Late transcription of bacteriophage Mu, which results in the expression of phage morphogenetic functions, is dependent on Mu C protein. Earlier experiments indicated that Mu late RNAs originate from four promoters, including the previously characterized mom promoter. Si nuclease protection experiments were used to map RNA 5' ends in the three new regions. Transcripts were initiated at these points only in the presence of C and were synthesized in a rightward direction on the Mu genome. Amber mutant marker rescue analysis of plasmid clones and limited DNA sequencing demonstrated that these new promoters are located between C and lys, upstream of I, and upstream of P within the N gene. A comparison of the promoter sequences upstream from the four RNA 5' ends yielded two conserved sequences: the first (tA . . cT, where capital and lowercase letters indicate 100 and 75% base conservation, respectively), at approximately -10, shares some similarity with the consensus Escherichia coli C70 -10 region, while the second (ccATAAc CcCPuG/Cac, where Pu indicates a purine), in the -35 region, bears no resemblance to the E. coli -35 consensus. We propose that these conserved Mu late promoter consensus sequences are important for C-dependent promoter activity. Plasmids containing transcription fusions of these late promoters to lacZ exhibited C-dependent lI-galactosidase synthesis in vivo, and C was the only Mu product needed for this transactivation. As expected, the late promoter-lacZ fusions were activated only at late times after induction of a Mu prophage. The C-dependent activation of lacZ fusions containing only a few bases of the 5' end of Mu late RNA and the presence of altered promoter sequences imply that C acts at the level of transcription initiation.Viruses have evolved a number of regulatory schemes to ensure efficient production of progeny particles during lytic development. A common strategy is to sequentially activate and inhibit transcription of different sets of viral genes. In general, genes for DNA replication are transcribed early, while those for'virion formation and release are expressed later. Activation of new sets of genes often involves utilization of new promoter sequences which are recognized by new virus-specific polymerases or by altered forms of the host polymerase. For example, Escherichia coli phages T7 and N4 synthesize their own RNA polyrmerases (12, 18), whereas Bacillus phage SPOl and E. coli phage T4 produce phage-encoded sigma factors which substitute for the host sigma subunit (14, 42). E. coli phage lambda uses two additional mechanisms: it produces accessory DNA-binding proteins, cI and cII, which enhance initiation by host RNA polymerase holoenzyme at specific promoters (55,69), and it modifies the host RNA polymerase complex with phage proteins N and Q to achieve antitermination and thus elongation of short constitutive leader transcripts (1,19
Previously, we reported cloning and characterization of the mouse gene, epitheliasin. In the present work we cloned the cDNA of the full‐length human orthologue and characterized its gene including 2 kb of 5′ flanking sequence. Analysis of epitheliasin gene expression in adult tissues shows that it is expressed as 3.4 kb and 2 kb transcripts. The major 3.4 kb transcript is observed in the following order: prostate > colon > small intestine > pancreas > kidney > lung > liver. Epitheliasin transcripts in fetal tissues are observed only in kidney and lung. In situ hybridization analysis of tissues revealed that epitheliasin was preferentially expressed in epithelial cells. The gene consists of 14 exons and 13 introns based on comparison with its cDNA sequence. In the 5′ flanking region, we identified two transcription start sites and three CpG islands encompassing a number of potential regulatory elements including SP1, SREBP, GRE/PRE and ERE. The region upstream of the transcription sites lacks a TATA box but contains an initiator‐like element as well as a downstream promoter‐like element. In vitro experiments with lymph node carcinoma of prostate (LNCaP) cells revealed that the epitheliasin gene was induced by androgens and the induction was not blocked by cycloheximide indicating that the induction required no intermediate protein factors. Immunoprecipitation analysis showed that androgens strongly increased epitheliasin protein levels.
Dipeptidyl-peptidase I, a lysosomal cysteine proteinase, is important in intracellular degradation of proteins and appears to be a central coordinator for activation of many serine proteinases in immune/inflammatory cells. Little is known about the molecular genetics of the enzyme. In the present investigation the gene for dipeptidyl-peptidase I was cloned and characterized. The gene spans approximately 3.5 kilobases and consists of two exons and one intron. The genomic organization is distinct from the complex structures of the other members of the papain-type cysteine proteinase family. By fluorescence in situ hybridization, the gene was mapped to chromosomal region 11q14.1-q14.3. Analysis of the sequenced 5 -flanking region revealed no classical TATA or CCAAT box in the GC-rich region upstream of cap site. A number of possible regulatory elements that could account for tissue-specific expression were identified. Northern analyses demonstrated that the dipeptidyl-peptidase I message is expressed at high levels in lung, kidney, and placenta, at moderate to low levels in many organs, and at barely detectable levels in the brain, suggesting tissue-specific regulation. Among immune/inflammatory cells, the message is expressed at high levels in polymorphonuclear leukocytes and alveolar macrophages and their precursor cells. Treatment of lymphocytes with interleukin-2 resulted in a significant increase in dipeptidyl-peptidase I mRNA levels, suggesting that this gene is subjected to transcriptional regulation. The results provide initial insights into the molecular basis for the regulation of human dipeptidylpeptidase I.
We report the isolation of a cDNA encoding a novel murine serine proteinase, epitheliasin. The cDNA spans 1753 bp and encodes a mosaic protein with a calculated molecular mass of 53 529 Da. Its domains include a cytoplasmic tail, a type II transmembrane domain, a low-density lipoprotein receptor class A domain, a cysteine rich scavenger receptor-like domain and a serine proteinase domain. The proteinase portion domain shows 46^53% identity with mouse neurotrypsin, acrosin, hepsin and enteropeptidase. The gene, located in the telomeric region in the long arm of mouse chromosome 16, consists of 14 exons and 13 introns and spans approximately 18 kb. Epitheliasin is expressed primarily in the apical surfaces of renal tubular and airway epithelial cells.z 2000 Federation of European Biochemical Societies.
Studies were designed to explore the possibility that sulfated polysaccharides had the potential to prevent human leukocyte elastase (HLE)-induced lung injury. Arteparon (GAGPS), heparin, heparan sulfate, chondroitin sulfate, and dextran sulfate, but not dextran, inhibited HLE-mediated hydrolysis of succinyl-ala2-val-pNA. GAGPS, used as a paradigmatic sulfated polysaccharide, was a potent inhibitor of elastolysis in vitro. GAGPS given intratracheally prevented acute injury and emphysema in hamsters when administered up to 8 h before HLE insufflation. The results suggest that sulfated polysaccharides may be potent inhibitors of HLE-mediated lung injury.
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