to various cellular functions, including stress signaling, cellular aging, and virulence (25)(26)(27)(28), the primary function of Ssd1 in S. cerevisiae is likely to modulate the expression and localization of mRNAs for cell wall proteins (29)(30)(31). Although Ssd1 phosphorylated by Cbk1 actively translates bound mRNAs, nonphosphorylated Ssd1 interacts with processing bodies (P bodies) and thus represses mRNA translation in S. cerevisiae (30)(31)(32)(33). These studies indicate that Cbk1 regulates Ssd1 activity to precisely coordinate cell wall remodeling during isotropic and polarized S. cerevisiae growth.…”

5′ untranslated regions: the next regulatory sequence in yeast synthetic biology

“…). However, it was shown that the RNA motif is not necessary or sufficient for Ssd1‐mediated control (Wanless, Lin, & Weiss, ). At present, it is likely that RBPs are one of the mechanisms able to control gene expression of targeted mRNAs.…”

“…for Ssd1-mediated control (Wanless, Lin, & Weiss, 2014). At present, it is likely that RBPs are one of the mechanisms able to control gene expression of targeted mRNAs.…”

Asymmetric secretion in budding yeast reinforces daughter cell identity

“…To minimize disruption to endogenous Cts1 expression, we tagged Cts1 internal to the coding sequence with a monomeric superfolding GFP (msGFP), which folds properly in the endomembrane system (Fitzgerald and Glick, 2014;Day et al, 2018). This methodology leaves the 5' and 3' regulatory elements intact (Dohrmann et al, 1996;Aulds et al, 2012;Wanless et al, 2014).…”