SUMOylation is essential for sex-specific assembly and function of the
            <i>Caenorhabditis elegans</i>
            dosage compensation complex on X chromosomes

Pferdehirt, Rebecca R.; Meyer, Barbara J

doi:10.1073/pnas.1315793110

Cited by 23 publications

(27 citation statements)

References 35 publications

(55 reference statements)

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“…The strategy permits the recovery of multiple classes of alleles, null, hypomorphic, hypermorphic, and antimorphic, in the same experiment. assembly of the DCC onto X chromosomes (Pferdehirt and Meyer 2013). To analyze in detail the role of SUMO in regulating the activities of these proteins, the lysine residues that serve as attachment sites for SUMO modifications must be identified.…”

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confidence: 99%

Precise and Heritable Genome Editing in Evolutionarily Diverse Nematodes Using TALENs and CRISPR/Cas9 to Engineer Insertions and Deletions

et al. 2013

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Exploitation of custom-designed nucleases to induce DNA double-strand breaks (DSBs) at genomic locations of choice has transformed our ability to edit genomes, regardless of their complexity. DSBs can trigger either error-prone repair pathways that induce random mutations at the break sites or precise homology-directed repair pathways that generate specific insertions or deletions guided by exogenously supplied DNA. Prior editing strategies using site-specific nucleases to modify the Caenorhabditis elegans genome achieved only the heritable disruption of endogenous loci through random mutagenesis by error-prone repair. Here we report highly effective strategies using TALE nucleases and RNA-guided CRISPR/Cas9 nucleases to induce error-prone repair and homology-directed repair to create heritable, precise insertion, deletion, or substitution of specific DNA sequences at targeted endogenous loci. Our robust strategies are effective across nematode species diverged by 300 million years, including necromenic nematodes (Pristionchus pacificus), male/female species (Caenorhabditis species 9), and hermaphroditic species (C. elegans). Thus, genome-editing tools now exist to transform nonmodel nematode species into genetically tractable model organisms. We demonstrate the utility of our broadly applicable genome-editing strategies by creating reagents generally useful to the nematode community and reagents specifically designed to explore the mechanism and evolution of X chromosome dosage compensation. By developing an efficient pipeline involving germline injection of nuclease mRNAs and single-stranded DNA templates, we engineered precise, heritable nucleotide changes both close to and far from DSBs to gain or lose genetic function, to tag proteins made from endogenous genes, and to excise entire loci through targeted FLP-FRT recombination. STRATEGIES to engineer heritable, site-directed mutations at endogenous loci have revolutionized our approach toward manipulating and dissecting genome function. Studies of plants and animals alike, whether conducted in whole organisms or cell lines, have benefitted greatly from these genomeediting approaches (Bibikova et al. 2002;Beumer et al. 2006;Doyon et al. 2008;Geurts et al. 2009;Hockemeyer et al. 2009;Holt et al. 2010;Zhang et al. 2010;Hockemeyer et al. 2011;Tesson et al. 2011;Wood et al. 2011;Young et al. 2011;Bedell et al. 2012;Bassett et al. 2013;Cong et al. 2013;Jinek et al. 2013;Mali et al. 2013;Wang et al. 2013;Zu et al. 2013). The most modern tools for modifying complex genomes at single-nucleotide resolution are site-specific nucleases that induce DNA double-strand breaks (DSBs) at specifically designated genomic locations. DSBs trigger repair pathways that can elicit targeted genetic reprogramming, primarily through two mechanisms: error-prone nonhomologous end joining (NHEJ) (Lieber 2010) and precise, homology-directed recombination or repair (HDR) (Chapman et al. 2012). NHEJ rejoins broken ends of chromosomes through an imprecise process that produces nucleo...

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Precise and Heritable Genome Editing in Evolutionarily Diverse Nematodes Using TALENs and CRISPR/Cas9 to Engineer Insertions and Deletions

et al. 2013

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“…Interestingly, Tup1 interaction partners histone H3, Cti6, Gcn5, Gal11, and Cyc8 also become SUMOylated under similar conditions as Tup1, and SUMO may generally hold together transcriptional repressor complexes in the same manner that SUMO protein functions to hold together DNA repair systems and PML nuclear bodies (Psakhye and Jentsch, 2012; Shen et al, 2006). In C. elegans , SUMOylation of multiple components of the dosage compensation complex is also required for complex assembly, but it is not required for initial targeting of the this complex to the X chromosome (Pferdehirt and Meyer, 2013). Our evidence that UNC-37 and SUMO pathway components regulate tbx-2 in a similar manner to TBX-2 is consistent with the notion that SUMO binding may hold together TBX-2 and the Groucho-like homolog UNC-37 in a repressor complex to regulate target genes.…”

Section: Discussionmentioning

confidence: 99%

Function of the C. elegans T-box factor TBX-2 depends on interaction with the UNC-37/Groucho corepressor

Huber

Crum

Okkema

2016

Developmental Biology

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T-box transcription factors are important regulators of development in all animals, and altered expression of T-box factors has been identified in an increasing number of diseases and cancers. Despite these important roles, the mechanism of T-box factor activity is not well understood. We have previously shown that the C. elegans Tbx2 subfamily member TBX-2 functions as a transcriptional repressor to specify ABa-derived pharyngeal muscle, and that this function depends on SUMOylation. Here we show that TBX-2 function also depends on interaction with the Groucho-family corepressor UNC-37. TBX-2 interacts with UNC-37 in yeast two-hybrid assays via a highly conserved engrailed homology 1 (eh1) motif located near the TBX-2 C-terminus. Reducing unc-37 phenocopies tbx-2 mutants, resulting in a specific loss of anterior ABa-derived pharyngeal muscles and derepression of the tbx-2 promoter. Moreover, double mutants containing hypomorphic alleles of unc-37 and tbx-2 exhibit enhanced phenotypes, providing strong genetic evidence that unc-37 and tbx-2 share common functions in vivo. To test whether interaction with UNC-37 is necessary for TBX-2 activity, we developed a transgene rescue assay using a tbx-2 containing fosmid and found that mutating the tbx-2 eh1 motif reduced rescue of a tbx-2 null mutant. These results indicate that TBX-2 function in vivo depends on interaction with UNC-37. As many T-box factors contain eh1 motifs, we suggest that interaction with Groucho-family corepressors is a common mechanism contributing to their activity.

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“…Robust X-chromosome specific binding of the DCC also requires sumoylation of several subunits, SDC-3, DPY-27 and DPY-28 [*26]. DPY-28 also functions in the context of condensin I, but it is only sumoylated when part of the DCC (Figure 2).…”

Section: The Dosage Compensation Complexmentioning

confidence: 99%

“…DPY-28 also functions in the context of condensin I, but it is only sumoylated when part of the DCC (Figure 2). Depletion of SUMO reduced X chromosome binding of condensin I DC , but not binding of SDC-2 and DPY-30, indicating that sumoylation is needed for stable interaction between X targeting proteins and other DCC members, rather than initial X recognition [*26]. Interestingly, SUMO depletion also leads to the appearance of novel DCC binding sites on autosomes [*26], suggesting that while SUMO may be required to restrict DCC binding to the X chromosome.…”

Section: The Dosage Compensation Complexmentioning

confidence: 99%

“…Depletion of SUMO reduced X chromosome binding of condensin I DC , but not binding of SDC-2 and DPY-30, indicating that sumoylation is needed for stable interaction between X targeting proteins and other DCC members, rather than initial X recognition [*26]. Interestingly, SUMO depletion also leads to the appearance of novel DCC binding sites on autosomes [*26], suggesting that while SUMO may be required to restrict DCC binding to the X chromosome. This latter interpretation is similar to a previous finding, where restriction of DCC binding to the X chromosome required the histone variant HTZ-1 [27].…”

Section: The Dosage Compensation Complexmentioning

confidence: 99%

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Balancing up and downregulation of the C. elegans X chromosomes

Lau

Csankovszki

2015

Current Opinion in Genetics & Development

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In Caenorhabditis elegans, males have one X chromosome and hermaphrodites have two. Emerging evidence indicates that the male X is transcriptionally more active than autosomes to balance the single X to two sets of autosomes. Because upregulation is not limited to males, hermaphrodites need to strike back and downregulate expression from the two X chromosomes to balance gene expression in their genome. Hermaphrodite-specific downregulation involves binding of the dosage compensation complex to both Xs. Advances in recent years revealed that the action of the dosage compensation complex results in compaction of the X chromosomes, changes in the distribution of histone modifications, and ultimately limiting RNA Polymerase II loading to achieve chromosome-wide gene repression.

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SUMOylation is essential for sex-specific assembly and function of the Caenorhabditis elegans dosage compensation complex on X chromosomes

Cited by 23 publications

References 35 publications

Precise and Heritable Genome Editing in Evolutionarily Diverse Nematodes Using TALENs and CRISPR/Cas9 to Engineer Insertions and Deletions

Precise and Heritable Genome Editing in Evolutionarily Diverse Nematodes Using TALENs and CRISPR/Cas9 to Engineer Insertions and Deletions

Function of the C. elegans T-box factor TBX-2 depends on interaction with the UNC-37/Groucho corepressor

Balancing up and downregulation of the C. elegans X chromosomes

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