Regulation of alternative splicing of Gtf2ird1 and its impact on slow muscle promoter activity

A hallmark of general transcription factor 3 (GTF3) is the presence of multiple GTF2I-like repeats that were suggested to mediate protein-protein interactions. However, we have recently demonstrated that repeat 4 is necessary and sufficient for binding of GTF3 to the bicoid-like motif of the Troponin I slow enhancer. Given the sequence similarity between different GTF2I-like repeats we hypothesized that DNA binding might be a common property of this domain type. We subjected five repeats of GTF3 to random oligonucleotide selection (SELEX) to assess their DNA binding potentials. We delineated the consensus sequence G TC G A GATTA G BG A for repeat 4 and showed that binding sites for GTF3 in enhancers for Troponin I and homeobox c8 (HOXc8) are in very good agreement with this motif. SELEX selections for repeats 5 and 2 enriched for oligonucleotides that were also bound by R4, suggesting that they share common sequence preferences, whereas repeat 3 exhibited relaxed sequence requirements for DNA binding. No binding was observed for repeat 1. We also show that GTF2I-like repeats 4 and 6 of transcription factor II-I (TFII-I) exhibit modest DNA binding properties. Lastly, we identified several amino acids of GTF3 repeat 4 required for high affinity protein-DNA interaction. Based on the ability of many repeats to bind DNA in vitro, we suggest that GTF2I-like domains evolved by duplication and diversification of a prototypic DNA-binding ancestor.

mentioning

confidence: 59%

Multiple GTF2I-like Repeats of General Transcription Factor 3 Exhibit DNA Binding Properties

Vullhorst

Buonanno

2005

“…This explanation is unlikely as transcription of Gtf2ird1 in the (Tg(Alb1-Myc)166.8) mutants was undetectable (24), and analysis of the mutant transcript produced in the Gtf2ird1 tm1Hrd mice reported here indicate very low levels of protein synthesis. All of the reported splice isoforms and (putative) alternative promoter-driven transcripts (18,27) include exons 2 and 3, and would, therefore, be subject to the same translation constraints as the 3␣7 isoform used in the above analysis. Therefore, our estimate of 6% total protein synthesis in homozygous Gtf2ird1 tm1Hrd mice applies, regardless of alternative splicing possibilities.…”

Section: Discussionmentioning

confidence: 99%

Negative Autoregulation of GTF2IRD1 in Williams-Beuren Syndrome via a Novel DNA Binding Mechanism

Palmer

Santucci

Widagdo

et al. 2010

Self Cite

The GTF2IRD1 gene is of principal interest to the study of Williams-Beuren syndrome (WBS). This neurodevelopmental disorder results from the hemizygous deletion of a region of chromosome 7q11.23 containing 28 genes including GTF2IRD1. WBS is thought to be caused by haploinsufficiency of certain dosage-sensitive genes within the deleted region, and the feature of supravalvular aortic stenosis (SVAS) has been attributed to reduced elastin caused by deletion of ELN. Human genetic mapping data have implicated two related genes GTF2IRD1 and GTF2I in the cause of some the key features of WBS, including craniofacial dysmorphology, hypersociability, and visuospatial deficits. Mice with mutations of the Gtf2ird1 allele show evidence of craniofacial abnormalities and behavioral changes. Here we show the existence of a negative autoregulatory mechanism that controls the level of GTF2IRD1 transcription via direct binding of the GTF2IRD1 protein to a highly conserved region of the GTF2IRD1 promoter containing an array of three binding sites. The affinity for this protein-DNA interaction is critically dependent upon multiple interactions between separate domains of the protein and at least two of the DNA binding sites. This autoregulatory mechanism leads to dosage compensation of GTF2IRD1 transcription in WBS patients. The GTF2IRD1 promoter represents the first established in vivo gene target of the GTF2IRD1 protein, and we use it to model its DNA interaction capabilities. Williams-Beuren syndrome (WBS)4 is a relatively rare disorder with an estimated incidence of 1:7,500 -1:20,000 (1). In 95% of cases, this deletion is 1.5 Mb long and results from illegitimate recombination during meiosis between two blocks of low copy repeats (LCRs) that flank the deletion domain (2, 3). To explain the causes of WBS in molecular terms, it is first necessary to identify the genes that underpin each of the disorders. The only symptom that fulfills this to date is SVAS. Haploinsufficiency of elastin due to loss of the ELN gene leads to narrowing of the large elastic aorta and may also affect the pulmonary, coronary, and carotid arteries (4). Accumulating evidence from patients with atypical hemizygous deletions within the critical region indicate that the many of the remaining symptoms, in particular the craniofacial abnormalities, the visuospatial construction deficit and the hypersociability, can be attributed to two genes at the telomeric end of the deletion region, GTF2IRD1 and GTF2I (5-7). These genes share sequence homology and are adjacent, indicating that they have arisen by duplication and divergence from a common ancestor. Functional evidence suggests that these genes encode nuclear proteins with DNA binding capabilities and are widely considered to be transcription factors with specific gene targets (8, 9).The first reported gene product of GTF2IRD1 was Mus-TRD1, which was isolated in a yeast one-hybrid screen for proteins that could bind to a DNA enhancer element present in the TNNI1 gene (10). Human, mouse, and Xenopus orthologs o...

“…BEN also contains a leucine zipper domain, important for multimerization, at its N terminus as well as a C-terminal region rich in serine residues. There are a number of BEN isoforms (at minimum 13) ranging in size from 65 to 160 kDa (51,52,57). Alternative splicing has been shown recently to alter the DNA binding properties of the protein (52).…”

Section: Discussionmentioning

confidence: 99%

Regulation of Immunoglobulin Promoter Activity by TFII-I Class Transcription Factors

Tantin

Tussie-Luna

Roy

et al. 2004

The restriction of immunoglobulin variable region promoter activity to B lymphocytes is a well known paradigm of promoter specificity. Recently, a cis-element, located downstream of the transcription initiation site of murine heavy chain variable promoters, was shown to be critical for B cell activity and specificity. Here we show that mutation of this element, termed DICE (Downstream Immunoglobulin Control Element), reduces in vivo activity in B cells. Gel mobility shift assays show that DICE forms B cell-specific complexes that were also sensitive to DICE mutation. DICE mutation strongly reduces the ability of a distal immunoglobulin heavy chain intronic enhancer to stimulate transcription. We also identify a DICE-interacting factor: a TFII-I-related protein known as BEN (also termed Mus-TRD1 and WBSCR11). Dominant-negative and RNA i -mediated knockdown experiments indicate that BEN can both positively and negatively regulate IgH promoter activity, depending on the cell line.During development, an organism must exclusively express some genes in particular tissues or cell types. A number of genes and their promoters have been studied in detail in order to obtain general principles from their mechanism of regulation. Among the most intensely studied are the immunoglobulin variable (V) 1 region promoters. Ig heavy chain (IgH) and light chain (Ig ) genes are selectively expressed in B lymphocytes, and their respective V region (V H and V ) promoters are normally inactive in all other tissues (reviewed in Ref. 1). The IgH and Ig loci are sequentially activated during B lymphocyte maturation. Activation includes three principal events. Transcription initiates at various positions within each locus, including the V H and V promoters (2, 3). Recombination of a V region gene segment with a joining (J) gene segment, in the case of the chain, and a diversity (D) and J segment, in the case of the heavy chain, forms an Ig variable region exon of a given specificity (4). Hyperacetylation of chromatin within the Ig loci also makes it less compact and more accessible to transacting factors (5, 6). The relative timing of these three events is unclear; however, it has been shown that non-coding (germ line) transcription precedes recombination for some V H segments (3, 7). The direct relationship between these events is also unclear; however, in the heavy chain locus, V-DJ recombination brings a downstream intronic enhancer into proximity with the V H promoter. The relocation of the enhancer greatly stimulates expression of the recombined IgH (reviewed in Ref. 8). These multiple interdependent layers of regulation serve to tightly control Ig expression and prevent activation outside of the normal B cell maturation pathway.Studies from a number of laboratories have determined that in isolation, V H and V promoters are preferentially active in B lymphocytes (9, 10). A feature of most V H and V promoters is the octamer motif (5Ј-ATGCAAAT-3Ј), located 10 -25 nucleotides upstream of the TATA box (and inverted in the case of V ) (9,(1...