Members of the zinc finger protein gene family sharing a conserved N-terminal module

We report the isolation of human members of a subfamily of structurally related finger protein genes. These potentially encode polypeptides containing finger motifs of the Krüppel type at the C-terminus, and a conserved amino acid module at the N-terminus; because of its invariant location the latter is referred to as finger preceding box (FPB). The FPB, detected also in previously described finger proteins from human, mouse and Xenopus, extends over approximately 65 amino acids and appears to be composed of two contiguous modules: FPB-A (residues 1-42) and FPB-B (residues 43-65). The latter is absent in some of the members analyzed. Elements A and B and the zinc finger domain are encoded by separate exons in the ZNF2 gene, a human member of this sub-family. The positioning of introns within this gene is remarkable. One intron flanks and a second interrupts the first codon of the FPB-A and FPB-B modules, respectively. A third intron occurs a few nucleotides downstream of FPB-B marking its separation from the remainder of the coding sequences. This organization, together with the absence of FPB-B in some cDNAs, supports the hypothesis that mRNAs encoding polypeptides that include one, both or none of the FPB-A and FPB-B modules may be assembled through alternative splicing pathways. Northern analyses showed that members of his sub-family are expressed as multiple transcripts in several cell lines. The sequences of distinct cDNAs homologous to the ZNF2 gene indicate that alternative splicing events adjoin either coding or non oding xons to the FPB sequences. © 1991 Oxford University Press

Enhanced Glutathione Levels and Oxidoresistance Mediated by Increased Glucose-6-phosphate Dehydrogenase Expression *

Glucose-6-phosphate dehydrogenase (G6PD) is the key enzyme of the pentose phosphate pathway that is responsible for the generation of NADPH, which is required in many detoxifying reactions. We have recently demonstrated that G6PD expression is induced by a variety of chemical agents acting at different steps in the biochemical pathway controlling the intracellular redox status. Although we obtained evidence that the oxidative stress-mediated enhancement of G6PD expression is a general phenomenon, the functional significance of such G6PD induction after oxidant insult is still poorly understood. In this report, we used a GSH-depleting drug that determines a marked decrease in the intracellular pool of reduced glutathione and a gradual but notable increase in G6PD expression. Both effects are seen soon after drug addition. Once G6PD activity has reached the maximum, the GSH pool is restored. We suggest and also provide the first direct evidence that G6PD induction serves to maintain and regenerate the intracellular GSH pool. We used HeLa cell clones stably transfected with the human G6PD gene that display higher G6PD activity than the parent HeLa cells. Although the activities of glutathione peroxidase, glutathione reductase, and catalase were comparable in all strains, the concentrations of GSH were significantly higher in G6PD-overexpressing clones. A direct consequence of GSH increase in these cells is a decreased reactive oxygen species production, which makes these cells less sensitive to the oxidative burst produced by external stimuli. Indeed, all clones that constitutively overexpress G6PD exhibited strong protection against oxidants-mediated cell killing. We also observe that NF-kappa B activation, in response to tumor necrosis factor-alpha treatment, is strongly reduced in human HeLa cells overexpressing G6PD

Molecular anatomy of the human glucose 6-phosphate dehydrogenase core promoter

The gene encoding glucose 6-phosphate dehydrogenase (G6PD), which plays a pivotal role in cell defense against oxidative stress, is ubiquitously expressed at widely different levels in various tissues; moreover, G6PD expression is regulated by a number of stimuli. In this study we have analyzed the molecular anatomy of the G6PD core promoter. Our results indicate that the G6PD promoter is more complex than previously assumed; G6PD expression is under the control of several elements that are all required for correct promoter functioning and, furthermore, a still unidentified mammalian specific factor is needed. Copyright (C) 1998 Federation of European Biochemical Societies

Bimodal strategy for excellent audiological rehabilitation in a subject with a novel nonsense mutation of the SLC26A4 gene: A case report

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Human Synaptobrevin-like 1 Gene Basal Transcription Is Regulated through the Interaction of Selenocysteine tRNA Gene Transcription Activating Factor-Zinc Finger 143 Factors with Evolutionary Conserved Cis-elements

The synaptobrevin-like 1 (SYBL1) gene is ubiquitously expressed and codes for an unusual member of the v-SNAREs molecules implicated in cellular exocytosis. This X-linked gene has the peculiarity of also being present on the Y chromosome in a transcriptional inactive status. Moreover, although ubiquitous, the function of SYBL1 is prominent in specific tissues, such as brain. As a first insight into the molecular mechanisms controlling SYBL1 expression, in this report we describe the extent and role of SYBL1 upstream regions and characterize the binding of trans-acting factors. In vivo foot-printing experiments identify three protected regions. Band shift and transient reporter gene assays indicate a strong role of two of these evolutionary conserved regions in regulating SYBL1 transcription. Because one site is the classical CAAT box, we characterized the binding to the other site of the mammalian homologues of the selenocysteine tRNA gene transcription activating factor (Staf) family, zinc-finger transcription factors, and their role in regulating SYBL1 expression. The results reported here clarify that a Staf-zinc finger family factor, together with the CAAT factor, is the major nuclear protein bound to the SYBL1 promoter region and is responsible for its regulation in HeLa cells, thus identifying the basic control of SYBL1 transcription. In vivo binding of Staf proteins to the SYBL1 promoter is confirmed by chromatin immunoprecipitation assays. Our results identify a fourth mRNA promoter stimulated by a member of the Staf-zinc finger family, the function of which on mRNA polymerase II promoters is still very poorly understood

An upsteam enhancer and a negative element in the 5' flanking region of the human urokinase plasminogen activator gene

The 5' flanking region of the human urokinase (uPA) gene has been fused to the reporter chloramphenicol acetyl transferase (CAT) gene and its activity assayed by transfection in two human cell lines. Progressive deletions of the uPA regulatory region from the 5' end maintain a high level of expression provided at least 1870 (in A1251 cells) or 1963 (in HFS10 cells) nucleotides of the 5' flanking region are retained. A DNA fragment from -2350 to -1824 has enhancer properties, stimulating transcription of an enhancerless SV40 early promoter independently of orientation and distance. Internal deletions that still retain the enhancer element reveal the presence of negative cis-acting sequences between -1824 and -1572. Their removal, in fact, increases uPA transcriptional activity. Differences of expression of the uPA-CAT fusion genes in the two cell lines are also observed, indicating the presence of cell-specific cis-acting sequence