Replication protein A binds to regulatory elements in yeast DNA repair and DNA metabolism genes.

Saccharomyces cerevisiae responds to DNA damage by arresting cell cycle progression (thereby preventing the replication and segregation of damaged chromosomes) and by inducing the expression of numerous genes, some of which are involved in DNA repair, DNA replication, and DNA metabolism. Induction of the S. cerevisiae 3-methyladenine DNA glycosylase repair gene (MAG) by DNA-damaging agents requires one upstream activating sequence (UAS) and two upstream repressing sequences (URS1 and URS2) in the MAG promoter. Sequences similar to the MAG URS elements are present in at least 11 other S. cerevisiae DNA repair and metabolism genes. Replication protein A (Rpa) is known as a single-stranded-DNA-binding protein that is involved in the initiation and elongation steps of DNA replication, nucleotide excision repair, and homologous recombination. We now show that the MAG URS1 and URS2 elements form similar double-stranded, sequence-specific, DNA-protein complexes and that both complexes contain Rpa. Moreover, Rpa appears to bind the MAG URS1-like elements found upstream of 11 other DNA repair and DNA metabolism genes. These results lead us to hypothesize that Rpa may be involved in the regulation of a number of DNA repair and DNA metabolism genes.     

The human enhancer-binding protein Gata3 binds to several T-cell receptor regulatory elements.

The tissue-specific developmental regulation of the alpha, beta, gamma and delta T-cell antigen receptor (TCR) genes is controlled by the corresponding distinct enhancers and their enhancer-binding proteins. To find a common TCR regulatory element, we have studied the ability of the newly described enhancer-binding protein Gata3 to bind to the sequence motif (A/T)GATA(G/A) shared between enhancer elements of all four TCR genes. Gata3 was shown in the chicken to be an enhancer-binding protein containing a zinc-finger domain. We have isolated a full-length human Gata3 cDNA and characterized its role in TCR gene regulation. Northern blot analysis showed that the Gata3 mRNA is present in T-cell, but not B-cell, macrophage or HeLa cell lines. The in vitro transcribed and translated Gata3 cDNA product bound specifically to the enhancer elements of at least three TCR genes containing the above consensus sequence. This binding specificity was consistent with the in vivo activity of the TCR enhancer-binding element in transient transfection assays. Hence, the Gata3 protein, together with other T-cell-specific enhancer-binding proteins, may form the basis of tissue-specific expression of the TCR genes.     

A short review of human gamma-globin gene anomalies

This paper summarizes various anomalies involving the duplicated gamma-globin genes which are normally arranged as -G gamma-A gamma-. Variations include G gamma----A gamma and A gamma----G gamma replacements; newborn heterozygotes for these anomalies have low G gamma (A gamma A gamma/G gamma A gamma) or high G gamma values (G gamma G gamma/G gamma A gamma). Additional abnormalities include the deletion of one gamma-globin gene (-A gamma-; gamma-thalassemia), gamma-globin gene triplications (G gamma-AG gamma-A gamma; G gamma-G gamma-A gamma) and gamma-globin gene quadruplications (G gamma-G gamma-G gamma-A gamma-); several of these conditions are found in different populations but at low frequencies. In adults, the G gamma to A gamma ratio in the Hb F and often also the Hb F level, are directly related to specific structural characteristics of the chromosome; specific mutations in the promoter sequences 5' to G gamma or to A gamma, for instance, may result in increased G gamma or A gamma chain production with an increase in Hb F levels (nondeletional G gamma- or A gamma-HPFH) or with only modest changes in the total Hb F (normal adults; Swiss-HPFH). Similar variations have been observed in SS patients.     

A protein that preferentially binds Drosophila satellite DNA.

Using a nitrocellulose filter binding assay, we have detected and partially purified a protein from embryos of Drosophila melanogaster that preferentially binds to a highly repeated satellite DNA of the same species. Formation of the satellite DNA-protein complex requires physiological conditions of salt and temperature, but once formed, the complex is stable in high salt (1 M NaCl) or at low temperature. Optimal formation of the specific complex also requires the satellite DNA to be in a supertwisted conformation. The protein interacts with a limited region within the 359-base-pair repeated sequence of the satellite DNA.     

Nucleotide sequence of the yeast regulatory gene STE7 predicts a protein homologous to protein kinases.

The nucleotide sequence of the STE7 gene from Saccharomyces cerevisiae has been determined. It is one of several regulatory genes implicated in the control of cell-type-specific gene expression in yeast. The predicted amino acid sequence shows homology to several members of the protein kinase family. These results suggest protein phosphorylation may play a key role in regulation leading to cell-type specialization in yeast.     

DNA gyrase binds to the family of prokaryotic repetitive extragenic palindromic sequences.

A family of repetitive extragenic palindromic (REP) sequences is composed of hundreds of copies distributed throughout the chromosome. Their palindromic nature and conservation suggested that they are specifically recognized by a protein(s). We have identified DNA gyrase [DNA topoisomerase (ATP-hydrolysing), EC 5.99.1.3] as one of the REP-binding proteins. Gyrase has at least a 10-fold higher affinity for DNA containing REP sequences than for DNA not containing REP sequences. Binding effectiveness correlates directly with the number of REP sequences in the DNA. DNase I footprinting shows that gyrase protects 205 base pairs on a REP-containing DNA fragment enclosing the REP sequences. In agreement with the above results, a comparison of the REP consensus sequence with the sequence of previously identified pBR322 "strong" gyrase cleavage sites reveals a high degree of homology. Because REP sequences are numerous and found throughout the genome, we suggest they have physiological functions mediated through their interaction with gyrase, such as being sites of action for the maintenance of DNA supercoiling. In addition, we speculate that these interactions may be of a structural nature, such as involvement in the higher-order structure of the bacterial chromosome.