HDAC1 SUMOylation promotes Argonaute-directed transcriptional silencing in C. elegans

  1. Heesun Kim
  2. Yue-He Ding
  3. Gangming Zhang
  4. Yong-Hong Yan
  5. Darryl Conte Jr
  6. Meng-Qiu Dong
  7. Craig C Mello  Is a corresponding author
  1. RNA Therapeutics Institute, University of Massachusetts Medical School, United States
  2. National Institute of Biological Sciences, China
  3. Howard Hughes Medical Institute, United States
7 figures, 1 table and 6 additional files


SUMOylation and chromatin remodeling factors promote piRNA-mediated silencing.

(A) Schematic of the piRNA sensor screen. The piRNA sensor strain contains a gfp::csr-1 transgene that is silenced by the piRNA pathway in the presence of an active oma-1::gfp transgene (Seth et al., 2018). OMA-1::GFP localizes to the cytoplasm of oocytes. Inactivation of the piRNA pathway (by RNAi, mutation, or auxin-inducible protein depletion) desilences the transgene, resulting in GFP::CSR-1 expression in perinuclear P-granules throughout the germline, as shown in (B). (B) Differential interference contrast and epifluorescence images of dissected gonads in wild-type (wt), prg-1(tm872), and wago-9/hrde-1(ne4769) worms. PRG-1 is required to initiate silencing, while WAGO-9 is required to maintain silencing. The percentage of desilenced worms and number of worms scored are shown. (C) Analysis of SUMO and chromatin remodeling factors required for piRNA-mediated silencing. Genes identified in the RNAi-based screen of chromatin factors are listed with their human homologs and with the percentage of worms that express GFP::CSR-1 among the total number of worms analyzed (n) when function is reduced by RNAi (blue column) or by either mutation or degron-dependent protein depletion (peach column).

SUMOylation of HDA-1 at K444 and K459 facilitates piRNA-mediated silencing.

(A) Domain structure of C. elegans type 1 histone deacetylase HDA-1 and C-terminal location of SUMO-acceptor sites. Sequence alignment showing poor conservation at the C-termini of C. elegans HDA-1 and Homo sapiens HDAC1. The human HDAC1 C-terminus possesses two consensus SUMO-acceptor sites, K444 and K476 (acceptor lysines in red; consensus SUMO acceptor motif in pink box). GPS-SUMO predicts two candidate non-consensus SUMOylation sites in HDA-1, both near the C-terminus, K444 and K459 (red lysines in green boxes). (B) Western blot analyses of HDA-1 and MRG-1 before (lanes 1–5) and after (lanes 6–10) affinity enrichment of SUMOylated proteins from wild-type (wt) or hda-1 SUMO acceptor-site mutants. SUMOylated proteins were enriched from worms expressing HIS10::SMO-1. Black arrowheads indicate SUMOylated HDA-1 and MRG-1; white arrowheads indicate unmodified forms of HDA-1 and MRG-1. Asterisks indicate non-specific bands. Additional higher forms (indicated by white star) were detected, suggesting Multi-monoSUMOylation or PolySUMOylation of HDA-1. (C) Analysis of piRNA-mediated silencing in SUMOylation-defective mutants and rescue by HDA-1::SMO-1 translational fusion. (Top) The color of each bar indicates the percentage of worms in which the piRNA sensor was silent (OFF, gray) or expressed (ON, green). Thirty (n = 30) worms of each genotype were examined. (Bottom) Differential interference contrast and epifluorescence images of dissected gonads from hda-1[KKRR] and hda-1[KKRR]::smo-1. (D) Western blots showing levels of HDA-1 and variants proteins expressed from the endogenous hda-1 locus. Tubulin was used as a loading control. (E) Brood size analysis of wt worms or HDA-1 SUMOylation-site mutants, and rescue by HDA-1::SMO-1 translational fusion. Worms were grown at 20°C or 25°C, as indicated. Statistical significance was determined by ordinary one-way ANOVA: *p<0.05; **p<0.01; ****p<0.0001; ns: not significant. (F) Mortal germline analyses of wt or HDA-1 SUMOylation-site mutant, and HDA-1::SMO-1 fusion worms. The wago-9/hrde-1 mutant has a severe mortal germline phenotype. Worms were passaged at 25°C for eight generations, and the average number of progeny from 10 individuals (n = 10) was determined at each generation. Error bars represent standard error of the mean (SEM).

SUMOylation of HDA-1 promotes its association with NuRD and other chromatin factors.

(A) Silver stained gel of proteins that co-immunoprecipitate with HDA-1::GFP. The indicated protein bands were excised from the gel and identified by mass spectrometry. (B) Scatter plot comparing the levels of proteins identified by mass spectrometry in HDA-1::GFP IPs from smo-1(RNAi) and wild-type (no RNAi) worms. The x axis shows the log value of spectral counts for each protein identified by IP-MS from wild-type worms. The y axis shows the log ratio of spectral counts for each protein in HDA-1::GFP IPs from smo-1(RNAi) vs. wild-type. (C) As in B, but comparing hda-1[KKRR]::gfp to hda-1[WT]::gfp. In (B) and (C), the spectral counts of HDA-1 (RED) were used to normalize between samples. A full list of the identified proteins is provided in Supplementary file 2. (D) Western blot analyses of proteins (indicated to left of blots) that associate with MEP-1::GTF (GFP IP) in hda-1[WT], hda-1[KKRR] and hda-1[KKRR]::smo-1 lysates. The detected proteins are indicated to the right (black arrowheads). The modified isoforms, HDA-1::SUMO and HDA-1::SUMO-UBIQUITIN are indicated with white arrow and white star, respectively. (E) Side-by-side comparison of HDA-1 isoforms detected in the HDA-1, SMO-1, and UBIQUITIN blots in (D). The black dot indicates an unknown HDA-1 isoform. The black star indicates an unknown SUMOylated protein.

Figure 4 with 1 supplement
HDA-1 SUMOylation is required for formation of germline heterochromatin.

(A, B) Immunofluorescence micrographs of (A) wild-type (wt) and (B) hda-1[KKRR] gonads stained with anti-H3K9me2 antibody and DAPI. The dashed boxes indicated ‘a’ and ‘b’ are enlarged as shown. Two representative gonads are shown for each strain. (C) Differential interference contrast and immunofluorescence micrographs of gonads from wt, hda-1[KKRR], mep-1::gfp::degron with auxin (50 µM) worms stained with anti-H3K9Ac antibody and DAPI. (D) Genome Browser tracks (Integrated Genomics Viewer [IGV]) showing ChIP-seq peaks for NuRD complex components (HDA-1, LET-418, and MEP-1) and three histone modifications (H3K9Ac, H3K9me2, and H3K9me3) along each C. elegans chromosome (I–V and X). MEP-1(gonad) data are from worms that express MEP-1::GTF only in the germline, using the wago-1 promoter, for germline-specific CHIP.

Figure 4—figure supplement 1
Increased levels of active H3K4me3 chromatin mark in hda-1[KKRR].

(A, B) Immunofluorescence micrographs of anti-H3K4me3 and DAPI staining in three adult gonads of (A) wild-type (wt) and (B) hda-1[KKRR].

Figure 5 with 5 supplements
The SUMO, NuRD, and piRNA pathways regulate the same group of targets.

(A) Schematic of mRNA-seq from dissected gonads. (B) Venn diagram showing overlap between upregulated genes in degron::hda-1, mep-1::degron, ubc-9(G56R), and hda-1[KKRR] germlines. Numbers in parentheses indicate total number of upregulated genes. (C) Scatter plot of upregulated genes in hda-1[KKRR]. The x-axis represents reads in wild-type (wt), and y-axis represents reads in hda-1[KKRR]. (D) Scatter plot showing the effect of hda-1[KKRR]::smo-1 on the 430 genes (from C) upregulated in hda-1[KKRR]. The x-axis represents reads in wt, and y-axis represents reads in hda-1[KKRR]::smo-1. (E) Venn diagram showing overlap between upregulated genes in hda-1[KKRR], prg-1, rde-3, and wago-9 mutants. (F) Bar graph showing fractions of upregulated genes involved in spermatogenesis, oogenesis, neutral, or other categories. ‘Other’ indicates genes that cannot be put into one of the other categories (Ortiz et al., 2014). Genes with >1 mRNA-seq reads in wt gonad were used to generate the ‘wild-type’ dataset as a reference. The number of genes in each dataset is labeled at the top. (G–I) Scatter plots comparing mRNA-seq reads in (G) hda-1[KKRR], (H) rde-3, and (I) prg-1(ne4766) to those in wt. The blue dashed lines indicate a twofold increase or decrease in mutant compared to wt.

Figure 5—figure supplement 1
Comparison of replicates in gonad mRNA-seq.

(A) Scatter plots comparing two independent replicates of mRNA-seq data in the indicated strains. Replicate 1 is plotted on the x-axes and replicate 2 on the y-axes. The dashed blue lines indicate a twofold change. (B) Bar graphs showing number of upregulated (green) and downregulated (orange) genes that change at least twofold in the mutants germline (p≤0.05). (C) Venn diagram showing the overlap of upregulated genes in degron::hda-1, prg-1, rde-3, and wago-9. Total number of the upregulated genes in each mutant is indicated in the parentheses.

Figure 5—figure supplement 2
Analysis of transposons.

(A) Bar graphs showing number of upregulated (green) or downregulated (orange) transposons that change at least twofold in the gonad mRNA-seq of the indicated mutants (p≤0.05). (B) Scatter plots displaying upregulated transposons in the indicated mutants. The x-axis shows log10 value of average read counts in repeats of control (WT), and the y-axis shows log10 value of read counts in the indicated mutants. Transposons that are increased at least twofold and adjusted p-value ≤0.05 are colored in red. Transposon families are labeled if available. WT was used as a control for all the mutants except degron::hda-1 and ubc-9(ne4446[G56R]). For degron::hda-1 with 100 µM auxin exposure, worms without auxin exposure were used as control, and for ubc-9(ne4446[G56R]);oma-1::gfp; gfp::csr-1(AE), oma-1::gfp; gfp::csr-1(AE) was used as control. The dashed lines indicate twofold changes.

Figure 5—figure supplement 3
Loss of HDA-1 SUMOylation leads to depletion of H3k9me2 on a selected set of genes.

(A) Box plots show levels of H3K9me2 on HDA-1(KKRR) targets (upregulated in mRNA-seq) in the indicated strains. (B–E) Examples of H3K9me2 peaks (ChIP-seq reads) over protein-coding genes (B, C) and transposons (D , E). Read counts (y-axis) were normalized by total mapped reads in each sample.

Figure 5—figure supplement 4
Upregulated spermatogenic genes in the mutants.

(A) Scatter plots comparing mRNA-seq reads in mutant gonads to those in wild-type. Axes show log10 values of average reads from replicates of control (x-axis) and the indicated mutant (y-axis). Wild-type gonads were used as control for all the mutants except degron::hda-1 and ubc-9(ne4446[G56R]). Worms without auxin exposure were used as control for degron::hda-1, and oma-1::gfp; gfp::csr-1(AE) was used as a control for ubc-9(ne4446[G56R]);oma-1::gfp; gfp::csr-1(AE). (B) Scatter plots comparing mRNA-seq reads in rde-3 vs. hda-1[KKRR], prg-1 vs. hda-1[KKRR], and prg-1 vs. rde-3. Dashed lines indicate a twofold change. A value of 0.1 was assigned to undetected genes.

Figure 5—figure supplement 5
Analysis of small RNAs targeting upregulated genes in hda-1[KKRR].

(A) Scatter plots comparing anti-sense small RNA reads targeting each gene in hda-1[KKRR] and wild-type. The upregulated genes in hda-1[KKRR] from mRNA-seq are indicated in red. Dashed lines represent a twofold change. (B) Venn diagram showing the overlap between upregulated genes in hda-1[KKRR] from mRNA-seq and genes with increased or decreased small RNAs in hda-1[KKRR]. The total number of genes in each category is indicated in the parentheses.

HDA-1 interacts with WAGO-9/HRDE-1, which requires HDA-1 SUMOylation.

(A) Western blot analysis showing that HDA-1 and HPL-2 co-immunoprecipitate with GFP::WAGO-9/HRDE-1. (B) Western blot analysis of WAGO-9 and HPL-2 in HDA-1::GFP immunoprecipitates from wild-type, HDA-1 SUMO acceptor-site mutant, or smo-1(RNAi) worms. Blotting with anti-SMO-1 antibody showed depletion of SMO-1 in the smo-1(RNAi) worms. A band the size of untagged HDA-1 (black arrowhead) in hda-1[KKRR]::gfp in input appears to be a cleavage product that removes the GFP tag. (C) Graph showing the levels of silencing induced by RNAi over three generations in wild-type, hda-1[KKRR], and hda-1[KKRR]::smo-1 worms. Worms were treated with gfp(RNAi) at P0, and F1 larvae were transferred to regular NGM plates. The percentage of worms that express OMA-1::GFP was scored (n≥22, two replicates). Error bars represent standard error of the mean (SEM).

Model: HDAC1 SUMOylation promotes Argonaute-directed transcriptional gene silencing.

SUMOylation of HDA-1 enables nucleosome remodeling and deacetylase (NuRD) complex assembly in the adult germline and WAGO-9 or other Argonautes recruit the NuRD complex to piRNA targets. The Argonaute/NuRD complex, along with other histone-modifying enzymes—for example, SPR-5, MET-2—removes the active histone marks (H3K9Ac and H3K4me2/3) and establishes silencing marks (H3K9me2/3) to suppress their transcription.


Appendix 1—key resources table
Reagent type (species) or resourceDesignationSource or referenceIdentifiersAdditional information
AntibodyMouse monoclonal anti-FLAG M2Sigma-AldrichCat# F1804; RRID:AB_262044IB(1:1000)
AntibodyRabbit polyclonal anti-GFPGenScriptCat# A01704; RRID:AB_2622199IB(1:1000)
AntibodyRabbit polyclonal anti-MRG-1Novus BiologicalsCat# 49130002; RRID:AB_10011724IB(1:1000)
AntibodyRabbit polyclonal anti-HPL-1Novus BiologicalsCat# 38620002;
AntibodyRabbit polyclonal anti-HPL-2Novus BiologicalsCat# 38630002;
AntibodyRabbit polyclonal anti-LIN-53Novus BiologicalsCat# 48710002;
AntibodyRabbit polyclonal anti-HDA-1Novus BiologicalsCat# 38660002;
AntibodyRabbit monoclonal anti-tubulinCell Signaling TechnologyCat# 9099, RRID:AB_10695471IB(1:2000)
AntibodyRabbit polyclonal anti-LET-418Novus BiologicalsCat# 48960002, RRID:AB_10708820IB(1:1000)
AntibodyMouse monoclonal anti-SMO-1Pelisch et al., 2014Gift from Ronald T HayIB(1:500) freshly purified from hybridoma cell culture
AntibodyRabbit polyclonal anti-ubiquitinAbcamCat# ab7780; RRID:AB_306069IB(1:1000)
AntibodyRabbit polyclonal anti-HRDE-1/WAGO-9Ashe et al., 2012Gift from Eric A MiskaIB(1:500)
AntibodyGoat anti-mouse IgG (HRP-conjugated)Thermo Fisher ScientificCat# 62-6520; RRID:AB_2533947IB(1:2500)
AntibodyMouse Anti-rabbit IgG light (HRP-conjugated)AbcamCat# ab99697; RRID:AB_10673897IB(1:3000)
AntibodyMouse monoclonal anti-histone H3, di methyl K9AbcamCat# ab1220;
AntibodyMouse monoclonal anti-histone H3, acetyl K9AbcamCat# ab12179; RRID:AB_298910IF(1:100)
AntibodyRabbit polyclonal anti-trimethyl histone H3, K4MilliporeCat# 07-473; RRID:AB_1977252IF(1:100)
AntibodyGoat anti-mouse IgG (H+L) Alexa Fluor 488Thermo Fisher ScientificCat# A-11001; RRID:AB_2534069IF(1:1000)
AntibodyGoat anti-mouse IgG (H+L) Alexa Fluor 594Thermo Fisher ScientificCat# A-11005; RRID:AB_2534073IF(1:1000)
AntibodyGoat anti-rabbit IgG (H+L) Alexa Fluor 568Thermo Fisher ScientificCat# A-11011; RRID:AB_143157IF(1:1000)
AntibodyMouse monoclonal anti-FLAGSigma-AldrichCat# F3165; RRID:AB_259529For CHIP
AntibodyMouse monoclonal anti-dimethyl histone H3, K9MBL internationalCat# MABI0307; RRID:AB_11124951For CHIP
AntibodyRabbit polyclonal anti-histone H3, trimethyl K9MilliporeCat# 07-523; RRID:AB_310687For CHIP
AntibodyRabbit polyclonal anti-histone H3, acetyl K9AbcamCat# ab4441; RRID:AB_2118292For CHIP
Strain, strain backgroundC. elegans strainsThis studySupplementary file 3
Strain, strain backgroundE. coli: Strain OP50Caenorhabditis Genetics CenterWormBase: OP50
Strain, strain backgroundE. coli: Strain HT115Caenorhabditis Genetics CenterWormBase: HT115
Strain, strain backgroundE. coli: Ahringer collectionLaboratory of C. MelloN/A
recombinant protein
Ex Taq DNA polymeraseTakaraCat# RR001C
recombinant protein
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recombinant protein
Alt-R S.p. Cas9 Nuclease V3Integrated DNA Technologies (IDT)Cat# 1081058CRISPR reagent
recombinant protein
Alt-R A.s. Cas12a (Cpf1) V3Integrated DNA Technologies (IDT)Cat# 1081068CRISPR reagent
recombinant protein
GFP-binding protein (GBP) beadsHomemadeN/A
Peptide, recombinant proteinHybridase Thermostable RNase HLucigen CorporationCat# H39500
Peptide, recombinant proteinTurbo DNaseThermo Fisher ScientificCat# AM2238
Peptide, recombinant proteinSuper Script III Reverse TranscriptaseThermo Fisher ScientificCat# 18080085
Peptide, recombinant proteinT4 RNA ligase 1New England BiolabsCat# M0437M
Peptide, recombinant proteinT4 RNA ligase 2New England BiolabsCat# M0242L
Peptide, recombinant proteinSuper Script III Reverse TranscriptaseThermo Fisher ScientificCat# 18080085
Chemical compound, drugEthidium bromideSigma-AldrichCat# E1510
Chemical compound, drugIsopropyl-β -D-thiogalactoside (IPTG)Sigma-AldrichCat# 11411446001
Chemical compound, drugAmpicillinSigma-AldrichCat# A9518
Chemical compound, drugTetracyclineSigma-AldrichCat# 87128
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Chemical compound, drugTetramisole hydrochlorideSigma-AldrichCat# L9756-5G
Chemical compound, drugParaformaldehyde 16% solutionElectron Microscopy ScienceCat# Nm15710
Chemical compound, drugFormaldehyde, 36.5–38% in H2OSigma-AldrichCat# F8775
Chemical compound, drugPBSLife TechnologiesCat# AM9615
Chemical compound, drugTween20Fisher BioReagentsCat# BP337-500
Chemical compound, drugBovine serum albumin (BSA)Life TechnologiesCat# AM2618
Chemical compound, drug1M HEPES, pH7.4TEKnovaCat# H1030
Chemical compound, drugSodium citrate dihydrateThermo Fisher ScientificCat# BP337500
Chemical compound, drugTriton X-100Sigma-AldrichCat# T8787-250ml
Chemical compound, drugcOmplete EDTA-free Protease Inhibitor CocktailRocheCat# 11836170001
Chemical compound, drugNP-40EMD MilliporeCat# 492018
Chemical compound, drugTris (Base)AvantorCat# 4099–06
Chemical compound, drugEthylenediaminetetraacetic acid disodium salt dihydrateSigma-AldrichCat# E1644
Chemical compound, drugTE buffer, pH 8.0Thermo Fisher ScientificCat# AM9858
Chemical compound, drugSodium dodecyl sulfate (SDS)Sigma-AldrichCat# L3771-100G
Chemical compound, drugSodium chloride (NaCl)Genesee ScientificCat# 18-214
Chemical compound, drugMagnesium chloride (MgCl2)Sigma-AldrichCat# M8266
Chemical compound, drugDL-Dithiothreitol (DTT)Sigma-AldrichCat# D0632-10G
Chemical compound, drugCalcium chloride (CaCl2)Sigma-AldrichCat# C5080
Chemical compound, drugPotassium chloride (KCl)Sigma-AldrichCat# P9541
Chemical compound, drugGuanidine-HClSigma-AldrichCat# G3272
Chemical compound, drugImidazoleSigma-AldrichCat# 792527
Chemical compound, drugβ-MercaptoethanolSigma-AldrichCat# M6250
Chemical compound, drugSodium phosphate, dibasicSigma-AldrichCat# S7907
Chemical compound, drugSodium phosphate, monobasicSigma-AldrichCat# S0751
Chemical compound, drugUreaThermo Fisher ScientificCat# Ac327380010
Chemical compound, drugTrichloroacetic acid (TCA)Sigma-AldrichCat# T0699
Chemical compound, drug1-Bromo-3-chloropropaneSigma-AldrichCat# B9673
Chemical compound, drugGlycineThermo Fisher ScientificCat# BP381-1
Chemical compound, drugTRI reagentSigma-AldrichCat# T9424
Chemical compound, drugTrypsinNew England BiolabsCat# P8101S
Commercial assay, kitSlowFade Diamond antifade Mountant with DAPILife TechnologiesCat# S36964
Commercial assay, kitQuick start Bradford 1xdye reagentBio-RadCat# 5000205
Commercial assay, kitNuPAGE LDS sample buffer (4x)Thermo Fisher ScientificCat# NP0008
Commercial assay, kitGlycoBlue CoprecipitantThermo Fisher ScientificCat# AM9515
Commercial assay, kitNi-NTA resinQiagenCat# 30210
Commercial assay, kitpCR-Blunt II Topo cloning kitThermo Fisher ScientificCat# K280020
Commercial assay, kitMinElute PCR purification KitQiagenCat# 28006
Commercial assay, kitChIP DNA clean and concentrator KitZymo ResearchCat# 5205
Commercial assay, kitProteoSilver Plus Silver Stain KitSigma-AldrichCat# PROT-SIL2
Commercial assay, kitTrans-blot Turbo Mini NC Transfer PacksBio-RadCat# 1704158
Commercial assay, kitLumi-Light Plus western blotting substrateSigma-AldrichCat# 12015196001
Commercial assay, kitHyperfilm ECLThermo Fisher ScientificCat# 45001507
Commercial assay, kitmirVana miRNA Isolation KitThermo Fisher ScientificCat# AM1561
Commercial assay, kitKAPA RNA HyperPrep with RiboErase (KK8560)RocheCat# 08098131702
Commercial assay, kitKAPA single-indexed adapter kit (KK8700)RocheCat# 08005699001
Commercial assay, kitChIP-Grade Protein A/G Magnetic BeadsThermo Fisher ScientificCat# 26162
Commercial assay, kitIllumina NextSeq 500/550 v2.5 kit (75 cycles)Illumina, Inc.Cat# 20024906
Commercial assay, kitIllumina NextSeq 500/550 v2.5 kit (150 cycles)Illumina, Inc.Cat# 20024907
Recombinant DNA reagentPeft3::cas9 vector (backbone: blunt II topo vector)Friedland et al., 2013N/ABackbone: blunt II topo vector (Kim et al., 2021)
Recombinant DNA reagentpRF4: injection marker, rol-6(su1006)Mello et al., 1991N/ABackbone: blunt II topo vector (Kim et al., 2021)
Recombinant DNA reagentsmo-1 sgRNA plasmidThis studySupplementary file 4
Recombinant DNA reagentmep-1 sgRNA plasmidKim et al., 2021Supplementary file 4
Sequence-based reagentList of gRNA sequencesThis studySupplementary file 4
Sequence-based reagentAlt-R CRISPR-Cas9 tracrRNAIntegrated DNA Technologies (IDT)Cat# 1072534CRISPR reagent
Sequence-based reagentAnti-rRNA Oligos for C. elegansThis study (homemade)N/A
Software, algorithmGraphPad Prism version 8.2.1GraphPad Softwarehttp://www.graphpad.com
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Software, algorithmDEseq2Love et al., 2014v1.26.0
Software, algorithmBowtie2Langmead and Salzberg, 2012v2.2.3
Software, algorithmPicard-toolsBroad Institute, 2019v1.131
Software, algorithmMACS2Feng et al., 2012v2.1.1.20160309
Software, algorithmBedToolsQuinlan and Hall, 2010v2.25.0
Software, algorithmIGVRobinson et al., 2017v2.7.2

Additional files

Supplementary file 1

Summary of RNAi-based genetic screen of chromatin factors and modifiers using a silenced piRNA sensor.

Supplementary file 2

HDA-1 interactors identified from HDA-1::GFP immunoprecipitation-mass spectrometry (IP-MS).

Supplementary file 3

List of C. elegans strains used in this study.

Supplementary file 4

List of gRNA and ssOligo donor sequences.

Supplementary file 5

RNA-seq data (deposited to Bioproject: PRJNA657279).

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  1. Heesun Kim
  2. Yue-He Ding
  3. Gangming Zhang
  4. Yong-Hong Yan
  5. Darryl Conte Jr
  6. Meng-Qiu Dong
  7. Craig C Mello
HDAC1 SUMOylation promotes Argonaute-directed transcriptional silencing in C. elegans
eLife 10:e63299.