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Transcriptional repression of Gata3 is essential for early B cell commitment.

ABSTRACT: The mechanisms underlying the silencing of alternative fate potentials in very early B cell precursors remain unclear. Using gain- and loss-of-function approaches together with a synthetic Zinc-finger polypeptide (6ZFP) engineered to prevent transcription factor binding to a defined cis element, we show that the transcription factor EBF1 promotes B cell lineage commitment by directly repressing expression of the T-cell-lineage-requisite Gata3 gene. Ebf1-deficient lymphoid progenitors exhibited increased T cell lineage potential and elevated Gata3 transcript expression, whereas enforced EBF1 expression inhibited T cell differentiation and caused rapid loss of Gata3 mRNA. Notably, 6ZFP-mediated perturbation of EBF1 binding to a Gata3 regulatory region restored Gata3 expression, abrogated EBF1-driven suppression of T cell differentiation, and prevented B cell differentiation via a GATA3-dependent mechanism. Furthermore, EBF1 binding to Gata3 regulatory sites induced repressive histone modifications across this region. These data identify a transcriptional circuit critical for B cell lineage commitment.

SUBMITTER: Banerjee A

PROVIDER: S-EPMC3664383 | biostudies-literature | 2013 May

REPOSITORIES: biostudies-literature

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Transcriptional repression of Gata3 is essential for early B cell commitment.

Banerjee Anupam A Northrup Daniel D Boukarabila Hanane H Jacobsen Sten Erik W SE Allman David D

Immunity 20130516 5

The mechanisms underlying the silencing of alternative fate potentials in very early B cell precursors remain unclear. Using gain- and loss-of-function approaches together with a synthetic Zinc-finger polypeptide (6ZFP) engineered to prevent transcription factor binding to a defined cis element, we show that the transcription factor EBF1 promotes B cell lineage commitment by directly repressing expression of the T-cell-lineage-requisite Gata3 gene. Ebf1-deficient lymphoid progenitors exhibited i ...[more]

PMID: 23684985

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Project description:Toxoplasma gondii is a widespread protozoan parasite that has a significant impact on human and veterinary health. The parasite undergoes a complex life cycle involving multiple hosts and developmental stages. How Toxoplasma transitions between life cycle stages is poorly understood yet central to controlling transmission. Of particular neglect are the factors that contribute to its sexual development, which takes place exclusively in feline intestines. While epigenetic repressors have been shown to play an important role in silencing the spurious gene expression of sexually committed parasites, the specific factors that recruit this generalized machinery to the appropriate genes remain largely unexplored. Here, we establish that a member of the AP2 transcription factor family, AP2XII-2, is targeted to genomic loci associated with sexually committed parasites along with epigenetic regulators of transcriptional silencing, HDAC3 and MORC. Despite its widespread association with gene promoters, AP2XII-2 is required for the silencing of relatively few genes. Using the CUT&Tag (cleavage under targets and tagmentation) methodology, we identify two major genes associated with sexual development downstream of AP2XII-2 control, AP2X-10 and the amino acid hydroxylase AAH1. Our findings show that AP2XII-2 is a key contributor to the gene regulatory pathways modulating Toxoplasma sexual development. IMPORTANCE Toxoplasma gondii is a parasite that undergoes its sexual stage exclusively in feline intestines, making cats a major source of transmission. A better understanding of the proteins controlling the parasite's life cycle stage transitions is needed for the development of new therapies aimed at treating toxoplasmosis and the transmission of the infection. Genes that regulate the sexual stages need to be turned on and off at the appropriate times, activities that are mediated by specific transcription factors that recruit general machinery to silence or activate gene expression. In this study, we identify a transcription factor called AP2XII-2 as being important for the repression of a subset of sexual stage genes, including a sexual stage-specific AP2 factor (AP2X-10) and a protein (AAH1) required to construct the infectious oocysts expelled from infected cats.

Project description:IntroductionThe transcription factor GATA3 is involved in mammary gland development and is crucial for the maintenance of the differentiated status of luminal epithelial cells. The role of GATA3 in breast cancer as a tumor suppressor has been established, although insights into the mechanism of GATA3 expression loss are still required.MethodsChromatin immunoprecipitation assays were conducted to study progestin modulation of recruitment of transcription factors to GATA3 promoter. We performed western blot and reverse RT-qPCR experiments to explore progestin regulation of GATA3 protein and mRNA expression respectively. Confocal microscopy and in vitro phosphorylation studies were conducted to examine progestin capacity to induce GATA3 serine phosphorylation in its 308 residue. GATA3 participation in progestin-induced breast cancer growth was addressed in in vitro proliferation and in vivo tumor growth experiments.ResultsIn this study, we demonstrate that progestin-activated progesterone receptor (PR) reduces GATA3 expression through regulation at the transcriptional and post-translational levels in breast cancer cells. In the former mechanism, the histone methyltransferase enhancer of zeste homolog 2 is co-recruited with activated PR to a putative progesterone response element in the GATA3 proximal promoter, increasing H3K27me3 levels and inducing chromatin compaction, resulting in decreased GATA3 mRNA levels. This transcriptional regulation is coupled with increased GATA3 protein turnover through progestin-induced GATA3 phosphorylation at serine 308 followed by 26S proteasome-mediated degradation. Both molecular mechanisms converge to accomplish decreased GATA3 expression levels in breast cancer cells upon PR activation. In addition, we demonstrated that decreased GATA3 levels are required for progestin-induced upregulation of cyclin A2, which mediates the G1 to S phase transition of the cell cycle and was reported to be associated with poor prognosis in breast cancer. Finally, we showed that downregulation of GATA3 is required for progestin stimulation of both in vitro cell proliferation and in vivo tumor growth.ConclusionsIn the present study, we reveal that progestin-induced PR activation leads to loss of GATA3 expression in breast cancer cells through transcriptional and post-translational regulation. Importantly, we demonstrate that GATA3 downregulation is required for progestin-induced upregulation of cyclin A2 and for progestin-induced in vitro and in vivo breast cancer cell growth.

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Transcriptional repression of Gata3 is essential for early B cell commitment.

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Transcriptional repression of Gata3 is essential for early B cell commitment.

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