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| Item Type: | Preprint |
|---|---|
| Title: | Analysis of developmental gene expression using smFISH and in silico staging of C. elegans embryos |
| Creators: |
Breimann, L.  ORCID: https://orcid.org/0000-0001-7702-5749, Bahry, E. ORCID: https://orcid.org/0000-0002-6358-2576, Zouinkhi, M. ORCID: https://orcid.org/0000-0002-9441-2908, Kolyvanov, K., Street, L.A. ORCID: https://orcid.org/0000-0002-5473-6713, Preibisch, S. ORCID: https://orcid.org/0000-0002-0276-494X and Ercan, S. ORCID: https://orcid.org/0000-0001-7297-1648
|
| Abstract: | Regulation of transcription during embryogenesis is key to development and differentiation. To study transcript expression throughout Caenorhabditis elegans embryogenesis at single-molecule resolution, we developed a high-throughput single-molecule fluorescence in situ hybridization (smFISH) method that relies on computational methods to developmentally stage embryos and quantify individual mRNA molecules in single embryos. We applied our system to sdc-2, a zygotically transcribed gene essential for hermaphrodite development and dosage compensation. We found that sdc-2 is rapidly activated during early embryogenesis by increasing both the number of mRNAs produced per transcription site and the frequency of sites engaged in transcription. Knockdown of sdc-2 and dpy-27, a subunit of the dosage compensation complex (DCC), increased the number of active transcription sites for the X chromosomal gene dpy-23 but not the autosomal gene mdh-1, suggesting that the DCC reduces the frequency of dpy-23 transcription. The temporal resolution from in silico staging of embryos showed that the deletion of a single DCC recruitment element near the dpy-23 gene causes higher dpy-23 mRNA expression after the start of dosage compensation, which could not be resolved using mRNAseq from mixed-stage embryos. In summary, we have established a computational approach to quantify temporal regulation of transcription throughout C. elegans embryogenesis and demonstrated its potential to provide new insights into developmental gene regulation. |
| Keywords: | Dosage Compensation, SmFISH, C. Elegans, X Chromosome, Sex Determination, Embryogenesis, Condensin, DCC, Transcription Burst, Transcription Frequency, Gene Regulation, Development, Animals, Worms |
| Source: | bioRxiv |
| Publisher: | Cold Spring Harbor Laboratory Press |
| Article Number: | 2024.05.15.594414 |
| Date: | 16 May 2024 |
| Official Publication: | https://doi.org/10.1101/2024.05.15.594414 |
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