Mitsuko Mori

Structure, chromosomal localization and expression of mouse genes encoding type III Reg, RegIIIα, RegIIIβ, RegIIIγ

Abstract Reg (regenerating gene), first isolated from a rat regenerating islet cDNA library, is expressed in regenerating islet β-cells. Recently, it has been revealed that Reg and Reg-related genes constitute a multigene family, Reg family, which consists of three subtypes (type I, II, III) based on the primary structures of the encoded proteins of the genes. In mouse, type I and type II Reg genes (i.e. RegI and RegII gene) have so far been isolated. In the present study, the complete nucleotide (nt) sequences of the cDNAs and genes encoding murine type III Reg (regenerating gene product), RegIIIα, RegIIIβ and RegIIIγ were determined. RegIIIα, RegIIIβ and RegIIIγ encode 175-, 175- and 174-amino acid (aa) proteins, respectively, with 60–70% homology. All three genes are composed of six exons and five introns spanning approx. 3 kb, and exhibit distinctive structural features unique for members of the Reg gene family. All the mouse Reg genes, RegIIIα, RegIIIβ, RegIIIγ, RegI and RegII, are assigned to the adjacent site of chromosome 6C by fluorescence in situ hybridization (FISH). RegIIIα, RegIIIβ and RegIIIγ were expressed weakly in pancreas, strongly in intestinal tract, but not in hyperplastic islets, whereas both RegI and RegII were expressed in hyperplastic islets. These results suggest that genes of the mouse Reg family are derived from a common ancestor gene by several gene duplications, and have obtained divergency in expression and function in the process of genetic evolution.

Human REG family genes are tandemly ordered in a 95-kilobase region of chromosome 2p12

Reg, first isolated from a rat regenerating islet cDNA library, is expressed in regenerating islet β‐cells. Recently, it has been revealed that Reg and Reg‐related genes constitute a multigene family, the Reg family. In human, the four REG family genes, i.e., REG Iα, REG Iβ, REG‐related sequence (RS) and HIPIPAP, have so far been isolated. In this study, we analyzed YAC clones containing the four genes and performed two‐color FISH to determine the map order of the genes. The human REG family genes are tandemly ordered in the 95‐kbp DNA region of chromosome 2p12 as follows: 2cen‐HIPIPAP‐RS‐REG Iα‐REG Iβ‐ptel.

Assignment of CD38, the gene encoding human leukocyte antigen CD38 (ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase), to chromosome 4p15

CD38 has been used as a phenotype marker of lymphocyte differentiation. Recently, we have demonstrated that cyclic ADP-ribose can be synthesized and hydrolyzed by CD38 and acts as a second messenger in insulin secretion from pancreatic beta-cells. We have mapped the CD38 gene to human chromosome 4p15 by fluorescence in situ hybridization.

Telomerase activity in human chorionic villi and placenta determined by TRAP and in situ TRAP assay

Telomerase activity (TA) was analysed in human chorionic villi and placenta in normal and abnormal pregnancy using the telomeric repeat amplification protocol (TRAP) and in situ TRAP assay. Twenty chorionic villi specimens and 25 placenta specimens from normal pregnancies were examined as well as placenta specimens from 10 cases of intrauterine growth retardation (IUGR; nine asymmetric and one symmetric). TA was detected in 18 of the 20 (90 per cent) chorionic villi specimens and in 18 of the 25 (72 per cent) placenta specimens from normal pregnancy. However, no or only weak TA was exhibited in the placenta specimens of the nine asymmetric IUGR cases. In situ TRAP assay detected TA in trophoblastic cells from normal pregnancy, but not in trophoblastic cells from cases of asymmetric IUGR.

Up-regulation of cyr61 in vascular smooth muscle cells of spontaneously hypertensive rats

In the present study, we applied a fluorescent differential display method to mRNAs from aortae of spontaneously hypertensive rats (SHRs), stroke-prone spontaneously hypertensive rats (SPSHRs), and their parental strain, Wistar Kyoto rats (WKYRs), to identify the genes involved in the development of hypertension. Through this screen we came across a gene that is consistently up-regulated in hypertensive rats. Nucleotide sequence determination of the corresponding cDNA revealed that the gene is the rat orthologue of cyr61. Northern blot analysis showed that cyr61 expression increases in SHR and SPSHR before the onset of hypertension and is sustained thereafter at higher levels than in age-matched WKYRs. In situ hybridization analysis demonstrated that cyr61 is expressed strongly in smooth muscle cells (SMCs) in media of SHR and SPSHR but not WKYR aorta. Fluorescent in situ hybridization mapped the cyr61 gene to rat chromosome 1p12–13, which is located in close proximity to a recently defined quantitative trait locus including NHE3 Na+/H+ exchanger. Overexpression of the cyr61 gene in stably transfected rat SMC line A7r5 caused rather inhibitory effects on the proliferation and DNA and protein synthesis. Our results thus demonstrate for the first time that cyr61 can also act as a growth inhibitor in SMC of genetically hypertensive rats. This may reveal a new route for investigation of the pathogenesis of hypertension.

Genotyping of Angiotensin-Converting Enzyme and Angiotensinogen Polymorphisms with the LightCycler System

The renin-angiotensin system regulates blood pressure, and maintains electrolyte homeostasis in humans [1]. In this system, angiotensinogen (AGT) is catalyzed by renin to form angiotensin I, which is then cleaved by angiotensin-converting enzyme (ACE) to yield angiotensin II, a potent vasopressor and effector on renal function [1]. Recently, several studies have focused on the correlation between physiological disorders and the genetic variation of peptides in the reninangiotensin system. A specific mutation in the angiotensinogen gene was reported to be associated with essential hypertension [2,3]. Individuals with homozygous deletion alleles of ACE were reported to have a higher level of serum ACE and an increased risk of ischemic heart disease, sudden death, left ventricular hypertrophy, increased blood glucose levels, diabetic nephropathy, and premature death [4–7]. Detection of mutations in genes that constitute the renin-angiotensin system may be important in the prevention and control of disorders in the cardiovascular system, glucose metabolism, and urinary function.