Flatworm-specific transcriptional regulators promote the specification of tegumental progenitors in Schistosoma mansoni
- UT Southwestern Medical Center, Texas
- Australian Institute of Tropical Health and Medicine, James Cook University, Australia
- Wellcome Genome Campus, United Kingdom
Figures
Figure 1 with 2 supplements
tsp-2+ neoblast progeny cells fuse with the tegumental syncytium in adult schistosomes.
(A) Cartoon depicting anatomy of the tegument and fluorescent dextran labeling. (B–D) Transverse planes through various levels of the tegument as indicated in (A). Phalloidin labels F-actin-rich (B) …
Figure 1—figure supplement 1
FISH examining the expression of several candidate tegument markers.
(A–C) FISH experiments on dextran-labeled worms demonstrating the localization of (A) annexin expression (n = 216 cells from three adult male parasites), (B) gtp-4 expression (n = 172 cells from …
Figure 1—figure supplement 2
Examination of TSP-2 protein localization.
(A) Western blot showing depletion of TSP-2 protein levels following tsp-2 RNAi. (B) Cartoon depicting dextran and TSP-2 labeling of the tegument. (C) Transverse image at the level indicated in (B) …
Figure 2 with 3 supplements
FACS purification and transcriptional profiling identifies molecules expressed in neoblasts and cells associated with the tegumental lineage.
(A) Cartoon depicting FACS purification strategy. (B) FACS plots showing various cell populations in control and following gamma-irradiation. Percentages represent fraction of the number of cells in …
Figure 2—figure supplement 1
Microscopic imaging of sorted ‘TSP-2 Intermediate’ cells.
Confocal images of sorted TSP-2 Intermediate cells. Arrows in the inset show cells with no TSP-2 labeling (red arrows), cells with pieces of TSP-2+ debris attached to them (yellow arrows), and …
Figure 2—figure supplement 2
Examination of the expression of genes expressed in TSP-2-enriched clusters.
FISH for tsp-2, a mixture of tegumental makers (Tegument), and panel of 15 genes from various clusters of gene expression (indicated at right of the image). Relative expression levels of each gene …
Figure 2—figure supplement 3
Graphical summary of genes expressed in TSP-2-enriched clusters.
A graphical summary indicating where in the tegumental lineage genes are expressed. An asterisk next to a gene name indicates that the gene is only expressed in a subset of the population. A …
Figure 3 with 3 supplements
An RNAi screen identifies zfp-1 and zfp-1–1 as genes required for the production of tsp-2+ cells.
(A) Cartoon depicting the RNAi screening strategy used to identify regulators of tegument progenitor specification. Candidate genes were knocked-down using RNAi, worms were pulsed with EdU for 4 hrs …
Figure 3—figure supplement 1
RNAi screen of candidate tegument biogenesis regulators.
Results of knocking down candidate transcripts that are dispensable for normal neoblast function and tsp-2+ cell production. Representative maximum intensity confocal projections are shown. Numbers …
Figure 3—figure supplement 2
Quantification of gene expression in zfp-1(RNAi) and zfp-1-1(RNAi) parasites.
Quantitative real time PCR analysis of the effects of zfp-1 and zfp-1–1 RNAi on the expression of (A) zfp-1, (B) zfp-1–1, (C) sm13, and (D) tsp-2. Each bar represents the expression of the indicated …
Figure 3—figure supplement 3
Gene expression patterns of sm13 and zfp-1–1.
Representative double FISH demonstrating the localization of sm13 expression relative to zfp-1–1 expression. 1 of 248 sm13+ cells was zfp-1–1 positive. Image represents a z-projection. Scale bar: 10 …
Figure 4
ZFP-1 and ZFP-1–1 are flatworm specific zinc finger proteins and are putative transcriptional regulators.
(A) Multiple protein sequence alignment of the C2H2 domain of several zfp-1 and zfp-1–1 homologs. Zinc coordinating residues are shown in black background. Conserved residues contributing to high …
Figure 5
ZFP-1 family proteins are required for the production of new tegumental cells.
(A) Cartoon depicting the strategy for fate-mapping by EdU pulse-chase experiments. (B) FISH for tsp-2 and tegumental markers with EdU detection in zfp-1(RNAi) or zfp-1-1(RNAi) parasites at day …
Figure 6
Model for the specification of new tegumental cells from neoblasts.
Neoblasts (magenta cells) expressing nanos2 and zfp-1 specify large numbers of tsp-2+ cells. Some fraction of tsp-2 cells express zfp-1–1. Within this tsp-2 compartment are cells that extend …
Videos
Video 1
Movie depicting the elaborate interconnected network of the parasite’s tegument and the attached tegumental cell bodies.
https://doi.org/10.7554/eLife.33221.006
Video 2
Movie depicting the localization of TSP-2 protein relative to the tegument and tsp-2 expressing cells.
https://doi.org/10.7554/eLife.33221.007Additional files
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Supplementary file 1
Pairwise comparisons of transcriptional profiles of neoblasts, TSP-2+ cells, and IR Rest cells.
- https://doi.org/10.7554/eLife.33221.019
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Supplementary file 2
Results of model-based clustering analysis to define genes whose expression is enriched in either neoblasts, TSP-2+ cells, or IR Rest cells.
- https://doi.org/10.7554/eLife.33221.020
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Supplementary file 3
Table of candidate tegument-associated genes, their abbreviations, their gene expression cluster, and their expression pattern.
- https://doi.org/10.7554/eLife.33221.021
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Supplementary file 4
Pairwise comparisons of transcriptional profiles of control(RNAi) parasites versus zfp-1-1(RNAi) parasites.
- https://doi.org/10.7554/eLife.33221.022
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Supplementary file 5
Table of gene names, abbreviations, and oligonucleotides sequences from this study.
- https://doi.org/10.7554/eLife.33221.023
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Supplementary file 6
Table of results from Fisher’s exact test to define how zfp-1–1 RNAi treatment affects genes expressed in various gene expression clusters.
- https://doi.org/10.7554/eLife.33221.024
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Supplementary file 7
FASTA formatted file with various flatworm ZFP-1 family protein sequences that were used for generating protein alignments and phylogenetic trees.
- https://doi.org/10.7554/eLife.33221.025
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Transparent reporting form
- https://doi.org/10.7554/eLife.33221.026
