Additional file 1 of Folate regulates RNA m5C modification and translation in neural stem cells
Creators: name: Xu, Xiguang
name: Johnson, Zachary
name: Wang, Amanda
name: Padget, Rachel L.
name: Smyth, James W.
name: Xie, Hehuang
Publisher: name: figshare
Year: 2022
Subjects: subject: Genetics
subject: FOS: Biological sciences

description: Additional file 1: Figure S1. Quality control of total mRNA-seq datasets. (a, b, c) Scatter plot showing the Pearson correlation between two biological replicates in total mRNA-seq datasets in NSCs. (d) Principal component analysis of total mRNA-seq datasets. Figure S2. Validation of differentially expressed genes (DEGs). (a-g) RT-qPCR was performed to validate the expression of DEGs identified by RNA-seq. (*&lt;0.05, **&lt;0.01, ***&lt;0.001.) Numerical data is provided in Additional file 7. The error bars indicate standard error of the mean (SEM). Figure S3. Distribution of m5C sites in total mRNA BS-seq datasets. (a) Pie chart showing the categories of RNA molecules with m5C modification. (b) Histogram showing the distribution of methylation level for each condition. Figure S4. Quality control of polysome mRNA-seq datasets. (a, b, c) Scatter plot showing the Pearson correlation between two biological replicates in polysome mRNA-seq datasets in NSCs. (d) Principal component analysis of polysome mRNA-seq datasets. Figure S5. Validation of differentially translated genes (DTGs). (a, b) Western blot was performed to validate the expression of DTGs identified by polysome profiling. Gng11 showed increased translation efficiency in LF (a), and Ptx3 showed increased translation efficiency in HF (b). Original blot images were provided in Additional file 8. Figure S6. Characterization of m5C sites in polysome mRNA BS-seq datasets. (a) Venn diagram showing the overlap of m5C sites between two replicates. (b) Dot plot showing the methylation level correlation of the overlapped m5C sites between two replicates. (c) Box plot showing the methylation level of overlapped and non-overlapped m5C sites in the two replicates. (d) Pie chart showing the categories of RNA molecules with m5C modification. (e) Histogram showing the distribution of methylation level for each condition. Figure S7. Functional annotation of m5C-modified mRNAs in polysome mRNAs. (a) Venn diagram showing the overlap of m5C sites among the three conditions (pLF, pMF, pHF) (b) Bubble plot showing the GO terms of m5C-modified mRNAs in pLF, pMF and pHF. Figure S8. Comparison between total and polysome methylomes. (a) Venn diagram showing the overlap of m5C sites between total and polysome mRNAs. (b) Box plot showing the methylation level of the union of m5C sites between total and polysome mRNAs with 20x coverage in both samples. A Wilcoxon signed-rank test was performed. Figure S9. Visualization of the correlation between m5C methylation and mRNA translation. (a, b, c) Left panel depicts transcript-level read pileup of the mRNAs: Hdgf (a), Glg1 (b), and Dgcr2 (c) using the IGV browser. The total and polysome mRNA-seq datasets were shown (green – LF, blue – MF, red –HF). Right panel depicted the methylation level of the m5C site identified in the respective transcript in both total and polysome mRNAs (t: total, p: polysome).