Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.17 (lysozyme)
21,489 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Expression of the lysozyme gene in chicken macrophages is regulated by an enhancer located 2.7 kilobases upstream of the transcription start site. The organization of this enhancer was analyzed by in vitro assays (DNase I footprinting, dimethyl sulfate methylation protection, and band shift assays) and in vivo footprinting experiments. The results show that the enhancer contains four regions (I-IV) of protein-DNA interactions. First, transient gene transfer experiments demonstrate that region II contributes most to the enhancer function. This region contains a recognition site for the macrophage- and B cell-specific transcription factor PU.1, a member of the Ets transcription factor family. This site plays a major role in the enhancer since a mutation of this site abolishes enhancer activity. Second, in competition band shift experiments, we show that the only myeloid-specific complexes in regions I and II with nuclear factors are formed on the PU.1 recognition site by a member of the Ets transcription factor family. Third in in vivo footprinting experiments, the only sign of a protein-DNA interaction is a dimethyl sulfate-hyperreactive guanine 3'-adjacent to the PU.1 recognition site.
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PMID:Characterization of a myeloid-specific enhancer of the chicken lysozyme gene. Major role for an Ets transcription factor-binding site. 802 33

We studied the effects of thyroid hormone (T3) on nuclear protein-DNA interactions by using dimethyl sulfate (DMS) and DNase I ligation-mediated PCR footprinting. We examined an endogenous gene the growth hormone (GH) gene, and a stably transfected plasmid containing the chicken lysozyme silencer (F2) T3 response element (TRE) gene, F2-TRE-TK-CAT, both in pituitary tumor (GC) cells. The 235-1 cell line, which expresses prolactin (PRL) and Pit-1, but not the T3 receptor (TR) or GH, was used as a control. DMS and DNase I footprinting identified protected G residues in the Pit-1, Sp1, and Zn-15 binding sites of the GH gene in GC, but not in 235-1, cells. There was no specific protection of the tripartite GH TRE at -180 bp against either DMS or DNase I in the absence or presence of T3 in either cell line. However, T3 increased protection of the Pit-1 and Sp1 binding sites against DMS in GC cells. In GC cells stably transfected with a plasmid containing F2-TRE-TK-CAT or TRalpha, chloramphenicol acetyltransferase expression was T3 inducible and DMS footprinting revealed both F2 TRE TR-binding half sites in a pattern suggesting the binding of TR homodimers before and during T3 exposure. We conclude that the GH gene is accessible to specific nuclear proteins in GC, but not in 235-1, cells and that T3 enhances this interaction, although there is no evidence of TR binding to the low-affinity rat GH TRE. The presence of TR binding to the high-affinity F2 TRE before and during T3 exposure suggests that reversible interaction of T3 with DNA-bound TRs, rather than transient T3-TR contact with TREs, determines the level of T3-stimulated transcriptional activation.
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PMID:In vivo genomic footprinting of thyroid hormone-responsive genes in pituitary tumor cell lines. 875 47

Differentiation of myeloid precursor cells results in transcriptional activation of the myeloid-specific murine M-lysozyme gene. M-lysozyme gene expression depends on the differentiation state of the myeloid cells and provides a marker for myeloid leukemias. The mouse lysozyme downstream enhancer (MLDE) was colocalized previously with the DNase I hypersensitive site in the chromatin of mature macrophages and shown to be macrophage differentiation-dependent. The correlation of the hypersensitive site appearance with expression of the M-lysozyme gene suggests that the enhancer becomes activated during macrophage differentiation. However, the predominant MLDE-binding protein GABP is ubiquitously expressed, indicating that additional regulatory mechanisms are required for restricting the tissue-specific activity of the enhancer. To demonstrate the specificity of the enhancer in vivo, we examined the in vivo interaction of factors with the MLDE in T cells, immature macrophage cells, and in macrophage cells. Although identical DNase I protection activity is present in extracts from all tested cell lines in vitro, the in vivo interaction of proteins is restricted to mature macrophage cells. The presence of factors capable of interacting with the enhancer is not sufficient for enhancer activity, suggesting that the process of differentiation results in factor accessibility for the MLDE. Analysis of the MLDE methylation state revealed a correlation between demethylation of the single CpG dinucleotide within the MLDE sequence and the in vivo interaction of proteins.
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PMID:In vivo protein interaction with the mouse M-lysozyme gene downstream enhancer correlates with demethylation and gene expression. 893 Apr 4

The complete chicken lysozyme locus is expressed in a position independent fashion in macrophages of transgenic mice and forms the identical chromatin structure as observed with the endogenous gene in chicken cells. Individual lysozyme cis -regulatory elements reorganize their chromatin structure at different developmental stages. Accordingly, their activities are developmentally regulated, indicating a differential role of these elements in locus activation. We have shown previously that a subset of enhancer elements and the promoter are sufficient to activate transcription of the chicken lysozyme gene at the correct developmental stage. Here, we analyzed to which grade the developmentally controlled chromatin reorganizing capacity of cis -regulatory elements in the 5'-region of the chicken lysozyme locus is dependent on promoter elements, and we examined whether the lysozyme locus carries a dominant chromatin reorganizing element. To this end we generated transgenic mouse lines carrying constructs with a deletion of the lysozyme promoter. Expression of the transgene in macrophages is abolished, however, the chromatin reorganizing ability of the cis -regulatory elements is differentially impaired. Some cis -elements require the interaction with the promoter to stabilize transcription factor complexes detectable as DNase I hypersensitive sites in chromatin, whereas other elements reorganize their chromatin structure autonomously.
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PMID:The developmental activation of the chicken lysozyme locus in transgenic mice requires the interaction of a subset of enhancer elements with the promoter. 922 98

The mouse lysozyme downstream enhancer was previously colocalized with the DNase I-hypersensitive site in the chromatin of mature macrophages. This hypersensitive site was shown to be macrophage differentiation-dependent. Demethylation of CpG sequences within the enhancer is correlated with lysozyme expression in mature macrophages. Binding of the GABP heterotetrameric transcription factor to the enhancer core element (MLDE), only seen in vivo on the demethylated MLDE element in macrophages, is inhibited by DNA methylation. Here, we analyzed the DNA sequences required for demethylation. In electrophoretic mobility shift experiments we found that in addition to the complete methylated MLDE the hemimethylated form of the lower strand inhibits GABP binding as well. Therefore, GABP is unlikely to be the mediator of demethylation. In addition, we show by stable DNA transfections of methylated mouse lysozyme enhancer sequences that MLDE-flanking sequences are required for demethylation. We narrowed down these DNA elements to two short regions of 163 and 79 base pairs on either side of the MLDE, each of which is sufficient to mediate demethylation of the GABP site.
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PMID:Cis-elements required for the demethylation of the mouse M-lysozyme downstream enhancer. 925 11

The mouse M-lysozyme downstream enhancer has been previously characterized on several levels of gene regulation. The enhancer was co-localized with a DNase I hypersensitive site in the chromatin of mature macrophages, the in vivo interaction of transcription factor GABP with the enhancer core (MLDE) demonstrated binding being restricted to mature macrophage cells, and analysis of the MLDE methylation state revealed a correlation between demethylation of CpG dinucleotides and the in vivo GABP binding. Here, we analyzed in detail the full-length enhancer in addition to the core element. We identified a total of nine binding sites for nuclear factors. Most of these factors are found ubiquitously in all cell types tested. These factors include several unknown proteins as well as the transcription factor NF-Y. In addition, three binding sites for a new single-stranded DNA binding protein were found. The presence of this factor in mature macrophages correlates with the in vivo DNA melting of one of the binding sites and with the enhancer strength.
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PMID:Complex protein binding to the mouse M-lysozyme gene downstream enhancer involves single-stranded DNA binding. 937 40

Refolding of denatured-reduced lysozyme and the effect of co-refolding it with other proteins such as RNase A, bovine serum albumin, histone, myelin basic protein, alcohol dehydrogenase and DNase I on the renaturation yield and the aggregation of lysozyme have been studied. Basic proteins consistently increase the renaturation yield of the basic protein lysozyme (10-20% more than in their absence) with little or no aggregation. On the other hand, co-refolding of lysozyme with acidic proteins leads to aggregation and a significant decrease in renaturation yields. Our results show that hetero-interchain interactions (non-specific interactions) occur when the basic protein lysozyme is refolded together with acidic proteins such as bovine serum albumin, alcohol dehydrogenase or DNase I. Our results also suggest that the net charge on proteins plays a significant role in such non-specific aggregation. These results should prove useful in understanding the hetero-interchain interactions between folding polypeptide chains.
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PMID:Co-refolding denatured-reduced hen egg white lysozyme with acidic and basic proteins. 942 46

We previously identified a broad initiation zone of DNA replication at the chicken lysozyme gene locus. However, the existence of a highly preferred origin of bidirectional replication (OBR), often found in initiation zones, remained elusive. In order to re-examine this issue we used a competitive PCR assay to determine the abundance of closely spaced genomic segments in a 1 kb size fraction of nascent DNA. A sharp peak of nascent strand abundance occurred at the 3" end of the gene, where initiation events were 17 times more frequent than upstream of the gene. This primary initiation site, active in lysozyme expressing myelomonocytic HD11 cells and non-expressing hepatic DU249 cells, was found to reside within an unusually located CpG island. While most CpG islands are found at the 5" end of genes, the lysozyme gene island extends from the 3" end of the second intron and includes approximately 1.2 kb of 3" flanking DNA. As diagnosed by methylation-sensitive restriction enzymes, the island is largely non-methylated in HD11 cells, DU249 cells and inactive chicken erythrocytes. Furthermore, a DNase I hypersensitive site (HS) that is composed of two subsites separated by approximately 100 bp, was localised very close to the segment with the highest initiation activity. Our results suggest that the non-methylated CpG island and the HS provide an accessible chromatin structure for the lysozyme gene origin of replication.
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PMID:An origin of bidirectional DNA replication is located within a CpG island at the 3" end of the chicken lysozyme gene. 1045 94

The chicken lysozyme gene -3.9 kb enhancer forms a DNase I hypersensitive site (DHS) early in macrophage differentiation, but not in more primitive multipotent myeloid precursor cells. A nucleosome becomes precisely positioned across the enhancer in parallel with DHS formation. In transfection assays, the 5'-part of the -3.9 kb element has ubiquitous enhancer activity. The 3'-part has no stimulatory activity, but is necessary for enhancer repression in lysozyme non-expressing cells. Recent studies have shown that the chromatin fine structure of this region is affected by inhibition of histone deacetylase activity after Trichostatin A (TSA) treatment, but only in lysozyme non-expressing cells. These results indicated a developmental modification of chromatin structure from a dynamic, but inactive, to a stabilised, possibly hyperacetylated, active state. Here we have identified positively and negatively acting transcription factors binding to the -3.9 kb enhancer and determined their contribution to enhancer activity. Furthermore, we examined the influence of TSA treatment on enhancer activity in macrophage cells and lysozyme non-expressing cells, including multipotent macrophage precursors. Interestingly, TSA treatment was able to restore enhancer activity fully in macrophage precursor cells, but not in non-macrophage lineage cells. These results suggest (i) that the transcription factor complement of multipotent progenitor cells is similar to that of lysozyme-expressing cells and (ii) that developmental regulation of the -3.9 kb enhancer is mediated by the interplay of repressing and activating factors that respond to or initiate changes in the chromatin acetylation state.
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PMID:Identification of factors mediating the developmental regulation of the early acting -3.9 kb chicken lysozyme enhancer element. 1171 4

The chicken lysozyme (cLys) locus has been shown to contain all of the cis-elements necessary for position-independent and tissue-specific expression entirely within a 24-kb region defined by general DNase I sensitivity and flanked by matrix attachment regions. As such, it has been viewed as an example of a functional chromatin domain, which is structurally and functionally isolated from neighbouring chromatin. We report here the identification and characterisation of the chicken glioma-amplified sequence (cGas41) locus, which though widely expressed, is contained entirely within the lysozyme chromatin domain. The cGas41 transcript encodes a putative transcription factor, starts 207 bp downstream of the cLys polyadenylation site and is preceded by a CpG island with proposed dual promoter/origin function. The location and differential expression of cGas41 compels re-evaluation of the accumulated literature on the lysozyme domain, and represents an example of two unrelated, differentially expressed vertebrate genes coexisting in the same functional chromatin domain.
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PMID:The chicken lysozyme chromatin domain contains a second, widely expressed gene. 1178 8


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