No abstract

Background Chronic lymphocytic leukemia (CLL) is the most frequent, and still incurable, form of leukemia in the Western World. It is widely accepted that cancer results from an evolutionary process shaped by the acquisition of driver mutations which confer selective growth advantage to cells that harbor them. Clear examples are missense mutations in classic RAS genes (KRAS, HRAS and NRAS) that underlie the development of approximately 13% of human cancers. Although autonomous B cell antigen receptor (BCR) signaling is involved and mutations in many tumor suppressor genes and oncogenes have been identified, an oncogenic driver gene has not still been identified for CLL. Methods Conditional knock-in mice were generated to overexpress wild type RRAS2 and prove its driver role. RT-qPCR analysis of a human CLL sample cohort was carried out to measure RRAS2 transcriptional expression. Sanger DNA sequencing was used to identify a SNP in the 3’UTR region of RRAS2 in human CLL samples. RNAseq of murine CLL was carried out to identify activated pathways, molecular mechanisms and to pinpoint somatic mutations accompanying RRAS2 overexpression. Flow cytometry was used for phenotypic characterization and shRNA techniques to knockdown RRAS2 expression in human CLL. Results RRAS2 mRNA is found overexpressed in its wild type form in 82% of the human CLL samples analyzed (n = 178, mean and median = 5-fold) as well as in the explored metadata. A single nucleotide polymorphism (rs8570) in the 3’UTR of the RRAS2 mRNA has been identified in CLL patients, linking higher expression of RRAS2 with more aggressive disease. Deliberate overexpression of wild type RRAS2 in mice, but not an oncogenic Q72L mutation in the coding sequence, provokes the development of CLL. Overexpression of wild type RRAS2 in mice is accompanied by a strong convergent selection of somatic mutations in genes that have been identified in human CLL. R-RAS2 protein is physically bound to the BCR and mediates BCR signals in CLL. Conclusions The results indicate that overexpression of wild type RRAS2 is behind the development of CLL.

To gain better insight into the insulin secretory activity of fetal b cells in response to glucose, the expression of glucose transporter 2 (GLUT-2), glucokinase and mitochondrial glycerol phosphate dehydrogenase (mGDH) were studied. Expression of GLUT-2 mRNA and protein in pancreatic islets and liver was signi®cantly lower in fetal and suckling rats than in adult rats. The glucokinase content of fetal islets was signi®cantly higher than of suckling and adult rats, and in liver the enzyme appeared for the ®rst time on about day 20 of extrauterine life. The highest content of hexokinase I was found in fetal islets, after which it decreased progressively to the adult values. Glucokinase mRNA was abundantly expressed in the islets of all the experimental groups, whereas in liver it was only present in adults and 20-day-old suckling rats. In fetal islets, GLUT-2 and glucokinase protein and their mRNA increased as a function of increasing glucose concentration, whereas reduced mitochondrial citrate synthase, succinate dehydrogenase and cytochrome c oxidase activities and mGDH expression were observed. These ®ndings, together with those reported by others, may help to explain the decreased insulin secretory activity of fetal b cells in response to glucose.

This study was conducted to determine the effects of triiodothyronine (T3) and bovine growth hormone (bGH) on the expression of glucose transporter-2 (GLUT-2) and of glucokinase (GK) from pancreatic islets of fetal and adult rats. Incubation of both sets of pancreatic islets with T3 did not modify GLUT-2 mRNA levels, but did reduce the content of GLUT-2 protein, while it reduced the expression of GK mRNAs in fetal and adult pancreatic islets. Treatment of fetal and adult pancreatic islets with 1 microg/ml bGH did not alter the expression of GLUT-2 mRNAs, but significantly increased GLUT-2 protein levels in adult islets by 50%. Also, bGH had no effect on the GK mRNA content of fetal and adult pancreatic islets whereas, in contrast, there was a significant reduction in the amount of GK protein in fetal islets cultured with that hormone but not in those corresponding to adult rats. These findings suggest that T3 and bGH are able to modulate the expression of GLUT-2 and GK mRNAs and proteins in pancreatic islets in a manner different from that in the liver, as previously reported by others. In addition, both hormones produced different responses in fetal and in adult pancreatic islets.