Low complexity RGG-motif containing proteins Scd6 and Psp2 act as suppressors of clathrin heavy chain deficiency.

Clathrin, made up of the heavy- and light-chains, constitutes one of the most abundant proteins involved in intracellular protein trafficking and endocytosis. YPR129W, which encodes RGG-motif containing translation repressor was identified as a part of the multi-gene construct (SCD6) that suppressed clathrin deficiency. However, the contribution of YPR129W alone in suppressing clathrin deficiency has not been documented. This study identifies YPR129W as a necessary and sufficient gene in a multi-gene construct SCD6 that suppresses clathrin deficiency. Importantly, we also identify cytoplasmic RGG-motif protein encoding gene PSP2 as another novel suppressor of clathrin deficiency. Detailed domain analysis of the two suppressors reveals that the RGG-motif of both Scd6 and Psp2 is important for suppressing clathrin deficiency. Interestingly, the endocytosis function of clathrin heavy chain assayed by internalization of GFP-Snc1 and α-factor secretion activity are not complemented by either Scd6 or Psp2. We further observe that inhibition of TORC1 compromises the suppression activity of both SCD6 and PSP2 to different extent, suggesting that two suppressors are differentially regulated. Scd6 granules increased based on its RGG-motif upon Chc1 depletion. Strikingly, Psp2 overexpression increased the abundance of ubiquitin-conjugated proteins in Chc1 depleted cells in its RGG-motif dependent manner and also decreased the accumulation of GFP-Atg8 foci. Overall based on our results using SCD6 and PSP2, we identify a novel role of RGG-motif containing proteins in suppressing clathrin deficiency. Since both the suppressors are RNA-binding proteins, this study opens an exciting avenue for exploring the connection between clathrin function and post-transcriptional gene control processes.

Elucidation of the RNA-granule inducing sodium azide stress response through transcriptome analysis.

Sodium azide is a commonly used cytochrome oxidase inhibitor that leads to translation repression and RNA granule assembly. The global changes in mRNA abundance in response to this stressor are unknown. RGG-motif proteins Scd6 and Sbp1 are translation-repressors and decapping-activators that localize to and affect the assembly of RNA granules in response to sodium azide stress. Transcriptome-wide effects of these proteins remain unknown. To address this, we have sequenced transcriptome of the: a) wild type strain under unstressed and sodium azide stress, b) Δscd6 and Δsbp1 strains under unstressed and sodium azide stress. Transcriptome analysis identified altered abundance of many transcripts belonging to stress-responsive pathways which were further validated by qRT-PCR results. Abundance of several transcripts was altered in Δscd6/Δsbp1 under normal conditions and upon stress. Overall, this study provides critical insights into transcriptome changes in response to sodium azide stress and the role of RGG-motif proteins in these changes.