Transcript accumulation rates in the early Caenorhabditis elegans embryo

Dynamic transcriptional changes are widespread in rapidly dividing developing embryos when cell fate decisions are made quickly. The Caenorhabditis elegans embryo overcomes these constraints partly through the rapid production of high levels of transcription factor mRNAs. Transcript accumulation rates for some developmental genes are known at single-cell resolution, but genome-scale measurements are lacking. We estimate zygotic mRNA accumulation rates from single-cell RNA sequencing data calibrated with single-molecule transcript imaging. Rapid transcription is common in the early C. elegans embryo with rates highest soon after zygotic transcription begins. High-rate genes are enriched for recently duplicated cell-fate regulators and share common genomic features. We identify core promoter elements associated with high rate and measure their contributions for two early endomesodermal genes, ceh-51 and sdz-31. Individual motifs modestly affect accumulation rates, suggesting multifactorial control. These results are a step toward estimating absolute transcription kinetics and understanding how transcript dosage drives developmental decisions.

A. Example for half-life estimation using one-phase decay for ceh-51 RNA using embryo time course data (37).B.
Half-lives by accumulation rate categories, median indicated for each category.C. Adjusted synthesis rate for medium-and high-rate genes using estimated half-lives by cell type (compare to Fig. S2B).D. Fold difference between accumulation rate and adjusted synthesis rate for high-and medium-rate genes across cells.E. Intensities of ceh-51 exon and intron smFISH spots for individual 14-or 15-cell stage embryos.Intensities are an average +/-SEM of the number of ATS (x-axis) for each embryo.

Figure S3 -
Figure S3 -Related to Fig. 3: Half-life estimates and adjusted synthesis rates.

Figure S4 -
Figure S4 -Related to Fig. 4: Paralogs or orthologs in low vs. high-rate genes.A. Proportion of all high-and low-rate genes with at least one human ortholog using Ortholist 2 (43).B. Paralogs (Blast result from a e-value threshold of 10 −15 ) that are also syn-expressed from Tintori et al. (20).C. Proportion of all high-rate vs. all genes that have paralogs that are also high-rate.p-values from Fisher's Exact Test.

Figure S5 -
Figure S5 -Related to Fig. 5: Trans-splicing and distribution of genes on the different chromosomes.A. Proportion of genes that are trans-spliced or not in each rate group using data from Saito et al. (49).Wilcoxon Test with adjusted p-values.B. Proportion of genes in each rate category normalized by the number of genes on each chromosome.C. Number of genes as a proportion of the genes in either low-or medium+high-rate categories found

Figure S6 -
Figure S6 -Related to Fig. 5F: Karyogram of clustered genes on all chromosomes for indicated cell types.A. Karyogram showing absolute change of genes over genomic position (coordinate) for genes in E cells with Fold Change > 5 over the parent EMS cell.B and C. Same as A but for MSx1 cell compared to MS parent (B) and for ABprx compared to ABpr parent (C).Genes are colored as high-or medium rate.

Figure S7 -
Figure S7 -Related to Fig. 6: Motif enrichment of lineage-specific TFs. A. SKN-1 motif enrichment in genes expressed in the EMS compared to those expressed in all cells except EMS.B, C. END and MED motif enrichment in the E lineage (E, Ea, Ep cells) compared with genes expressed in all but the E lineage.C shows enrichment in genes unique to E vs. not.D. Motif occurrences for all lineage-specific motifs (in Fig. 6A) combined across the three rate categories.p-values from Chi-squared Test, FDR adjusted.

Figure S8 -
Figure S8 -Related to Fig. 7: ceh-51 and sdz-31 promoter region and survival of motif deletion mutants.A. Detailed sequence of up to -200bp from the ATG start codon of the ceh-51 promoter.Motifs indicated in filled colored boxes.Colored boxes around the sequence shows the deleted regions in indicated mutants.B. Embryonic and larval lethality after treatment with sptf-3 RNAi (SP1), vector only (C) or pop-1 RNAi, N=at least 4 biological replicates.C. Survival in motif mutants at 20°C and 25°C, N=at least 4 biological replicates.D. smFISH counts for sdz-31 over developmental time comparing WT and the germline cas9 strain (EG9882).E. 200bp upstream region (from translation start ATG) of sdz-31 showing important motifs and deletion in the Inr mutant.F. smFISH counts of sdz-31 at the indicated stages in WT and the Inr promoter mutant.