Re-utilization of pyrimidine nucleotides during rat liver regeneration

Abstract

The changes in the specific radioactivities of the pool of total acid-soluble uridine nucleotides and of uridine and cytidine components of total cellular and nuclear RNA were monitored in regenerating rat liver for 12 days after partial hepatectomy. Evidence is presented for the re-utilization of pyrimidine nucleotides derived from cytoplasmic RNA degradation for the synthesis of new RNA. The extent of recycling was assessed and the true rate of rRNA turnover determined more accurately. The reutilization of the uridine components of RNA was 7.0%/day during the proliferative and 3.2%/day during the post-proliferative phase, whereas that of the cytidine nucleotides was more pronounced (9.6%/day and 18.1%/day respectively). The results reveal the existence of partial compartmentalization of pyrimidine ribonucleoside triphosphate pools in the nucleus and cytoplasm of rat liver cells.

Pyrimidine nucleotide metabolism in rat hepatocytes: evidence for compartmentation of nucleotide pools

“…The opposite is true for the nucleus-localized synthesis of RNA. This is in agreement with results found by others in various cell types [26,[31][32][33][34]. Compartmentation of the de novo and salvage pathways has also been described in the nucleus [16,30,35].…”

“…The concentration of the molecules in the overflow pool determines the extent of uptake and feedback inhibition and thus the flow through the salvage route. Published results by other authors concerning pyrimidine compartmentation fit this model [18,19,[29][30][31].…”

Heterogeneous Nuclear Ribonucleoprotein (hnRNP) E1 Binds to hnRNP A2 and Inhibits Translation of A2 Response Element mRNAs

“…To determine whether reduced GFP expression in the presence of hnRNP E1 was due to RNA degradation, we used a nuclear/cytoplasmic partition assay based on the observation that intact RNA is excluded from the nucleus, whereas products of RNA degradation diffuse into the nucleus (Nikolov and Dabeva, 1985;Ifrim, personal communication). Fluorescently tagged GFP-A2RE RNA injected with hnRNP E1 ( Figure 5E, b and e) and without hnRNP E1 ( Figure 5E, a and d) into the cytoplasm of B104 cells was excluded from the nucleus up to 6 h after injection, indicating that hnRNP E1 does not increase degradation of GFP-A2RE RNA.…”

“…To analyze stability of the injected RNA, a nuclear/cytoplasmic partition assay, based on the observations of Nikolov and Dabeva (1985), was used. B104 cells were injected with fluorescent cyanine (Cy)-5-UTP-labeled GFP-A2RE RNA with or without recombinant hnRNP E1 protein or RNAse A. GFP-A2RE RNA concentration in the injection mixes was 50 ng/l and that of hnRNP E1 and RNAse A, 0.06 g/l and 0.2 g/l, respectively.…”

Determination of in vivo RNA kinetics using RATE-seq

“…Pulse-chase labeling with 4tU (Munchel et al 2011) represents a promising approach to estimating mRNA degradation rates. However, internal recycling of labeled nucleotides (Puckett et al 1975;Nikolov and Dabeva 1985) may result in an incomplete chase thereby confounding the estimation of mRNA degradation rates. Alternatively, comparative Dynamic Transcriptome Analysis (cDTA) (Sun et al 2012) (an updated version of Dynamic Transcriptome Analysis [DTA]) (Miller et al 2011) estimates rates of mRNA degradation by determining the ratio of labeled to total RNA using hybridization to a DNA microarray at a single time point following addition of 4tU.…”