Experiments were designed to test the effect of introns on gene expression in transgenic mice. Four different pairs of gene constructs, which were identical except that one member of each pair lacked all introns, were compared for expression of mRNA after introduction into the murine germ line by microiqjection of fertilized eggs. The expression of two chimeric genes, made by fusing either the mouse metallothionein I or the rat elastase 1 promoter/enhancer to the rat growth hormone gene, was assayed in fetal liver or pancreas, respectively, while two natural genes, an oligonucleotidemarked mouse metallothionein I gene and the human .8-globin gene, were assayed in fetal liver. In each case there was, on average, 10-to 100-fold more mRNA produced from the intron-containing construct. Moreover, mRNA levels were proportional to the relative rates of transcription that were measured in isolated nuclei. However, when the expression of the two mouse metallothionein I gene-based constructs was tested after transfection into cultured cells, little difference was observed. These observations suggest that introns play a role in facilitating transcription of microinjected genes and that this effect may be manifest only on genes exposed to developmental influences.Most mammalian genes coding for mRNA are interrupted by noncoding sequences known as introns, many of which are larger than the exons; thus, the entire gene may span tens or even hundreds of kilobases (1). For some genes, introns clearly separate functional or structural domains of the proteins encoded by the exons. This observation, and the fact that similar domains can be found within different proteins, has led to the suggestion that one function of introns may be to accelerate the evolution of proteins with different properties (2, 3). Introns also allow differential joining of exons during splicing, which can -result in the synthesis of variant proteins with new properties (4, 5).The possibility that introns may be necessary for efficient processing and transport of mRNA to the cytoplasm has been examined. Hamer and Leder (6) showed that a series of simian virus 40 viruses that contained various combinations of simian virus 40 and mouse j3-globin splice sites produced stable mRNAs only if at least one splice site, derived from either the virus or P-globin, was retained. In another experiment, a simian virus 40 mutant missing a late gene intron failed to produce stable transcripts but could be rescued by addition of an heterologous intron (7,8). These early observations suggested that splicing was obligatory for mRNA accumulation in the cytoplasm. As a consequence, the first-generation cDNA expression vectors usually included a heterologous intron in addition to promoter and polyadenylylation sequences (9). Subsequently, it was discovered that deletion of introns does not always result in loss of mRNA production. For instance, several viral genes, including those for ElA protein (10) and polyoma large tumor (T) antigen (11) function without introns. ...
BackgroundThe mechanisms underlying chronic obstructive pulmonary disease (COPD) remain unclear. MicroRNAs (miRNAs or miRs) are small non-coding RNA molecules that modulate the levels of specific genes and proteins. Identifying expression patterns of miRNAs in COPD may enhance our understanding of the mechanisms of disease. A study was undertaken to determine if miRNAs are differentially expressed in the lungs of smokers with and without COPD. miRNA and mRNA expression were compared to enrich for biological networks relevant to the pathogenesis of COPD. Methods Lung tissue from smokers with no evidence of obstructive lung disease (n¼9) and smokers with COPD (n¼26) was examined for miRNA and mRNA expression followed by validation. We then examined both miRNA and mRNA expression to enrich for relevant biological pathways.
The identification of genes with selective expression in specific organs or cell types provides an entry point for understanding biological processes that occur uniquely within a particular tissue. Using a subtraction approach designed to identify genes preferentially expressed in specific tissues, we have identified prostase, a human serine protease with prostate-restricted expression. The prostase cDNA encodes a putative 254-aa polypeptide with a conserved serine protease catalytic triad and an amino-terminal pre-propeptide sequence, indicating a potential secretory function. The genomic sequence comprises five exons and four introns and contains multiple copies of a chromosome 19q-specific minisatellite repeat. Northern analysis indicates that prostase mRNA is expressed in hormonally responsive normal and neoplastic prostate epithelial tissues, but not in prostate stromal constituents. Prostase shares 35% amino acid identity with prostate-specific antigen (PSA) and 78% identity with the porcine enamel matrix serine proteinase 1, an enzyme involved in enamel matrix degradation and with a putative role in the disruption of intercellular junctions. Radiation-hybrid-panel mapping localized prostase to chromosome 19q13, a region containing several other serine proteases, including protease M, pancreatic͞renal kallikrein hK1, and the prostate-specific kallikreins hK2 and hK3 (PSA). The sequence homology between prostase and other well-characterized serine proteases suggests several potential functional roles for the prostase protein that include the degradation of extracellular matrix and the activation of PSA and other proteases.
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