Modular, cascade-like transcriptional program of regeneration in Stentor

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Abstract

The giant ciliate Stentor coeruleus is a classical model system for studying regeneration and morphogenesis at the level of a single cell. The anterior of the cell is marked by an array of cilia, known as the oral apparatus, which can be induced to shed and regenerate in a series of reproducible morphological steps, previously shown to require transcription. If a cell is cut in half, each half will regenerate an intact cell, including a new oral apparatus in the posterior half. We used RNAseq to assay the dynamic changes in Stentor's transcriptome during regeneration, after both oral apparatus shedding and bisection, allowing us to identify distinct temporal waves of gene expression including kinases, RNA binding proteins, centriole biogenesis factors, and orthologs of human ciliopathy genes implicated in Meckel and Joubert syndromes. By comparing transcriptional profiles of different regeneration events in the same species, we were able to identify distinct modules of gene expression corresponding to oral apparatus regeneration, posterior holdfast regeneration, and recovery after wounding. By measuring gene expression in cells in which translation is blocked, we show that the sequential waves of gene expression involve a cascade mechanism in which later waves of expression are triggered by translation products of early-expressed genes. Among the early-expressed genes, we identified an E2F transcription factor and the conserved RNA binding protein Pumilio as potential regulators of regeneration based on the expression pattern of their predicted target genes. RNAi mediated knockdown experiments indicate that Pumilio is required for regenerating oral structures of the correct size. We show that E2F is involved in the completion of regeneration but is dispensable for earlier steps. This work allows us to classify regeneration genes into groups based on their potential role for regeneration in distinct cell regeneration paradigms, and provides insight into how a single cell can coordinate complex morphogenetic pathways to regenerate missing structures.

Data availability

transcriptomic data have been deposited in GEO under accession code GSE186036

The following data sets were generated

1. Sood P
2. Marshall W
3. Lin A
4. McGillivary R
(2021) Modular, Cascade-like Transcriptional Program of Regeneration in Stentor
NCBI Gene Expression Omnibus, GSE186036.

https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE186036

Article and author information

Author details

Pranidhi Sood

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States

Competing interests
The authors declare that no competing interests exist.
Athena Lin

Department of Biochemistry and BioPhysics, University of California, San Francisco, San Francisco, United States

Competing interests
The authors declare that no competing interests exist.
Connie Yan

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States

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The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-9961-0671
Rebecca McGillivary

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States

Competing interests
The authors declare that no competing interests exist.
Ulises Diaz

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States

Competing interests
The authors declare that no competing interests exist.
Tatyana Makushok

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States

Competing interests
The authors declare that no competing interests exist.
Ambika Nadkarni

Department of Mechanical Engineering, Stanford University, palo alto, United States

Competing interests
The authors declare that no competing interests exist.
Sindy KY Tang

Department of Mechanical Engineering, Stanford University, Palo Alto, United States

Competing interests
The authors declare that no competing interests exist.
Wallace F Marshall

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States

For correspondence
wallace.marshall@ucsf.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-8467-5763

Funding

National Institutes of Health (R35 GM130327)

Wallace F Marshall

National Science Foundation (MCB-1938102)

Sindy KY Tang
Wallace F Marshall

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.