Figure S1.
Rie1 binds RNA and is involved in meiotic entry. (A) Overview of protein characteristics of screened RBPs. For our forward screen for meiotic entry mutants, we considered 94 total genes encoding RBPs harboring at least one of the following domains: RRM, zinc finger (ZnF), K Homology domains (KH), Pumilio Family domains (PUF), double-stranded RNA binding domains (dsRBD), or pentatricopeptide repeat domains (PPR). Mutants in non-essential genes (green) were included in the screen. Mutants in essential genes (magenta) or petites (orange) were not included. (B and C) Strains harboring RIE1-3V5 (B3114, blue), rie1 F240L-3V5 (B2674, yellow), rie1 F298L-3V5 (B2816, pink), or rie1 604L-3V5 (B2680, purple) were induced to sporulate at 30°C. (B) Cells were collected either with or without UV crosslinking at 2 h in SPO. Samples were treated with DNAse and Rie1 was IPed under denaturing conditions, and bound RNA was assessed by a polynucleotide labeling assay. Shown are autoradiograms (top) indicating bound RNA and immunoblot (IB; bottom) indicating IPed Rie1. Quantifications of RNA-binding signal/IPed Rie1 protein are indicated below. (C) Protein levels of Rie1 and Pgk1 (loading) were determined by immunoblot at the indicated times. Quantification of Rie1 normalized to Pgk1 (three technical replicates) is shown below. (D) Strains harboring RIE1-3V5 were grown in either YPD (log phase), BYTA (stationary phase), or induced to sporulate at 30°C. Samples were collected at the indicated times, and Rie1 and Pgk1 (loading) protein levels were determined by immunoblot. Quantifications of Rie1/Pgk1 compared with YPD (set at 1) are indicated below. Biological replicates = 3. (E) Strains harboring HTB2-mCherry and either RIE1-ENVY (B1554) or wild type RIE1 (no tag control, B3680) were induced to sporulate at 30°C. Cells were collected at the indicated times and imaged by fluorescence microscopy. Rie1 is shown in green and Htb2 (histone indicating nucleus) is shown in magenta. Green signal in no tag likely represents autofluorescence from bud scars and mitochondria. Scale bar, 2 µm. Source data are available for this figure: SourceData FS1.

Rie1 binds RNA and is involved in meiotic entry. (A) Overview of protein characteristics of screened RBPs. For our forward screen for meiotic entry mutants, we considered 94 total genes encoding RBPs harboring at least one of the following domains: RRM, zinc finger (ZnF), K Homology domains (KH), Pumilio Family domains (PUF), double-stranded RNA binding domains (dsRBD), or pentatricopeptide repeat domains (PPR). Mutants in non-essential genes (green) were included in the screen. Mutants in essential genes (magenta) or petites (orange) were not included. (B and C) Strains harboring RIE1-3V5 (B3114, blue), rie1 F240L-3V5 (B2674, yellow), rie1 F298L-3V5 (B2816, pink), or rie1 604L-3V5 (B2680, purple) were induced to sporulate at 30°C. (B) Cells were collected either with or without UV crosslinking at 2 h in SPO. Samples were treated with DNAse and Rie1 was IPed under denaturing conditions, and bound RNA was assessed by a polynucleotide labeling assay. Shown are autoradiograms (top) indicating bound RNA and immunoblot (IB; bottom) indicating IPed Rie1. Quantifications of RNA-binding signal/IPed Rie1 protein are indicated below. (C) Protein levels of Rie1 and Pgk1 (loading) were determined by immunoblot at the indicated times. Quantification of Rie1 normalized to Pgk1 (three technical replicates) is shown below. (D) Strains harboring RIE1-3V5 were grown in either YPD (log phase), BYTA (stationary phase), or induced to sporulate at 30°C. Samples were collected at the indicated times, and Rie1 and Pgk1 (loading) protein levels were determined by immunoblot. Quantifications of Rie1/Pgk1 compared with YPD (set at 1) are indicated below. Biological replicates = 3. (E) Strains harboring HTB2-mCherry and either RIE1-ENVY (B1554) or wild type RIE1 (no tag control, B3680) were induced to sporulate at 30°C. Cells were collected at the indicated times and imaged by fluorescence microscopy. Rie1 is shown in green and Htb2 (histone indicating nucleus) is shown in magenta. Green signal in no tag likely represents autofluorescence from bud scars and mitochondria. Scale bar, 2 µm. Source data are available for this figure: SourceData FS1.

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