Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Yeast mutants assigned to the pet complementation group G104 were found to lack alpha-ketoglutarate dehydrogenase activity as a result of mutations in the dihydrolipoyl transsuccinylase (KE2) component of the complex. The nuclear gene KGD2, coding for yeast KE2, was cloned by transformation of E250/U6, a G104 mutant, with a yeast genomic library. Analysis of the KGD2 sequence revealed an open reading frame encoding a protein with a molecular weight of 52,375 and 42% identities to the KE2 component of Escherichia coli alpha-ketoglutarate dehydrogenase complex. Disruption of the chromosomal copy of KGD2 in a respiratory-competent haploid yeast strain elicited a growth phenotype similar to that of G104 mutants and abolished the ability to mitochondria to catalyze the reduction of NAD+ by alpha-ketoglutarate. The expression of KGD2 was transcriptionally regulated by glucose. Northern (RNA) analysis of poly(A)+ RNA indicated the existence of two KGD2 transcripts differing in length by 150 nucleotides. The concentrations of both RNAs were at least 10 times lower in glucose (repressed)- than in galactose (derepressed)-grown cells. Different 5'-flanking regions of KGD2 were fused to the lacZ gene of E. coli in episomal plasmids, and the resultant constructs were tested for expression of beta-galactosidase in wild-type yeast cells and in hap2 and hap3 mutants. Results of the lacZ fusion assays indicated that transcription of KGD2 is activated by the HAP2 and HAP3 proteins. The regulated expression of KGD2 was found to depend on sequences that map to a region 244 to 484 nucleotides upstream of the structural gene. This region contains two short sequence elements that differ by one nucleotide from the consensus core (5'-TN[A/G]TTGGT-3') that has been proposed to be essential for binding of the HAP activation complex. These data together with earlier reports on the regulation of the KGD1 and LPD1 genes for the alpha-ketoglutarate and dihydrolipoyl dehydrogenases indicate that all three enzyme components of the complex are catabolite repressed and subject to positive regulation by the HAP2 and HAP3 proteins.
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PMID:Structure and regulation of KGD2, the structural gene for yeast dihydrolipoyl transsuccinylase. 211 21

Transcriptional activation by the yeast CYC1 upstream activation site UAS2UP1 requires the products of both the HAP2 and HAP3 regulatory genes. We show here that both HAP2 and HAP3 in yeast extracts bind to UAS2UP1 and give rise to a single protein-DNA complex, termed C, in nondenaturing polyacrylamide gels. That both products are a part of complex C was shown by altering the mobility of the complex by fusing either HAP2 or HAP3 to beta-galactosidase. Further, methylation interference footprinting showed that sequences in UAS2UP1 contacted in complex C were identical to those contacted in either fusion protein complex. Binding was centered on the sequence TGATTGGT, also found in the UASs of other genes subject to activation by the HAP2-HAP3 system and homologous to the CCAAT box sequence found in higher cells. The binding of either HAP2 or HAP3 was abolished when synthesized in a strain mutant in the complementary HAP gene. Thus the binding of HAP2 and HAP3 to UAS2UP1 is interdependent. The involvement of multiple gene products in binding to a single site is discussed with reference to other systems in yeast and higher cells.
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PMID:Yeast HAP2 and HAP3 activators both bind to the CYC1 upstream activation site, UAS2, in an interdependent manner. 282 15

We generated neurotropic herpes simplex type 1 viruses expressing human placental alkaline phosphatase and studied the utility of this enzyme as a marker of infected neurons. The neurotropism of these viruses was assessed by their ability to infect sympathetic preganglionic neurons after adrenal injection in hamsters. The transneuronal transfer of these viruses was examined by their ability to cross the peripheral synapse from the kidney to renal preganglionic neurons or to cross the central synapse from the adrenal gland to the medulla oblongata. Finally, we injected an alkaline phosphatase-expressing herpes simplex virus into the adrenal gland and a beta-galactosidase-expressing herpes simplex virus (US5gal) into the muscular wall of the small intestine to label two neural circuits in one animal and to assess the feasibility of a dual-virus labelling system. The alkaline phosphatase gene was inserted into the glycoprotein J locus or the virus-induced host shut-off locus in the herpes simplex genome to create viruses which replicate (gJHAP HSV or vhsHAP HSV) or into the thymidine kinase locus to generate a virus that does not replicate in neurons in vivo (TK- HAP HSV). Each of the three viruses was retrogradely transported from the adrenal gland of hamsters to sympathetic preganglionic neurons, suggesting that the neurotropism of these viruses was maintained. gJHAP HSV travelled transneuronally from the kidney to sympathorenal preganglionic neurons and from the adrenal gland to neurons in the rostral ventrolateral medulla. Neuronal infection with alkaline phosphatase-expressing virus could be identified using histochemistry but detailed morphology of these neurons was not revealed. However, staining by anti-herpes simplex virus immunoperoxidase demonstrated that they had normal morphology. Identification of two distinct neural circuits in one animal was achieved with our dual-virus labelling system. The nonreplicating TK- HAP HSV was used in combination with US5gal to identify intestinal and adrenal sympathetic preganglionic neurons. The beta-galactosidase-expressing intestinal neurons were labelled bilaterally in the nucleus intermediolateralis, pars principalis, and alkaline phosphatase-expressing adrenal neurons were found ipsilaterally. Some clusters of sympathetic preganglionic neurons in the nucleus intermediolateralis, pars principalis contained mostly intestinal sympathetic preganglionic neurons and a few adrenal sympathetic preganglionic neurons. In other areas, the opposite pattern occurred. About 3-7% of the labelled sympathetic preganglionic neurons were double-labelled by both markers. The distinct and crisp morphology and dendritic processes of neurons stained by beta-galactosidase histochemistry contrasted with the partial staining of neurons by alkaline phosphatase, revealing beta-galactosidase as a better marker of infected neurons. In conclusion, alkaline phosphatase-expressing herpes simplex viruses are yet neurotropic after insertion of this marker enzyme into any of three different loci of the herpes simplex genome. One replicating alkaline phosphatase-expressing virus travelled transneuronally. These alkaline phosphatase-expressing herpes simplex virus can be used together with beta-galactosidase-expressing herpes simplex viruses to determine the target specificity of sympathetic preganglionic neurons controlling visceral organs or can be used to express two different recombinant genes in two targeted neuronal populations. This study suggests that sympathetic preganglionic neurons controlling the intestine and adrenal gland are almost completely distinct.
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PMID:Simultaneous identification of two populations of sympathetic preganglionic neurons using recombinant herpes simplex virus type 1 expressing different reporter genes. 946 44

The SLTM [SAF (scaffold attachment factor)-like transcription modulator] protein contains a SAF-box DNA-binding motif and an RNA-binding domain, and shares an overall identity of 34% with SAFB1 {scaffold attachment factor-B1; also known as SAF-B (scaffold attachment factor B), HET [heat-shock protein 27 ERE (oestrogen response element) and TATA-box-binding protein] or HAP (heterogeneous nuclear ribonucleoprotein A1-interacting protein)}. Here, we show that SLTM is localized to the cell nucleus, but excluded from nucleoli, and to a large extent it co-localizes with SAFB1. In the nucleus, SLTM has a punctate distribution and it does not co-localize with SR (serine/arginine) proteins. Overexpression of SAFB1 has been shown to exert a number of inhibitory effects, including suppression of oestrogen signalling. Although SLTM also suppressed the ability of oestrogen to activate a reporter gene in MCF-7 breast-cancer cells, inhibition of a constitutively active beta-galactosidase gene suggested that this was primarily the consequence of a generalized inhibitory effect on transcription. Measurement of RNA synthesis, which showed a particularly marked inhibition of [(3)H]uridine incorporation into mRNA, supported this conclusion. In addition, analysis of cell-cycle parameters, chromatin condensation and cytochrome c release showed that SLTM induced apoptosis in a range of cultured cell lines. Thus the inhibitory effects of SLTM on gene expression appear to result from generalized down-regulation of mRNA synthesis and initiation of apoptosis consequent upon overexpressing the protein. While indicating a crucial role for SLTM in cellular function, these results also emphasize the need for caution when interpreting phenotypic changes associated with manipulation of protein expression levels.
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PMID:A novel member of the SAF (scaffold attachment factor)-box protein family inhibits gene expression and induces apoptosis. 1763 Sep 52