Imcorporation of Orotic Acid into Nucteotides and RNA in Mouse Organs during 60 Minutes

Lewan, Lillemor; Petersen, Inga; Yngner, Torsten

doi:10.1515/bchm2.1975.356.1.425

Cited by 16 publications

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References 2 publications

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“…Orotate (orotic acid, 6-carboxyuracil), a normal intermediate in pyrimidine metabolism, is converted to uridine 5 0 -triphosphate (UTP), which is utilized for the synthesis of RNA and for the synthesis of uridine 5 0 -diphosphohexoses and uridine 5 0 -diphosphohexosamines (UDP sugars) [1][2][3]. Since UDP sugars are used in the glycosylation of the basement membrane collagen, increased bioavailability of these compounds is involved in the pathogenesis of renal hypertrophy with concomitant thickening of the glomerular and tubular basement membranes observed in human diabetes mellitus [4,5].…”

Section: Introductionmentioning

confidence: 99%

Human urate transporter 1 (hURAT1) mediates the transport of orotate

et al. 2011

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Orotate is a precursor of pyrimidine synthesis. The kidney uses exogenous orotate for the synthesis of uridine diphosphosugars, which are used in the glycosylation of collagen in glomerular and tubular basement membranes. Orotate uptake occurs in the liver and kidney, but its molecular mechanism is largely unknown. Since orotate has been shown to be a substrate of the renal urate/anion exchanger in brush border membrane vesicle studies, we investigated whether human URAT1 (hURAT1) mediates the transport of orotate using HEK293 cells expressing hURAT1 (HEK-hURAT1). hURAT1 mediated a time- and dose-dependent uptake of orotate (K (m) 5.2 μM). hURAT1-mediated [(3)H]orotate transport was inhibited strongly by non-labeled (cold) orotate and the uricouric agent benzbromarone, and moderately inhibited by urate, nicotinate, and another uricouric agent, probenecid. This is the first report demonstrating that hURAT1 mediates the transport of orotate. hURAT1 may function as one of the entrance pathways in renal proximal tubular cells.

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Section: Introductionmentioning

confidence: 99%

Human urate transporter 1 (hURAT1) mediates the transport of orotate

et al. 2011

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“…It has been shown by biochemical method that RNA in various tissues may be derived from labeled uridine (22)(23)(24)(25)(26)(27)(28) and labeled orotic acid (22,23,29,30) . In the small intestine of the rat, however, Raisonnier et al (31) reported incorporation of ['°C]uridine, but of only traces of [ t 'C]orotic acid, which implies that orotic acid was not a suitable precursor for studies of RNA in rat small intestine .…”

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confidence: 99%

Radioautographic visualization of differences in the pattern of [3H]uridine and [3H]orotic acid incorporation into the RNA of migrating columnar cells in the rat small intestine.

Uddin¹,

Altmann²,

Leblond³

1984

The Journal of Cell Biology

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The epithelium of rat small intestine was radioautographed to examine whether RNA is synthesized by the salvage pathway as shown after [3H]uridine injection or by the de novo pathway as shown after [3H]orotic acid injection. The two modes of RNA synthesis were thus investigated during the migration of columnar cells from crypt base to villus top, and the rate of synthesis was assessed by counting silver grains over the nucleolus and nucleoplasm at six levels along the duodenal epithelium--that is, in the base, mid, and top regions of the crypts and in the base, mid, and top regions of the villi. Concomitant biochemical analyses established that, after injection of either [5- 3H]uridine or [5-3H]orotic acid: (a) buffered glutaraldehyde fixative was as effective as perchloric acid or trichloracetic acid in insolubilizing the nucleic acids of rat small intestine; (b) a major fraction of the nucleic acid label was in RNA, that is, 91% after [3H]uridine and 72% after [3H]orotic acid, with the rest in DNA; and (c) a substantial fraction of the RNA label was in poly A+ RNA (presumed to be messenger RNA). In radioautographs of duodenum prepared after [3H] uridine injection, the count of silver grains was high over nucleolus and nucleoplasm in crypt base cells and gradually decreased at the upper levels up to the villus base. In the rest of the villus, the grain count over the nucleolus was negligible, while over the nucleoplasm it was low but significant. After [3H]-orotic acid injection, the number of silver grains over the nucleolus was negligible at all levels, whereas over the nucleoplasm the number was low in crypt cells, but high in villus cells with a peak in mid villus. The interpretation is that, except for a small amount of label incorporated into DNA from either precursor by crypt cells, the bulk of the label is incorporated into RNA as follows. In the crypts, cells make almost exclusive use of uridine, that is, of the salvage pathway, for the synthesis of ribosomal RNA in the nucleolus and of messenger and transfer RNA in the nucleoplasm. However, when cells pass from crypt to villus, they mainly utilize orotic acid--i.e., the de novo pathway--for the synthesis of messenger and transfer RNA within the nucleoplasm.

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“…Orotic acid has been actively utilized for the synthesis of pyrimidine nucleotides in the liver and kidney of rats [15,16]. Other organs characterized by rapid nucleic acid synthesis…”

Section: Resultsmentioning

confidence: 99%

“…Orotic acid has been actively utilized for the synthesis of pyrimidine nucleotides in the liver and kidney of rats [15,16]. Other organs characterized by rapid nucleic acid synthesis expt (a) 8.1 x lo6 counts/min and in expt (b) 12.3 x lo6 counts/min of [2-'4C]orotic acid was injected per The specific activity of DNA cytosine and thymine on the 11th and 14th day after the administration of [2-'4C]orotic acid is higher than the specific activity of the pyrimidine components of DNA on the 6th day after the administration of the labeled precursor.…”

Section: Resultsmentioning

confidence: 99%

Reutilization of Preformed Pyrimidines for the Synthesis of DNA in Rat Liver and Kidney

Seifert

2005

European Journal of Biochemistry

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The specific activities of rat liver and kidney DNA pyrimidines increase in animals which have had [2-14C]-orotic acid administered several days before killing. This phenomenon is caused by the reutilization of the preformed labeled pyrimidine components originating mainly from degraded RNAs of the organism.The quantitative ratio of the de now synthesis/salvage pathways (reutilization) of the low-molecular-weight building components of macromolecules is difficult to follow in experiments in vivo. Part of the radioactivity of the low-molecularweight components derived from the degraded macromolecules is reincorporated into the de novo synthesized macromolecules. This makes the exact determination of their halflife in the cell almost impossible.The pyrimidine components originating predominantly from the degradation of cellular RNAs are also reutilized [l ,2]. Whereas the ribonucleic acids (depending on their kind) undergo an intracellular turnover, the DNA of the cell is metabolically stable and remains in the cell nucleus during the whole life of the cell. The period of existence of the rat liver hepatocyte is assumed to be 160 -400 days [3,4] and the percentage of new liver DNA synthesized per day is similar to the percentage increase of liver weight [3]. Omitting the repair synthesis which could lead to an incorporation of the labeled precursor into the DNA, then this increase represents the de novo synthesis, preceding the proliferation.Labeled thymidine is the compound most often used in studies of DNA biosynthesis in vivo. This deoxyribonucleoside is formed by the removal of the phosphate group from thymidylic acid; it is therefore a catabolic product apart from the de novo pathways of the biosynthesis of pyrimidine deoxynucleoside phosphates. The utilization of thymidine for the synthesis can be influenced by numerous factors, such as metabolic changes in blood plasma, transport across the cell membrane, rate of phosphorylation, concentration changes in the pool caused by the administration of various drugs, etc. [5 -1 I]. It has therefore been recommended with experiments in vivo to determine simultaneously the degree of DNA biosynthesis by the administration of labeled orotic acid [12], which is a precursor of all pyrimidine components de novo.If the reutilization of the pyrimidine components did not exist, then the decrease of radioactivity of prelabeled DNA pyrimidines would be proportional to the growth of the organs. The present paper, however, shows an increase of the specific activity of the liver and kidney DNA pyrimidine components in animals administered [2-'4C]orotic acid several days before killing. This increase of the radioactivity of pyrimidine nucleotides can serve as a measure of the reutilization (mainly of the cytidine components) for the synthesis of DNA under both normal and specific conditions. MATERIALS AND METHODSMale rats (Wistar strain) weighing 180-190 g at the beginning of the experiments were used. The pelleted diet (produced by Velaz, Prague) was composed of the following ing...

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Imcorporation of Orotic Acid into Nucteotides and RNA in Mouse Organs during 60 Minutes

Cited by 16 publications

References 2 publications

Human urate transporter 1 (hURAT1) mediates the transport of orotate

Human urate transporter 1 (hURAT1) mediates the transport of orotate

Radioautographic visualization of differences in the pattern of [3H]uridine and [3H]orotic acid incorporation into the RNA of migrating columnar cells in the rat small intestine.

Reutilization of Preformed Pyrimidines for the Synthesis of DNA in Rat Liver and Kidney

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