Natural depletion of histone H1 in sex cells causes DNA demethylation, heterochromatin decondensation and transposon activation
- John Innes Centre, United Kingdom
Figures
Figure 1 with 1 supplement
VC-activated TEs are heterochromatic and demethylated by DME.
(A) Expression and locations of VC-activated TEs along Chromosome 5. The purple ellipse and red bars indicate the centromere and borders of pericentromeric regions, respectively. (B) Box plots …
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Figure 1—source data 1
List of VC-activated TEs.
- https://doi.org/10.7554/eLife.42530.005
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Figure 1—source data 2
List of VC DME targets.
- https://doi.org/10.7554/eLife.42530.006
Figure 1—figure supplement 1
VC-activated TEs are heterochromatic and demethylated by DME.
(A) Chromosomal view of VC-activated TEs, similar to Figure 1A. (B) Box plots showing the enrichment of euchromatic histone modifications at TEs, similar to Figure 1B. Difference between the two …
Figure 2 with 2 supplements
Ectopic H1 expression in the vegetative cell impedes DME at the most heterochromatic loci.
(A–B) Confocal images showing H1 localization under native promoter (A) and VC-specific promoter (pVC, B) during male gametogenesis. Msp, microspore; BCP, bicellular pollen; TCP, tricellular pollen. …
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Figure 2—source data 1
List of H1 hyperDMRs.
- https://doi.org/10.7554/eLife.42530.010
Figure 2—figure supplement 1
H1 ectopic expression in the vegetative cell causes pollen defect and reduced fertility.
pVC::H1 (pLAT52::H1-mRFP line #2) plants show reduced silique length (A) and an increased proportion of malformed pollen grains (B), which are indicated by red arrows in the SEM image (C). (D) Box …
Figure 2—figure supplement 2
H1 ectopic expression in the vegetative cell causes DNA hypermethylation at DME targets.
(A,C,D) Kernel density plots showing frequency distribution of methylation differences between VCs from pVC::H1 and WT in all 50 bp windows (blue traces) and windows overlapping VC DME targets …
Figure 3 with 1 supplement
Vegetative-cell-expressed H1 represses heterochromatic TEs in a dosage-dependent manner.
pVC::H1 refers to line #2 except as specified in A) and F). (A,F) quantitative RT-PCR demonstrating H1.1 (A) or TE (F) expression in pollen from WT and two independent pVC::H1 transgenic lines. …
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Figure 3—source data 1
List of pollen-expressed genes and their expression in WT and pVC::H1 pollen.
- https://doi.org/10.7554/eLife.42530.013
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Figure 3—source data 2
List of imprinted genes and their expression in WT and pVC::H1 pollen.
- https://doi.org/10.7554/eLife.42530.014
Figure 3—figure supplement 1
Two genes suppressed by VC-expressed H1 that gain methylation at nearby DME targets.
Top two traces show expression (Log2RPKM) in pVC::H1 (line #2) or WT pollen; three traces at the bottom show differential DNA methylation between pVC::H1 (line #2) and WT VC. The red lines indicate …
Figure 4
H1 suppresses TEs in the vegetative cell via two mechanisms.
(A) Heat map demonstrating DNA methylation differences between pVC::H1 and WT VCs within 500 bp of the TSS of H1-repressed TEs. Asterisks mark TEs whose suppression is not caused by changes in DNA …
Figure 5
Depletion of H1 decondenses heterochromatin in leaves and late microspores.
(A) Immunostaining with GFP and H3K9me2 antibodies showing the co-localization of H1 and H3K9me2-enriched chromocenters. (B) Examples of leaf nuclei with condensed, intermediate or decondensed …
Figure 6
Model depicting four mechanisms underlying TE activation in the VC.
The number of TEs in each group is shown on the top. Significantly less heterochromatic than TEs in other groups (Figure 3H), Group I TEs are activated by DME-directed DNA demethylation. Group II …
Author response image 1
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Gene (Arabidopsis thaliana) | H1.1 | NA | AT1G06760 | |
| Gene (A. thaliana) | H1.2 | NA | AT2G30620 | |
| Gene (A. thaliana) | DEMETER | NA | AT5G04560 | |
| Genetic reagent (A. thaliana) | h1.1–1 | Nottingham Arabidopsis Stock Centre | SALK_128430C | |
| Genetic reagent (A. thaliana) | h1.2–1 | GABI-Kat | GABI_406H11 | |
| Genetic reagent (A. thaliana) | dme-7 | Nottingham Arabidopsis Stock Centre | SALK_107538 | |
| Genetic reagent (A. thaliana) | pLAT52::H1.1-mRFP | this paper | LAT52 promoter from tomato is used | |
| Genetic reagent (A. thaliana) | pMSP1::H1.1-mRFP | this paper | MSP1 (AT5G59040) promoter is used | |
| Biological sample (A. thaliana) | sperm nuclei from Col-0 | this paper | bisulfite-sequencing | |
| Biological sample (A. thaliana) | vegetative nuclei from Col-0 | this paper | bisulfite-sequencing | |
| Biological sample (A. thaliana) | vegetative nuclei from pLAT52::H1.1-mRFP | this paper | bisulfite-sequencing | |
| Biological sample (A. thaliana) | pollen from Col-0 | this paper | RNA-sequencing | |
| Biological sample (A. thaliana) | pollen from pLAT52::H1.1-mRFP | this paper | RNA-sequencing | |
| Biological sample (A. thaliana) | pollen from dme-7/+ | this paper | RNA-sequencing | |
| Antibody | GFP | Abcam | Cat# ab290, RRID:AB_303395 | (1:100) |
| Antibody | H3K9me2 | Abcam | Cat# ab1220, RRID:AB_449854 | (1:100) |
| Antibody | Alexa 488- secondaries | ThermoFisher Scientific | Cat# A-11034, RRID:AB_2576217 | (1:200) |
| Antibody | Alexa 555- secondaries | ThermoFisher Scientific | Cat# A28180, RRID:AB_2536164 | (1:200) |
| Software, algorithm | Mikado | Venturini et al., 2018 | RRID:SCR_016159 | |
| Software, algorithm | Kallisto | Bray et al., 2016 | RRID:SCR_016582 | |
| Software, algorithm | Sleuth | Pimentel et al., 2017 | RRID:SCR_016883 | |
| Other | Hoechst 33342 | ThermoFisher Scientific | Cat# H3570 | (1:1000) |
| Other | DAPI | ThermoFisher Scientific | Cat# D1306, RRID:AB_2629482 | (100 ng/mL) |
Additional files
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Supplementary file 1
Sequencing summary statistics for bisulfite sequencing libraries.
Mean DNA methylation (Met) was calculated by averaging methylation of individual cytosines in each context, and chloroplast CHH methylation was used as a measure of cytosine non-conversion and other errors. SN, sperm nuclei; VN, vegetative nuclei.
- https://doi.org/10.7554/eLife.42530.018
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Supplementary file 2
List of primers for quantitative RT-PCR.
- https://doi.org/10.7554/eLife.42530.019
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Transparent reporting form
- https://doi.org/10.7554/eLife.42530.020
