1.Horizontal RNA transfer mediates platelet-induced hepatocyte proliferation.
Kirschbaum M1, Karimian G1, Adelmeijer J1, Giepmans BN2, Porte RJ3, Lisman T4. Blood. 2015 Aug 6;126(6):798-806. doi: 10.1182/blood-2014-09-600312. Epub 2015 Jun 8.
Liver regeneration is stimulated by blood platelets, but the molecular mechanisms involved are largely unexplored. Although platelets are anucleate, they do contain coding or regulatory RNAs that can be functional within the platelet or, after transfer, in other cell types. Here, we show that platelets and platelet-like particles (PLPs) derived from the megakaryoblastic cell line MEG-01 stimulate proliferation of HepG2 cells. Platelets or PLPs were internalized within 1 hour by HepG2 cells and accumulated in the perinuclear region of the hepatocyte. Platelet internalization also occurred following a partial hepatectomy in mice. Annexin A5 blocked platelet internalization and HepG2 proliferation. We labeled total RNA of MEG-01 cells by incorporation of 5-ethynyluridine (EU) and added EU-labeled PLPs to HepG2 cells. PLP-derived RNA was detected in the cytoplasm of the HepG2 cell. We next generated PLPs containing green fluorescent protein (GFP)-tagged actin messenger RNA.
2.A possible cause and remedy for the clinical failure of 5-fluorouracil plus eniluracil.
Spector T1, Cao S. Clin Colorectal Cancer. 2010 Jan;9(1):52-4. doi: 10.3816/CCC.2010.n.007.
BACKGROUND: Eniluracil is a potent inactivator of dihydropyrimidine dehydrogenase (uracil reductase), the enzyme that rapidly catabolizes 5-fluorouracil (5-FU). Although eniluracil in combination with 5-FU was promising in phase I and II studies, in 2 multicenter phase III colorectal cancer studies, eniluracil dosed in a 10-to-1 ratio to 5-FU produced less antitumor benefit than the standard regimen of 5-FU/leucovorin without eniluracil. The current study tested whether the high eniluracil doses caused the clinical failure. The effect of excess eniluracil versus adequate eniluracil on 5-FU antitumor activity was studied in rats bearing large transplanted colon tumors.
3.Exploring RNA transcription and turnover in vivo by using click chemistry.
Jao CY1, Salic A. Proc Natl Acad Sci U S A. 2008 Oct 14;105(41):15779-84. doi: 10.1073/pnas.0808480105. Epub 2008 Oct 7.
We describe a chemical method to detect RNA synthesis in cells, based on the biosynthetic incorporation of the uridine analog 5-ethynyluridine (EU) into newly transcribed RNA, on average once every 35 uridine residues in total RNA. EU-labeled cellular RNA is detected quickly and with high sensitivity by using a copper (I)-catalyzed cycloaddition reaction (often referred to as "click" chemistry) with fluorescent azides, followed by microscopic imaging. We demonstrate the use of this method in cultured cells, in which we examine the turnover of bulk RNA after EU pulses of varying lengths. We also use EU to assay transcription rates of various tissues in whole animals, both on sections and by whole-mount staining. We find that total transcription rates vary greatly among different tissues and among different cell types within organs.
4.Genome-wide technology for determining RNA stability in mammalian cells: historical perspective and recent advantages based on modified nucleotide labeling.
Tani H1, Akimitsu N. RNA Biol. 2012 Oct;9(10):1233-8. doi: 10.4161/rna.22036. Epub 2012 Oct 1.
Changing the abundance of transcripts by regulated RNA degradation is a critical step in the control of various biological pathways. Recently, genome-wide inhibitor-free technologies for determining RNA stabilities in mammalian cells have been developed. In these methods, endogenous RNAs are pulse labeled by uridine analogs [e.g., 4-thiouridine (4sU), 5-etyniluridine (EU) and 5'-bromo-uridine (BrU)], followed by purification of labeled de novo RNAs. These technologies have revealed that the specific half-life of each mRNA is closely related to its physiological function. Genes with short-lived mRNAs are significantly enriched among regulatory genes, while genes with long-lived mRNAs are enriched among housekeeping genes. This review describes the recent progress of experimental procedures for measuring RNA stability.