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The oncoprotein BCL11A binds to orphan nuclear receptor TLX and potentiates its transrepressive function.

Estruch SB, Buzón V, Carbó LR, Schorova L, Lüders J, Estébanez-Perpiñá E - PLoS ONE (2012)

Bottom Line: This interaction was validated by expression and coimmunoprecipitation in human cells.BCL11A potentiated the transrepressive function of TLX in an in vitro reporter gene assay.Our work suggests that BCL11A is a novel TLX coregulator that might be involved in TLX-dependent gene regulation in the brain.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology and Institute of Biomedicine from the University of Barcelona, University of Barcelona, Barcelona, Spain.

ABSTRACT
Nuclear orphan receptor TLX (NR2E1) functions primarily as a transcriptional repressor and its pivotal role in brain development, glioblastoma, mental retardation and retinopathologies make it an attractive drug target. TLX is expressed in the neural stem cells (NSCs) of the subventricular zone and the hippocampus subgranular zone, regions with persistent neurogenesis in the adult brain, and functions as an essential regulator of NSCs maintenance and self-renewal. Little is known about the TLX social network of interactors and only few TLX coregulators are described. To identify and characterize novel TLX-binders and possible coregulators, we performed yeast-two-hybrid (Y2H) screens of a human adult brain cDNA library using different TLX constructs as baits. Our screens identified multiple clones of Atrophin-1 (ATN1), a previously described TLX interactor. In addition, we identified an interaction with the oncoprotein and zinc finger transcription factor BCL11A (CTIP1/Evi9), a key player in the hematopoietic system and in major blood-related malignancies. This interaction was validated by expression and coimmunoprecipitation in human cells. BCL11A potentiated the transrepressive function of TLX in an in vitro reporter gene assay. Our work suggests that BCL11A is a novel TLX coregulator that might be involved in TLX-dependent gene regulation in the brain.

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The oncoprotein BCL11A binds to human TLX.(A) Schematic diagram of the different isoforms of BCL11A (XL, L, S) and corresponding exons (1, E2, E3, E4, E5L, E5S). All BCL11A identified clones share a 159 amino acid overlapping region (residues 586–744), whose amino acid sequence is shown. Previously BCL11A regions identified to bind to COUP-FT and named ID-1 and ID-2 are highlighted in light grey. Clone 1 features a novel combination of exons, while clones 3, and 4 contain an intronic sequence. All the identified clones contain ID-2, but only some of them also have ID-2 as well. (B) Validation of the interaction between TLX and the identified BCL11A clones by forward one-to-one Y2H assay. TLX constructs FL-TLX (1–385), TLX-LBD (172–385), TLX-H-LBD (94–385) and TLX-DBD (1–95) (baits) were tested for interaction with the BCL11A clones identified (preys). Yeast transformants were plated on a control plate (lacking Trp and Leu) and plated on a selective plate (lacking Trp, Leu, His supplemented with 50 mM 3AT).
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pone-0037963-g002: The oncoprotein BCL11A binds to human TLX.(A) Schematic diagram of the different isoforms of BCL11A (XL, L, S) and corresponding exons (1, E2, E3, E4, E5L, E5S). All BCL11A identified clones share a 159 amino acid overlapping region (residues 586–744), whose amino acid sequence is shown. Previously BCL11A regions identified to bind to COUP-FT and named ID-1 and ID-2 are highlighted in light grey. Clone 1 features a novel combination of exons, while clones 3, and 4 contain an intronic sequence. All the identified clones contain ID-2, but only some of them also have ID-2 as well. (B) Validation of the interaction between TLX and the identified BCL11A clones by forward one-to-one Y2H assay. TLX constructs FL-TLX (1–385), TLX-LBD (172–385), TLX-H-LBD (94–385) and TLX-DBD (1–95) (baits) were tested for interaction with the BCL11A clones identified (preys). Yeast transformants were plated on a control plate (lacking Trp and Leu) and plated on a selective plate (lacking Trp, Leu, His supplemented with 50 mM 3AT).

Mentions: When we analyzed the remaining clones, we identified the oncoprotein BCL11A as a novel interactor of TLX (Figure 2A and Text S1-B). We obtained several overlapping clones of BCL11A in independent screens using both baits (FL-TLX and TLX-LBD). All BCL11A clones shared a region comprising residues 586 to 744 (Figure 2A and Text S1-B). Secondary structure predictions indicate that this fragment might acquire an α-helical structure flanked by two short ß-sheets. This region contained the COUP-TFII interaction domain ID1 (Figure 2A) [27]. A second BCL11A domain that was described to interact with COUP-TFII and termed ID2 (residues 264–378) [27]–[28], was only present in some of the TLX-interacting clones (Figure 2A).


The oncoprotein BCL11A binds to orphan nuclear receptor TLX and potentiates its transrepressive function.

Estruch SB, Buzón V, Carbó LR, Schorova L, Lüders J, Estébanez-Perpiñá E - PLoS ONE (2012)

The oncoprotein BCL11A binds to human TLX.(A) Schematic diagram of the different isoforms of BCL11A (XL, L, S) and corresponding exons (1, E2, E3, E4, E5L, E5S). All BCL11A identified clones share a 159 amino acid overlapping region (residues 586–744), whose amino acid sequence is shown. Previously BCL11A regions identified to bind to COUP-FT and named ID-1 and ID-2 are highlighted in light grey. Clone 1 features a novel combination of exons, while clones 3, and 4 contain an intronic sequence. All the identified clones contain ID-2, but only some of them also have ID-2 as well. (B) Validation of the interaction between TLX and the identified BCL11A clones by forward one-to-one Y2H assay. TLX constructs FL-TLX (1–385), TLX-LBD (172–385), TLX-H-LBD (94–385) and TLX-DBD (1–95) (baits) were tested for interaction with the BCL11A clones identified (preys). Yeast transformants were plated on a control plate (lacking Trp and Leu) and plated on a selective plate (lacking Trp, Leu, His supplemented with 50 mM 3AT).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3366998&req=5

pone-0037963-g002: The oncoprotein BCL11A binds to human TLX.(A) Schematic diagram of the different isoforms of BCL11A (XL, L, S) and corresponding exons (1, E2, E3, E4, E5L, E5S). All BCL11A identified clones share a 159 amino acid overlapping region (residues 586–744), whose amino acid sequence is shown. Previously BCL11A regions identified to bind to COUP-FT and named ID-1 and ID-2 are highlighted in light grey. Clone 1 features a novel combination of exons, while clones 3, and 4 contain an intronic sequence. All the identified clones contain ID-2, but only some of them also have ID-2 as well. (B) Validation of the interaction between TLX and the identified BCL11A clones by forward one-to-one Y2H assay. TLX constructs FL-TLX (1–385), TLX-LBD (172–385), TLX-H-LBD (94–385) and TLX-DBD (1–95) (baits) were tested for interaction with the BCL11A clones identified (preys). Yeast transformants were plated on a control plate (lacking Trp and Leu) and plated on a selective plate (lacking Trp, Leu, His supplemented with 50 mM 3AT).
Mentions: When we analyzed the remaining clones, we identified the oncoprotein BCL11A as a novel interactor of TLX (Figure 2A and Text S1-B). We obtained several overlapping clones of BCL11A in independent screens using both baits (FL-TLX and TLX-LBD). All BCL11A clones shared a region comprising residues 586 to 744 (Figure 2A and Text S1-B). Secondary structure predictions indicate that this fragment might acquire an α-helical structure flanked by two short ß-sheets. This region contained the COUP-TFII interaction domain ID1 (Figure 2A) [27]. A second BCL11A domain that was described to interact with COUP-TFII and termed ID2 (residues 264–378) [27]–[28], was only present in some of the TLX-interacting clones (Figure 2A).

Bottom Line: This interaction was validated by expression and coimmunoprecipitation in human cells.BCL11A potentiated the transrepressive function of TLX in an in vitro reporter gene assay.Our work suggests that BCL11A is a novel TLX coregulator that might be involved in TLX-dependent gene regulation in the brain.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology and Institute of Biomedicine from the University of Barcelona, University of Barcelona, Barcelona, Spain.

ABSTRACT
Nuclear orphan receptor TLX (NR2E1) functions primarily as a transcriptional repressor and its pivotal role in brain development, glioblastoma, mental retardation and retinopathologies make it an attractive drug target. TLX is expressed in the neural stem cells (NSCs) of the subventricular zone and the hippocampus subgranular zone, regions with persistent neurogenesis in the adult brain, and functions as an essential regulator of NSCs maintenance and self-renewal. Little is known about the TLX social network of interactors and only few TLX coregulators are described. To identify and characterize novel TLX-binders and possible coregulators, we performed yeast-two-hybrid (Y2H) screens of a human adult brain cDNA library using different TLX constructs as baits. Our screens identified multiple clones of Atrophin-1 (ATN1), a previously described TLX interactor. In addition, we identified an interaction with the oncoprotein and zinc finger transcription factor BCL11A (CTIP1/Evi9), a key player in the hematopoietic system and in major blood-related malignancies. This interaction was validated by expression and coimmunoprecipitation in human cells. BCL11A potentiated the transrepressive function of TLX in an in vitro reporter gene assay. Our work suggests that BCL11A is a novel TLX coregulator that might be involved in TLX-dependent gene regulation in the brain.

Show MeSH
Related in: MedlinePlus