Loss of Bright/ARID3a function promotes developmental plasticity.

B-cell regulator of immunoglobulin heavy chain transcription (Bright)/ARID3a, an A+T-rich interaction domain protein, was originally discovered in B lymphocyte lineage cells. However, expression patterns and high lethality levels in knockout mice suggested that it had additional functions. Three independent lines of evidence show that functional inhibition of Bright results in increased developmental plasticity. Bright-deficient cells from two mouse models expressed a number of pluripotency-associated gene products, expanded indefinitely, and spontaneously differentiated into cells of multiple lineages. Furthermore, direct knockdown of human Bright resulted in colonies capable of expressing multiple lineage markers. These data suggest that repression of this single molecule confers adult somatic cells with new developmental options.

Transgenic mice expressing dominant-negative bright exhibit defects in B1 B cells.

The transcription factor Bright up-regulates Ig H chain production from select V region promoters and requires Bright dimerization, Bruton's tyrosine kinase (Btk), and the Btk substrate, TFII-I, for this activity. Defects in Btk cause X-linked immunodeficiency disease in mice and humans. Btk-deficient mice exhibit decreased serum IgM production, B cell developmental blocks, absence of peritoneal B1 cells, and subnormal immune responses against Ags, including phosphorylcholine, which confer protection against Streptococcus pneumoniae. Transgenic mice expressing dominant-negative Bright share similarities with Btk-deficient mice, including decreased serum IgM, poor anti-phosphorylcholine responses, and slightly reduced numbers of mature B cells. Although dominant-negative Bright mice developed B1 B cells, these were functionally deficient in Ig secretion. These data suggest a mechanistic explanation for the abnormal responses to phosphorylcholine observed in Btk-deficient mice, and indicate that Bright functions in a subset of Btk-dependent pathways in vivo, particularly those responses dominated by B1 B cells.

Anti-nuclear antibody production and autoimmunity in transgenic mice that overexpress the transcription factor Bright.

The B cell-restricted transcription factor, B cell regulator of Ig(H) transcription (Bright), up-regulates Ig H chain transcription 3- to 7-fold in activated B cells in vitro. Bright function is dependent upon both active Bruton's tyrosine kinase and its substrate, the transcription factor, TFII-I. In mouse and human B lymphocytes, Bright transcription is down-regulated in mature B cells, and its expression is tightly regulated during B cell differentiation. To determine how Bright expression affects B cell development, transgenic mice were generated that express Bright constitutively in all B lineage cells. These mice exhibited increases in total B220(+) B lymphocyte lineage cells in the bone marrow, but the relative percentages of the individual subpopulations were not altered. Splenic immature transitional B cells were significantly expanded both in total cell numbers and as increased percentages of cells relative to other B cell subpopulations. Serum Ig levels, particularly IgG isotypes, were increased slightly in the Bright-transgenic mice compared with littermate controls. However, immunization studies suggest that responses to all foreign Ags were not increased globally. Moreover, 4-wk-old Bright-transgenic mice produced anti-nuclear Abs. Older animals developed Ab deposits in the kidney glomeruli, but did not succumb to further autoimmune sequelae. These data indicate that enhanced Bright expression results in failure to maintain B cell tolerance and suggest a previously unappreciated role for Bright regulation in immature B cells. Bright is the first B cell-restricted transcription factor demonstrated to induce autoimmunity. Therefore, the Bright transgenics provide a novel model system for future analyses of B cell autoreactivity.

Induction of immunoglobulin heavy-chain transcription through the transcription factor Bright requires TFII-I.

Bright/ARID3a/Dril1, a member of the ARID family of transcription factors, is expressed in a highly regulated fashion in B lymphocytes, where it enhances immunoglobulin transcription three- to sixfold. Recent publications from our lab indicated that functional, but not kinase-inactive, Bruton's tyrosine kinase (Btk) is critical for Bright activity in an in vitro model system, yet Bright itself is not appreciably tyrosine phosphorylated. These data suggested that a third protein, and Btk substrate, must contribute to Bright-enhanced immunoglobulin transcription. The ubiquitously expressed transcription factor TFII-I was identified as a substrate for Btk several years ago. In this work, we show that TFII-I directly interacts with human Bright through amino acids in Bright's protein interaction domain and that specific tyrosine residues of TFII-I are essential for Bright-induced activity of an immunoglobulin reporter gene. Moreover, inhibition of TFII-I function in a B-cell line resulted in decreased heavy-chain transcript levels. These data suggest that Bright functions as a three-component protein complex in the immunoglobulin locus and tie together previous data indicating important roles for Btk and TFII-I in B lymphocytes.

Bruton's tyrosine kinase regulates immunoglobulin promoter activation in association with the transcription factor Bright.

Bright (B-cell regulator of immunoglobulin heavy chain transcription) binding to immunoglobulin heavy chain loci after B-cell activation is associated with increased heavy chain transcription. Our earlier reports demonstrated that Bright coimmunoprecipitates with Bruton's tyrosine kinase (Btk) and that these proteins associate in a DNA-binding complex in primary B cells. B cells from immunodeficient mice with a mutation in Btk failed to produce stable Bright DNA-binding complexes. In order to determine if Btk is important for Bright function, a transcription activation assay was established and analyzed using real-time PCR technology. Cells lacking both Bright and Btk were transfected with Bright and/or Btk along with an immunoglobulin heavy chain reporter construct. Immunoglobulin gene transcription was enhanced when Bright and Btk were coexpressed. In contrast, neither Bright nor Btk alone led to activation of heavy chain transcription. Furthermore, Bright function required both Btk kinase activity and sequences within the pleckstrin homology domain of Btk. Bright was not appreciably phosphorylated by Btk; however, a third tyrosine-phosphorylated protein coprecipitated with Bright. Thus, the ability of Bright to enhance immunoglobulin transcription critically requires functional Btk.

Mutations in the DNA-binding domain of the transcription factor Bright act as dominant negative proteins and interfere with immunoglobulin transactivation.

Bright, for B cell regulator of immunoglobulin heavy chain transcription, binds A+T-rich sequences in the intronic enhancer regions of the murine heavy chain locus and 5'-flanking sequences of some variable heavy chain promoters. Most resting B cells do not express Bright; however, it is induced after stimulation with antigen or polyclonal mitogens. Bright activation results in up-regulation of mu transcription; however, it is not clear whether Bright function is critical for normal B cell development. To begin to address Bright function during B cell development, seven mutated forms of Bright were produced. Five of the seven mutants revealed little or no DNA binding activity. Furthermore, because Bright binds DNA as a dimer, two of the mutants formed complexes with wild type Bright and acted in a dominant negative fashion. Dominant negative Bright prevented the up-regulation of mu transcription in transfected Chinese hamster ovary cells transfected with wild type Bright. These data identify regions within Bright that are required for the DNA binding activity of Bright and for its function as a transcription factor.

The transcription factor, Bright, is not expressed in all human B lymphocyte subpopulations.

Bright is an ARID family transcription factor that increases immunoglobulin heavy chain transcription. In the mouse, Bright expression is tightly regulated and B cell-restricted and the Bright protein associates with Bruton's tyrosine kinase (Btk), the defective enzyme in X-linked immunodeficiency. Human X-linked agammaglobulinemia results from defects in Btk and leads to early blocks in B lymphocyte development. Because so little is known about human Bright, we sought to determine where human Bright is expressed in normal B cell differentiation and whether it also forms complexes with Btk. Although human and mouse Bright exhibited similar expression patterns in normal B cells, many human transformed B cell lines did not express Bright protein. However, the human protein bound prototypic Bright DNA-binding motifs and, like mouse Bright, was capable of associating with Btk. These data suggest potentially important similarities exist in Bright expression and activity in human and mouse B lymphocytes.