RNA polymerases of cyanobacteria contain a novel core subunit, y, which is absent from the RNA polymerases of other eubacteria. The genes encoding the three largest subunits of RNA polymerase, including y, have been isolated from the cyanobacterium Anabaena sp. strain PCC 7120. The genes are linked in the order rpoB, rpoCl, rpoC2 and encode the 1, y, and B' subunits, respectively. RNA polymerase in prokaryotes generally consists of a catalytic core of four subunits (W313'a2) and a dissociable sigma factor, which confers promoter specificity (7,16,44). The basic structure of the enzyme was believed to be the same in all eubacteria until the RNA polymerase of the filamentous cyanobacterium Anabaena sp. strain PCC 7120 was purified (37). The cyanobacterial core RNA polymerase was found to contain, in addition to a ,B, a ,3', and two a's, a novel core subunit of 70 kDa designated y, which is absent from the RNA polymerases of other eubacteria. Western immunoblotting with antiserum to -y has shown that a serologically related -y protein is present in over 30 different cyanobacteria, representing the five major taxonomic subgroups (4a, 36). Thus, the -y subunit is a common feature of cyanobacterial RNA polymerases.Anti--y serum cross-reacts with the Escherichia coli ' subunit protein, suggesting homology between these subunits (36). A region of DNA homologous to part of the E. coli gene encoding 3' (rpoC) has been isolated from another cyanobacterium, Nostoc commune UTEX 584 (47). Sequencing of this region showed that homology to E. coli rpoC is split between two linked genes, rpoCl and rpoC2, in the Nostoc DNA (47). The DNAs of several plant chloroplasts have likewise been found to contain regions of sequence homology with E. coli rpoC (19,20,30,38,39). In chloroplasts, the blocks of homology are also distributed between two genes, rpoCl and rpoC2. These genes are linked and are thought to encode the P' and ,B" subunits of chloroplast RNA polymerase, respectively.The exact subunit composition of chloroplast RNA polymerase is unclear. Spinach chloroplast RNA polymerase was found to contain seven prominent polypeptides, some of which were serologically related to subunits of the E. coli RNA polymerase (26). DNA sequences potentially encoding protein equivalents to the E. coli a and ,B subunits (rpoA and * Corresponding author. rpoB) have been found in chloroplast genomes, in addition to the rpoC homologs (19,20,30,39,40). In a few cases, these genes have been shown to be expressed in chloroplasts. The protein product of the rpoA gene has been identified in maize and pea chloroplasts (33,34 Isolation ofAnabaena sp. strain PCC 7120 RNA polymerase genes. The 2.8-and 2.3-kb EcoRI fragments containing parts of the E. coli rpoB and rpoC genes, respectively, were isolated from pGB218 (2), which was a gift from C. Squires. The fragments were labeled with [a-32P]dCTP (3,000 Ci/ mmol; New England Nuclear) by the random priming method (11) and used to probe a Southern blot of Anabaena sp. strain PCC 7120 chromosomal DNA digested ...

. Evidence for increased postprandial distal nephron calcium delivery in hypercalciuric stone-forming patients. Am J Physiol Renal Physiol 295: F1286 -F1294, 2008. First published August 20, 2008 doi:10.1152/ajprenal.90404.2008.-A main mechanism of idiopathic hypercalciuria (IH) in calcium stoneforming patients (IHSF) is postprandial reduction of renal tubule calcium reabsorption that cannot be explained by selective reduction of serum parathyroid hormone levels; the nephron site(s) responsible are not as yet defined. Using fourteen 1-h measurements of the clearances of sodium, calcium, and endogenous lithium during a three-meal day in the University of Chicago General Clinical Research Center, we found reduced postprandial proximal tubule reabsorption of sodium and calcium in IHSF vs. normal subjects. The increased distal sodium delivery is matched by increased distal reabsorption so that urine sodium excretions do not differ, but distal calcium reabsorption does not increase enough to match increased calcium delivery, so hypercalciuria results. In fact, urine calcium excretion and overall renal fractional calcium reabsorption both are high in IHSF vs. normal when adjusted for distal calcium delivery, strongly suggesting a distal as well as proximal reduction of calcium reabsorption. The combination of reduced proximal tubule and distal nephron calcium reabsorption in IHSF is a new finding and indicates that IH involves a complex, presumably genetic, variation of nephron function. The increased calcium delivery into the later nephron may play a role in stone formation via deposition of papillary interstitial apatite plaque. nephrolithiasis; tubule reabsorption; lithium clearance; hypercalciuria; proximal tubule IDIOPATHIC HYPERCALCIURIA (IH) is best described as a state of high urine calcium excretion commonly found in patients who form calcium renal stones and thought to play a causal role in such stones (22). Although gastrointestinal calcium absorption is above normal, offering an obvious mechanism for hypercalciuria, the kidney also participates. After eating common foods, patients with calcium stones and IH (IHSF) reduce their overall renal calcium reabsorption more than well-matched normal control subjects (23). This reduction permits high urine calcium losses despite unchanged serum calcium and calcium filtered load. Although serum parathyroid hormone decreases with meals, levels do so equally in IHSF patients and normal subjects, and therefore cannot be the main mechanism for the reduced tubule calcium reabsorption (23). Because the differential increase in urine calcium occurs without a corresponding difference of urine sodium between IHSF and normal, the cortical thick ascending limb of the loop of Henle was considered an attractive site of reduced calcium reabsorption (1) along with the distal convoluted tubule (8). Since filtered load of calcium did not differ between patients and normal subjects, hypercalciuria essentially arose from the change in tubule reabsorption.Which nephron sites participat...

Regulation of acid-base metabolism maintains the pH of body fluids within a tight range. Urine pH (UpH) is also regulated under normal conditions. Median pH of 24-h urines is ~6, but others have noted that UpH in women is higher than men, which has been attributed to differences in diet. If true, it would help to explain the fact that calcium phosphate stones, which form at higher urine pH, are much more common in women than in men. We studied 14 normal subjects (7 men and 7 women) fed identical meals in a Clinical Research Center. Urine and blood samples were collected during fasting and after meals. UpH of women (6.74 ± 0.11) exceeded that of men (6.07 ± 0.17) fed, but not fasting, and UpH rose significantly with meals in women but not men. Serum and urine total CO rose with meals in women but not men, and in women net acid excretion fell to zero during the fed period. In a general linear model adjusted for age, sex, and weight, net gastrointestinal anion uptake was the main predictor of UpH and was significantly higher in women (3.9 ± 0.6) than men (1.8 ± 0.7) in the fed period. Urine citrate, an anion absorbed by the gastrointestinal tract, was higher in women than men in the fed state, and fractional excretion of citrate was higher in women than men. The higher fed UpH in women is related to a greater absorption of food anions and raises 24-h UpH.

Bergsland KJ, Worcester EM, Coe FL. Role of proximal tubule in the hypocalciuric response to thiazide of patients with idiopathic hypercalciuria. Am J Physiol Renal Physiol 305: F592-F599, 2013. First published May 29, 2013 doi:10.1152/ajprenal.00116.2013The most common metabolic abnormality found in calcium (Ca) kidney stone formers is idiopathic hypercalciuria (IH). Using endogenous lithium (Li) clearance, we previously showed that in IH, there is decreased proximal tubule sodium absorption, and increased delivery of Ca into the distal nephron. Distal Ca reabsorption may facilitate the formation of Randall's plaque (RP) by washdown of excess Ca through the vasa recta toward the papillary tip. Elevated Ca excretion leads to increased urinary supersaturation (SS) with respect to calcium oxalate (CaOx) and calcium phosphate (CaP), providing the driving force for stone growth on RP. Thiazide (TZ) diuretics reduce Ca excretion and prevent stone recurrence, but the mechanism in humans is unknown. We studied the effect of chronic TZ administration on renal mineral handling in four male IH patients using a fixed three meal day in the General Clinical Research Center. Each subject was studied twice: once before treatment and once after 4 -7 mo of daily chlorthalidone treatment. As expected, urine Ca fell with TZ, along with fraction of filtered Ca excreted. Fraction of filtered Li excreted also fell sharply with TZ, as did distal delivery of Ca. Unexpectedly, TZ lowered urine pH. Together with reduced urine Ca, this led to a marked fall in CaP SS, but not CaOx SS. Since CaOx stone formation begins with an initial CaP overlay on RP, by lowering urine pH and decreasing distal nephron Ca delivery, TZ might diminish stone risk both by reducing CaP SS, as well as slowing progression of RP. calcium oxalate; calcium phosphate; idiopathic hypercalciuria; kidney calculi; thiazide THE MOST COMMON METABOLIC abnormality found in calcium (Ca) stone formers is so-called idiopathic hypercalciuria (IH) (40). Patients with IH absorb more Ca from a given meal than normal people (N), but also have abnormally decreased renal tubule Ca reabsorption, so that they often excrete more Ca than they have absorbed, exhibit decreased bone density, and have a propensity for increased fractures (22,42). Elevated urine Ca excretion leads to increased urinary supersaturation (SS) with respect to both calcium oxalate (CaOx) and calcium phosphate (CaP), providing the driving force for kidney stone formation (36). In addition, urine Ca excretion is correlated with the amount of interstitial apatite deposits (called Randall's plaque) found on the surface of the renal papillae (19). These deposits are the attachment sites for most CaOx stones and appear to be critical for stone formation (14). A plausible mechanism by which increased Ca reaches the deep papillae is via washdown from reabsorption in the thick ascending limb; an increase in delivery of Ca to this site would tend to increase reabsorption and washdown into the papilla and potentially increase...

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