The availability of the complete sequence of the Bacillus subtilis chromosome (F. Kunst et al., Nature 390:249-256, 1997) makes possible the construction of genome-wide DNA arrays and the study of this organism on a global scale. Because we have a long-standing interest in the effects of scoC on late-stage developmental phenomena as they relate to aprE expression, we studied the genome-wide effects of a scoC null mutant with the goal of furthering the understanding of the role of scoC in growth and developmental processes. In the present work we compared the expression patterns of isogenic B. subtilis strains, one of which carries a null mutation in the scoC locus (scoC4). The results obtained indicate that scoC regulates, either directly or indirectly, the expression of at least 560 genes in the B. subtilis genome. ScoC appeared to repress as well as activate gene expression. Changes in expression were observed in genes encoding transport and binding proteins, those involved in amino acid, carbohydrate, and nucleotide and/or nucleoside metabolism, and those associated with motility, sporulation, and adaptation to atypical conditions. Changes in gene expression were also observed for transcriptional regulators, along with sigma factors, regulatory phosphatases and kinases, and members of sensor regulator systems. In this report, we discuss some of the phenotypes associated with the scoC mutant in light of the transcriptome changes observed.

Cosmid clones containing 250 kilobases of genomic DNA from the human amylase gene cluster have been isolated. These clones contain seven distinct amylase genes which appear to comprise the complete multigene family. By sequence comparison with the cDNAs, we have identified two pancreatic amylase genes and three salivary amylase genes. Two truncated pseudogenes were also recovered. Intergenic distances of 17 to 22 kilobases separate the amylase gene copies. Within the past 10 million years, duplications, gene conversions, and unequal crossover events have resulted in a very high level of sequence similarity among human amylase gene copies. To identify sequence elements involved in tissue-specific expression and hormonal regulation, the promoter regions of the human amylase genes were sequenced and compared with those of the corresponding mouse genes. The promoters of the human and mouse pancreatic amylase genes are highly homologous between nucleotide -160 and the cap site. Two sequence elements thought to influence pancreas-specific expression of the rodent genes are present in the human genes. In contrast, similarity in the 5' flanking sequences of the salivary amylase genes is limited to several short sequence elements whose positions and orientations differ in the two species. Some of these sequence elements are also associated with other parotid-specific genes and may be involved in their tissue-specific expression. A glucocorticoid response element and a general enhancer element are closely associated in several of the amylase promoters.

Genetic studies have demonstrated that salivary and pancreatic amylase genes are closely linked in human and mouse. To analyze the arrangement of genes within the amylase cluster, a library of YBR mouse genomic DNA was cloned in the cosmid vector pJB8. Clones containing amylase genes were identified by hybridization with amylase cDNA probes. Salivary and pancreatic amylase genes were isolated on separate cosmid clones, but no overlapping clones were evident from the initial screening. A strategy for the rapid isolation of terminal noncoding fragments from the cosmid clones was developed. By using these terminal fragments for chromosome "walking," a map of 78 kilobases of the amylase gene region was constructed. The salivary and pancreatic amylase genes are present within this region in the same 5'-to-3' orientation, separated by 22 kilobases of genomic DNA. A truncated amylase pseudogene is located 10 kilobases downstream from the pancreatic amylase gene.In the human and mouse genomes, distinct genes encode the a-amylases (EC 3.2.1.1) produced in salivary gland and in pancreas (1-4). The extensive sequence homology between salivary (Amy-1) and pancreatic (Amy-2) amylase cDNAs (>90%) indicates that the two genes, Amy-i and Amy-2, are derived from a common ancestral gene (1, 4). Close linkage ofAmy-i and Amy-2 has been established by genetic analysis of the segregation of electrophoretic variants (3, 5). The offspring of more than 1500 meioses have been studied in crosses between inbred mice without observation of recombination between these genes (6). This data suggested that the intergenic distance between Amy-i and Amy-2 may be less than 150 kilobases (kb). Consistent with their close linkage and sequence homology, the amylase genes appear to have undergone "correction" events during mammalian evolution (4, 7).The present study was undertaken to determine the precise organization of amylase gene copies within the mouse multigene cluster. To facilitate analysis of an extended chromosome region, cloning was carried out in a cosmid vector that accommodates 35-to 45-kb inserts of genomic DNA. We chose mouse strain YBR for these studies because of our interest in the independent regulation of its two active pancreatic amylase genes, Amy-2.1 and Amy-2.2 (3, 8-10). We report here the characterization of a 78-kb portion of the amylase gene region that includes one salivary amylase gene, the Amy-2.1 pancreatic amylase gene, and an apparent amylase pseudogene. A subcloning strategy that facilitates chromosome "walking" in cosmid libraries is also described. MATERIALS AND METHODSGenomic DNA libraries were constructed from partially Mbo I-digested DNA isolated from livers of YBR/Ki mice. Restriction fragments of 35-50 kb were size-selected by centrifugation through a neutral sucrose gradient and inserted into' the BamHI site of the cosmid vector pJB8 (11, 12). Recombinant molecules were packaged in vitro into X phage heads and used to infect Escherichia coli strain 490A (13). The yield of infectious phage particles...

SUMMARY We studied the role of renal papillae in the mechanism of increased sodium excretion during acute increase in mean arterial pressure (MAP). Sodium excretion increased dramatically in normal rats after acute increase in MAP by epinephrine (E) infusion (0.4 ^g/min/100g). Glomerular filtration rate (GFR), renal blood flow (RBF), and papillary plasma flow (PPF) remained unchanged after the E administration. To define the role of the medulla in the mechanism of pressure-induced natriuresis, experiments were performed in a group of rats 8 to 12 days after the development of papillary necrosis induced by bromoethylamine hydrobromide. Urinary sodium and fractional sodium excretions were 2.00 ± 0.34 /iEq/ min and 2.37 ± 0.53% (n = 7), respectively, in papillary necrosis rats infused with saline. Administration of E to papillary necrosis rats, however, failed to increase both urinary sodium (2.89 ± 0.61 /xEq/min) and fractional sodium (FE Nl , 2.82 ± 0.63%, n = 6) excretions despite a marked increase in MAP (129 vs 150 mm Hg, p < 0.01). The RBF increased slightly after E infusion (4.42 vs 3.24 ml/min/100 g, p < 0.05), but the GFR was not different between the control (0.39 ± 0.05 ml/min/100 g, n = 7) and the Etreated rats (0.43 ± 0.06, n = 6). Failure to increase sodium excretion during acute increase in MAP was not due to the decreased GFR, since control rats with bilateral partial nephrectomy were able to increase sodium excretion from 1.92 ± 0.33to7.76 ± 1.63/xEq/min(p< 0.01) after E infusion. These findings, therefore, suggest that renal papillae play a major role in the mechanism of natriuresis during acute increase in MAP. T HE mechanism of natriuresis that accompanies acute elevation of renal perfusion pressure has been studied extensively; however it remains undetermined. Previous studies have indicated that acute increase in perfusion pressure is associated with an increase in intratubular pressure and a decrease in fractional sodium reabsorption in the proximal tubule.

Renal tubular reabsorption of phosphate was studied in unanesthetized spontaneously hypertensive (SHR) and normotensive Wistar Kyoto rats (WKY). Three-hour-urinary phosphate excretions of 12-week-old-male SHR (53.0 +/- 13.1 micrograms/mg creatinine, N = 7) and 14-week-old female SHR (81.8 +/- 12.8, N = 8) were significantly lower than those of age sex-matched WKY (12 week, 435.8 +/- 73.2, N = 8, P less than 0.01; 14 week, 423.3 +/- 75.9, N = 8, P less than 0.01). Renal clearances were performed in 14-week-old female rats after an overnight fast. The serum phosphate concentration of SHR (4.61 +/- 0.25 mg/dl) was lower than that of WKY (5.44 +/- 0.14, P less than 0.02). Urinary phosphate excretion (U Pi V = 6.85 +/- 1.83 micrograms/min) and fractional phosphate excretion (FE Pi = 13.7 +/- 2.3%) of SHR were lower than those of WKY (U Pi V = 15.9 +/- 1.87 micrograms/min, P less than 0.01, FE Pi 22.8 +/- 2.6%, P less than 0.02). Acute hyperventilation could not account for the lower excretion of phosphate in SHR, since arterial pH and PCO2 were not different between WKY and SHR. The low renal phosphate clearance of SHR was noted at a very early age; the U Pi V of 5-week-old SHR was already lower than that of WKY. Maximum tubular phosphate reabsorption (TmPi) was studied in the 12-week-old SHR and WKY after acute thyroparathyroidectomy. TmPi of SHR (241 +/- 3.0 micrograms/ml GFR/100 g, N = 7) was greater than that of WKY (204 +/- 7.0, N = 8, P less than 0.01). However, the differences in 3-hr urinary phosphate excretion and clearances of phosphate were abolished between SHR and WKY after their blood pressures were lowered by chronic hydralazine treatment. The results indicate that hypertension is responsible for the lower renal phosphate clearances in SHR.