The transcription factor Krox20 is an E3 ligase that sumoylates its Nab coregulators

García-Gutiérrez, Pablo; Juárez-Vicente, Francisco; Gallardo-Chamizo, Francisco; Charnay, Patrick; García-Domínguez, Mario

doi:10.1038/embor.2011.152

Cited by 30 publications

(19 citation statements)

References 20 publications

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“…P19 cells were cultured in α-modified Eagle's medium supplemented with 7.5% (vol/vol) calf and 2.5% (vol/vol) fetal bovine sera. Sumoylation assays in cells and purification of endogenous sumoylated proteins from 293T cells were performed as described (35). In ovo electroporation and embryo immunofluorescence were carried out as described (33).…”

Section: Methodsmentioning

confidence: 99%

Control of neuronal differentiation by sumoylation of BRAF35, a subunit of the LSD1-CoREST histone demethylase complex

Ceballos-Chávez

Rivero

García-Gutiérrez³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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The LSD1-CoREST histone demethylase complex is required to repress neuronal genes in nonneuronal tissues. Here we show that sumoylation of Braf35, one of the subunits of the complex, is required to maintain full repression of neuron-specific genes and for occupancy of the LSD1-CoREST complex at its gene targets. Interestingly, expression of Braf35 was sufficient to prevent neuronal differentiation induced by bHLH neurogenic transcription factors in P19 cells and in neuronal progenitors of the chicken embryo neural tube. Sumoylation of Braf35 is required for this antineurogenic activity. We also show that iBraf, a paralogue of Braf35, forms heterodimers with Braf35. Braf35-iBraf heterodimerization impairs Braf35 interaction with the LSD1-CoREST complex and inhibits Braf35 sumoylation. Consistent with these results, iBraf prevents the antineurogenic activity of Braf35 in vivo. Our data uncover a mechanism of regulation of the LSD1-CoREST complex and provide a molecular explanation for the antagonism between Braf35 and iBraf in neuronal differentiation.C ell differentiation involves large modifications of gene expression that require extensive changes of chromatin epigenetic marks (1). Epigenetic marks are DNA or histone posttranslational modifications that are inherited through cell division and that inform about the transcriptional state of loci. Among histone modifications, histone lysine methylation is of particular interest in development for the broad range of processes in which it is involved, including maintenance of stem cell pluripotency, germ-line determination, cell differentiation, control of HOX genes expression, and so forth (2, 3). Histone lysine methylation was considered a stable posttranslational modification until the discovery of histone demethylases. LSD1/KDM1 (lysine-specific demethylase 1) was the first demethylase identified and catalyzes demethylation of both di-and monomethylated lysine 4 (K4) or lysine 9 (K9) of histone H3 (H3K4me2/1 or H3K9me2/1) (4, 5). The lysine specificity of LSD1 seems to depend on its molecular partners. Thus, when LSD1 is associated with CoREST in the LSD1-CoREST corepressor complex (also called BHC, BRAF-histone deacetylase complex), the preferred substrate is H3K4me2/1, consistent with the fact that methylation of H3K4 is a mark of transcriptionaly active genes. In addition to LSD1 and CoREST, the LSD1-CoREST complex also contains HDAC1-2, BHC80, and BRAF35 (also called HMG20B) (6-9). BRAF35 contains a high-mobility group (HMG) domain and a coiled-coil domain, but its function within the complex is not well-understood (7, 10). Several functions of the LSD1-CoREST complex in differentiation and development have been reported (2). One of the best-characterized functions of the complex is its role in repression of neuronal genes in nonneuronal tissues and neuronal progenitors through its interaction with repressor factor REST (RE1 silencing transcription factor) (11,12). iBRAF (inhibitor of BRAF35, also called HMG20A) is a close paralogue of BRAF35 (13). As BRAF35...

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Section: Methodsmentioning

confidence: 99%

Control of neuronal differentiation by sumoylation of BRAF35, a subunit of the LSD1-CoREST histone demethylase complex

Ceballos-Chávez

Rivero

García-Gutiérrez³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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“…Other TFs, such as Pbx/Meis, are also essential for hindbrain patterning via their synergistic activities with Hox factors (Aamar and Frank, 2004;Elkouby et al, 2012;Vlachakis et al, 2001;Wassef et al, 2008). Additional factors also modulate the expression of these TFs, such as Nab and Nlz (also known as Neurl1a), which colocalize with Krox20 but repress its transcription, leading to Krox20 restriction to the correct rhombomeres (García-Gutiérrez et al, 2011;Mechta-Grigoriou et al, 2000;Runko and Sagerström, 2003). Finally, Eph-ephrin signaling acts at the rhombomere interface and restricts cell intermixing by mediating repulsion.…”

Section: Introductionmentioning

confidence: 99%

A novel role for Pax6 in the segmental organization of the hindbrain

et al. 2013

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SUMMARYComplex patterns and networks of genes coordinate rhombomeric identities, hindbrain segmentation and neuronal differentiation and are responsible for later brainstem functions. Pax6 is a highly conserved transcription factor crucial for neuronal development, yet little is known regarding its early roles during hindbrain segmentation. We show that Pax6 expression is highly dynamic in rhombomeres, suggesting an early function in the hindbrain. Utilization of multiple gain-and loss-of-function approaches in chick and mice revealed that loss of Pax6 disrupts the sharp expression borders of Krox20, Kreisler, Hoxa2, Hoxb1 and EphA and leads to their expansion into adjacent territories, whereas excess Pax6 reduces these expression domains. A mutual negative cross-talk between Pax6 and Krox20 allows these genes to be co-expressed in the hindbrain through regulation of the Krox20-repressor gene Nab1 by Pax6. Rhombomere boundaries are also distorted upon Pax6 manipulations, suggesting a mechanism by which Pax6 acts to set hindbrain segmentation. Finally, FGF signaling acts upstream of the Pax6-Krox20 network to regulate Pax6 segmental expression. This study unravels a novel role for Pax6 in the segmental organization of the early hindbrain and provides new evidence for its significance in regional organization along the central nervous system.

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“…The RING finger protein TOPORS (topoisomerase I-binding, arginine/serine-rich, E3 ubiquitin protein ligase), a ubiquitin ligase for p53, was also found as a Ubc9-and SUMO-interacting protein that enhances the SUMOylation of p53 as well as other chromatin-associated proteins in a RING-independent manner [28,29]. Other examples include the RING finger proteins MAPL (mitochondria-anchored protein ligase) [30], CCNB1IP1 (cyclin B1-interacting protein 1, E3 ubiquitin protein ligase) [31] and the TRIM (tripartite motif-containing) family [32], the tumour suppressors p14 Arf (ADP-ribosylation factor) [33,34] and TLS (translocated in liposarcoma)/FUS (fused in sarcoma) [35], the histone deacetylase HDAC7 (histone deacetylase 7) [36], the RNA-binding protein RAX/PACT (protein activator of the interferon-induced protein kinase) [37], the transcription factor Krox20 [38], and the transcriptional co-repressor KAP1 (KRABassociated protein 1) [39]. Many of these were initially identified as interacting with Ubc9, although in most cases an affinity for SUMO itself has not been tested.…”

Section: Discussionmentioning

confidence: 99%

SIM-dependent enhancement of substrate-specific SUMOylation by a ubiquitin ligase in vitro

Parker

Ulrich

2014

Biochemical Journal

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SIMs (SUMO-interaction motifs), which mediate the non-covalent binding of SUMO (small ubiquitin-related modifier) to other proteins, are usually involved in the recognition of SUMOylated substrates by downstream effectors that transmit the biological signal of the modification. In ubiquitin ligase Rad18 (radiation-sensitive 18) from Saccharomyces cerevisiae, a SIM, contributes to the recognition of SUMOylated PCNA (proliferating-cell nuclear antigen) as its physiological ubiquitylation target. In the present study we show that Rad18 is also capable of enhancing PCNA SUMOylation in a SIM-dependent manner in vitro, most probably by means of directing SUMO-loaded Ubc9 (ubiquitin-conjugating enzyme 9) towards the substrate. The process shares important features with Rad18-dependent ubiquitylation, such as an exquisite specificity for the modification site on PCNA and the requirement of DNA, and the reaction proceeds under conditions that are widely used in other in vitro assays for SUMO ligase activity. However, there is no evidence that Rad18 contributes to PCNA SUMOylation in vivo. The findings of the present study therefore illustrate the problematic nature of in vitro SUMOylation assays and highlight the danger of extrapolating from this type of experiment to the biological function of a SUMO-interacting protein.

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The transcription factor Krox20 is an E3 ligase that sumoylates its Nab coregulators

Cited by 30 publications

References 20 publications

Control of neuronal differentiation by sumoylation of BRAF35, a subunit of the LSD1-CoREST histone demethylase complex

Control of neuronal differentiation by sumoylation of BRAF35, a subunit of the LSD1-CoREST histone demethylase complex

A novel role for Pax6 in the segmental organization of the hindbrain

SIM-dependent enhancement of substrate-specific SUMOylation by a ubiquitin ligase in vitro

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