Structure, chromosomal localisation and expression of the murine dominant negative helix-loop-helix Id4 gene

In the elderly patients a greater loss of teeth is frequently seen in combination with periodontal diseases. This article presents an overview of the changes of oral structures during ageing as well as immunology, physical and psychological aspects. Also summarised are complications by systemic illnesses, drug medications and periodontal alterations. If possible illnesses and medications are taken into consideration, there should be no special problem for the elderly patients.     

The chromosomal distribution of mouse odorant receptor genes

Odorant receptors (ORs) on nasal olfactory sensory neurons are encoded by a large multigene family. Each member of the family is expressed in a small percentage of neurons that are confined to one of several spatial zones in the nose but are randomly distributed throughout that zone. This pattern of expression suggests that when the sensory neuron selects which OR gene to express it may be confined to a particular zonal gene set of several hundred OR genes but select from among the members of that set via a stochastic mechanism. Both locus-dependent and locus-independent models of OR gene choice have been proposed. To investigate the feasibility of these models, we determined the chromosomal locations of 21 OR genes expressed in four different spatial zones. We found that OR genes are clustered within multiple loci that are broadly distributed in the genome. These loci lie within paralogous chromosomal regions that appear to have arisen by duplications of large chromosomal domains followed by extensive gene duplication and divergence. Our studies show that OR genes expressed in the same zone map to numerous loci; moreover, a single locus can contain genes expressed in different zones. These findings raise the possibility that OR gene choice may be locus-independent or involve consecutive stochastic choices.     

Reconstitution of the functional mouse oncostatin M (OSM) receptor: molecular cloning of the mouse OSM receptor beta subunit

Oncostatin M (OSM) is a member of the interleukin-6 (IL-6) family of cytokines that share the gp130 receptor subunit. Of these family members, leukemia inhibitory factor (LIF) is most closely related to OSM, and various overlapping biologic activities have been described between human LIF and OSM (hLIF and hOSM). Two types of functional hOSM receptors are known: the type I OSM receptor is identical to the LIF receptor that consists of gp130 and the LIF receptor beta subunit (LIFRbeta), and the type II OSM receptor consists of gp130 and the OSM receptor beta subunit (OSMRbeta). It is thus conceivable that common biologic activities between hLIF and hOSM are mediated by the shared type I receptor and OSM-specific activities are mediated by the type II receptor. However, in contrast to the human receptors, recent studies have demonstrated that mouse OSM (mOSM) does not activate the type I receptor and exhibits unique biologic activity. To elucidate the molecular structure of the functional mOSM receptor, we cloned a cDNA encoding mOSMRbeta, which is 55.5% identical to the hOSMRbeta at the amino acid level. mOSM-responsive cell lines express high-affinity mOSM receptors, as well as mOSMRbeta, whereas embryonic stem cells, which are responsive to LIF but not to mOSM, do not express mOSMRbeta. mOSMRbeta alone binds mOSM with low affinity (kd = 13.0 nmol/L) and forms a high-affinity receptor (kd = 606 pmol/L) with gp130. Ba/F3 transfectants expressing both mOSMRbeta and gp130 proliferated in response to mOSM, but failed to respond to LIF and human OSM. Thus, the cloned mOSMRbeta constitutes an essential and species-specific receptor component of the functional mOSM receptor. Reminiscent of the colocalization of the mOSM and mLIF genes, the mOSMRbeta gene was found to be located in the vicinity of the LIFRbeta locus in the proximal end of chromosome 15.     

Consistent loss of functional transforming growth factor beta receptor expression in murine plasmacytomas

SPR Online (http:@www.pedrad.org) is a recently developed digital representation of the Society for Pediatric Radiology (SPR) that enables physicians to access pertinent information and services on the Internet. SPR Online was organized on the basis of the five main services of the SPR, which include Administration, Patient Care, Education, Research, and Meetings. For each service, related content from the SPR was digitized and placed onto SPR Online. Usage over a 12-month period was evaluated with server log file analysis. A total of 3,209 users accessed SPR Online, viewing 11,246 pages of information. A wide variety of information was accessed, with that from the Education, Administration, and Meetings services being the most popular. Fifteen percent of users came from foreign countries. As a virtual professional society, SPR Online greatly enhances the power and scope of the SPR and has proved to be a popular resource, meeting the diverse information needs of an international community of pediatric radiologists.     

Evidence for uniparental, paternal expression of the human GABAA receptor subunit genes, using microcell-mediated chromosome transfer

We have constructed mouse A9 hybrids containing a single normal human chromosome 15, via microcell-mediated chromosome transfer. Cytogenetic and DNA-polymorphic analyses identified mouse A9 hybrids that contained either a paternal or maternal human chromosome 15. Paternal specific expression of the known imprinted genes SNRPN (small nuclear ribonucleoprotein-associated polypeptide N gene) and IPW (imprinted gene in the Prader-Willi syndrome region) was maintained in the A9 hybrids. Using this system, we first demonstrated that human GABAAreceptor subunit genes, GABRB3 , GABRA5 and GABRG3 , were expressed exclusively from the paternal allele and that E6-AP (E6-associated protein or UBE3A ) was biallelically expressed. Moreover, the 5' portion of the GABRB3 gene was found to be hypermethylated on the paternal allele. Our data imply that GABAAreceptor subunit genes are imprinted and are possible candidates for Prader-Willi syndrome, and that this human monochromosomal hybrid system enables the efficient analysis of imprinted loci.     

Saturation mutagenesis of the beta subunit of the human granulocyte-macrophage colony-stimulating factor receptor shows clustering of constitutive mutations, activation of ERK MAP kinase and STAT pathways, and differential beta subunit tyrosine phosphorylation

The high-affinity receptors for human granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5 are heterodimeric complexes consisting of cytokine-specific alpha subunits and a common signal-transducing beta subunit (hbetac). We have previously demonstrated the oncogenic potential of this group of receptors by identifying constitutively activating point mutations in the extracellular and transmembrane domains of hbetac. We report here a comprehensive screen of the entire hbetac molecule that has led to the identification of additional constitutive point mutations by virtue of their ability to confer factor independence on murine FDC-P1 cells. These mutations were clustered exclusively in a central region of hbetac that encompasses the extracellular membrane-proximal domain, transmembrane domain, and membrane-proximal region of the cytoplasmic domain. Interestingly, most hbetac mutants exhibited cell type-specific constitutive activity, with only two transmembrane domain mutants able to confer factor independence on both murine FDC-P1 and BAF-B03 cells. Examination of the biochemical properties of these mutants in FDC-P1 cells indicated that MAP kinase (ERK1/2), STAT, and JAK2 signaling molecules were constitutively activated. In contrast, only some of the mutant beta subunits were constitutively tyrosine phosphorylated. Taken together, these results highlight key regions involved in hbetac activation, dissociate hbetac tyrosine phosphorylation from MAP kinase and STAT activation, and suggest the involvement of distinct mechanisms by which proliferative signals can be generated by hbetac.     

Functional comparison of bovine, murine, and human beta2-microglobulin: interactions with murine MHC I molecules

Fetal calf serum is a well known source of bovine beta2-microglobulin (beta2m) which can exchange with endogenous beta2m from, as well as promote peptide binding to, class I major histocompatibility (MHC I) molecules on cells cultured in vitro. Recombinant bovine beta2m was expressed and purified for direct functional comparison to human and murine beta2m for interactions with murine MHC I molecules H-2Kb, Db, Kd, Ld, and Dd. Bovine and human beta2m were equivalent in stabilizing MHC I heavy chains and facilitating peptide loading, suggesting similar affinities for murine MHC I heavy chains. The activity of murine beta2m was significantly weaker, consistent with previous work that demonstrated the lower affinity of murine human beta2m for murine heavy chains compared to human beta2m. Analysis of bovine beta2m in fetal calf serum revealed ten-fold higher concentrations than in adult bovine serum, levels shown to significantly affect MHC I stability and peptide loading. The ramifications for the study of MHC I molecules from cells in culture and the evolutionary implications of the higher affinity interactions of human and bovine beta2m are discussed.     

Structural and functional characterization of human potassium channel subunit beta 1 (KCNA1B)

Voltage-activated Shaker-related potassium channels (kv1) consist of alpha and beta subunits. We have analysed the structure of the human KCNA1B (hKv beta 1) gene. KCNA1B is > 250 kb in size and encodes at least three Kv beta 1 splice variants. The Kv beta 1 open reading frame is divided into 14 exons. In contrast, genes coding for family members of KCNA (Kv 1 alpha) subunits are markedly smaller and have intronless open reading frames. The expression of Kv 1 alpha and Kv beta mRNA was compared in Northern blots of poly(A+) RNA isolated from various human brain tissues. The results suggest an intricate and cell-specific regulation of Kv 1 alpha and Kv beta mRNA synthesis such that distinct combinations of alpha and beta subunits would occur in different nuclei of the brain. The splice variants hKv beta 1.1 and hKv beta 1.2 were functionally characterized in coexpression studies with hKv 1.5 alpha subunits in 293 cells. It is shown that the confer rapid inactivation on hKv 1.5 channels with different potencies. This may be due to differences in their amino terminal sequences and/or inactivating domains. It is also shown that the amino terminal Kv beta 1.1 and Kv 1.4 alpha inactivating domains compete with each other, probably for the binding to the same receptor site(s) on Kv 1 alpha-subunits.     

The cloning and chromosomal mapping of two novel human opioid-somatostatin-like receptor genes, GPR7 and GPR8, expressed in discrete areas of the brain

Following the cloning of the opioid receptors mu, kappa, and delta, we conducted a search for related receptors. Using oligonucleotides based on the opioid and also the structurally related somatostatin receptors, we amplified genomic DNA using the polymerase chain reaction and isolated fragments of novel G protein-coupled receptor genes. Two of these gene fragments designated clones 12 and 11 were used to isolate the full-length genes. The intronless coding sequences of these genes, named GPR7 and GPR8, shared 70% identity with each other, and each shared significant similarity with the sequences encoding transmembrane regions of the opioid and somatostatin receptors. GPR7 was mapped to chromosome 10q11.2-q21.1 and GPR8 to chromosome 20q13.3. Northern blot analysis using human mRNA demonstrated expression of GPR7 mainly in cerebellum and frontal cortex, while GPR8 was located mainly in the frontal cortex. In situ hybridization revealed expression of GPR7 in the human pituitary. A partial sequence of the mouse orthologue of GPR7 was obtained, and in situ hybridization demonstrated expression in discrete nuclei of brain, namely suprachiasmatic, arcuate, and ventromedial nuclei of hypothalamus. A stable cell line expressing the GPR7 gene was created, but expression levels of the receptor were low. The available pharmacology indicated binding to several opioid drugs such as bremazocine, levorphanol, and beta-FNA, but not to the opioid receptor subtype-selective mu, delta, or kappa agonists.     

Functional analysis of myocardial performance in murine hearts overexpressing the human beta 2-adrenergic receptor

Transgenic mice overexpressing the human beta 2-adrenergic receptor gene were compared with wild mice type in terms of cardiac function, using a modified work-performing isolated murine heart preparation and on-line computer analysis. A preload-dependent experiment was performed, in which venous return was gradually increased in 5 mmHg increments from 5 mmHg to 25 mmHg. At each preload, aortic flow, left atrial pressure and aortic pressure were measured in all hearts, and from these parameters stroke volume, contractility, and cardiac index (cardiac output divided by body weight in g) were calculated and compared between groups. At increasing preload levels, the heart rates ranged from 322 beats/min (+/-29) to 369 beats/min (+/-39) in control mice and from 469 beats/min (+/-36) to 540 beats/min (+/-39) in transgenic mice. Cardiac index increased from 138 microliters/min/g (+/-13) and 48 microliters/min/g (+/-5) for transgenic and control mice, respectively at 5 mmHg preload to 262 microliters/min/g (+/-51) and 167 microliters/min/g (+/-15), respectively at 20 mmHg preload. The contractility in the transgenic mice were significantly increased at lower preload levels compared to control mice (1420 mmHg/s +/- 204 v 1187 mmHg/s +/- 127). An increase in myocardial adrenergic receptor density (100-200 fold) leads to significantly higher indices of cardiac function in transgenic mice compared to control mice. The increased heart rate leading to a positive inotropic effect in the hearts of transgenic mice is, at least in part, due to the overexpression of adrenergic receptors. These findings suggest a possible alternative method of establishing a positive chronotropic and inotropic state without the use of pharmacological agents.     

Evolution of the structure and chromosomal distribution of histidine biosynthetic genes

A database of more than 100 histidine biosynthetic genes from different organisms belonging to the three primary domains has been analyzed, including those found in the now completely sequenced genomes of Haemophilus influenzae, Mycoplasma genitalium, Synechocystis sp., Methanococcus jannaschii, and Saccharomyces cerevisiae. The ubiquity of his genes suggests that it is a highly conserved pathway that was probably already present in the last common ancestor of all extant life. The chromosomal distribution of the his genes shows that the enterobacterial histidine operon structure is not the only possible organization, and that there is a diversity of gene arrays for the his pathway. Analysis of the available sequences shows that gene fusions (like those involved in the origin of the Escherichia coli and Salmonella typhimurium hisIE and hisB gene structures) are not universal. In contrast, the elongation event that led to the extant hisA gene from two homologous ancestral modules, as well as the subsequent paralogous duplication that originated hisF, appear to be irreversible and are conserved in all known organisms. The available evidence supports the hypothesis that histidine biosynthesis was assembled by a gene recruitment process.     

The effect of track structure on the induction of chromosomal aberrations in murine cells

Maternal pre-pregnancy obesity is a risk factor for pre-eclampsia (proteinuric hypertension in pregnancy) among North American and European women. We studied the relationship between maternal obesity and risk of pre-eclampsia among Zimbabwean women. A case-control study was conducted at Harare Maternity Hospital, Harare, Zimbabwe, between June 1995 and April 1996. Study participants were 144 women with pre-eclampsia and 194 normotensive women serving as controls. Maternal weight, height and mid-arm circumference were measured and recorded during study participants' postpartum hospital admission. Maternal mid-arm circumference, considered to be relatively stable during pregnancy among women of developing countries, was used as the primary indicator of maternal pre-pregnancy obesity. Logistic regression procedures were used to estimate odds ratios and 95% confidence intervals. There were linear trends in risk of pre-eclampsia with increasing mid-arm circumference, increasing weight and increasing body mass index. After adjusting for potential confounding factors, women in the highest quintile for mid-arm circumference (28-39 cm) were 4.4 times more likely to have had their pregnancy complicated by pre-eclampsia than women in the lowest quintile (21-23 cm). Odds ratios of similar magnitude were observed for the other anthropometric measures. To our knowledge, this is the first study to demonstrate a positive association between maternal obesity and pre-eclampsia risk in a black African population. Biological mechanisms thought to explain this relatively consistent epidemiological finding include endothelial cell injury, possibly resulting from hyperlipidaemia.     

The e subunit gene of murine F1F0-ATP synthase. Genomic sequence, chromosomal mapping, and diet regulation

Genomic sequences encoding murine Lfm1, whose predicted protein sequence is 96% and 98% similar to bovine and rat F1F0-ATP synthase e subunits (respectively), have been amplified from BALB/cByJ DNA, cloned, and sequenced. The 1.1-kilobase gene has 3 introns and 4 exons, and its coding sequence differs by two nucleotides compared to the previously published BALB/cHnn Lfm1 cDNA sequence. A PstI restriction site polymorphism in intron 2 between C57BL/6J and Mus spretus was used to map this gene to Chromosome 5 near D5Mit9. Related sequences were mapped on Chromosomes 8, 11, and 2 unlinked loci on Chromosome 2 using Southern blot analyses with the 1. 1-kilobase gene as probe. Previous studies from this laboratory indicated that the Lfm1/e subunit was regulated by the level of dietary fat and carbohydrate. Northern hybridization analyses demonstrated that e subunit mRNA abundance showed statistically significant differences (p < 0.025) between hearts of BALB/c mice fed 3% and those fed 20% corn oil for 2 weeks and in liver (p < 0. 05) from the same animals. Significant differences were also observed in hepatic and heart mRNA expression at different times after eating in animals subjected to a fast/refeed regimen. The implications of the high degree of sequence similarity to the e subunit for rat and bovine F1F0-ATP synthase and its regulation by diet are discussed.     

Molecular cloning and regional distribution of a human proton receptor subunit with biphasic functional properties

BACKGROUND: At present no satisfying treatment for subfoveal choroidal neovascularization (CNV) secondary to age-related macular degeneration (AMD) is available. Visual results after successful surgical removal of subfoveal CNV are disappointing. This has been explained by a primary dysfunction of the retinal pigment epithelium (RPE) in the macular region and the surgical trauma to the RPE in patients with AMD. Therefore, Machemer and Steinhorst developed a technique for macular translocation after surgical removal of subfoveal CNV. We report our first experiences with this technique in patients with subfoveal CNV secondary to AMD. METHODS: Seven patients aged between 71 and 83 years with subfoveal CNV were included in the study. Visual acuity of the fellow eyes was below 20/400. All patients underwent pars plana vitrectomy. Retinal detachment was produced by subretinal infusion of balanced salt solution and a 360 degrees retinotomy at the base of the vitreous was performed. After removal of the CNV, retinal rotation and reattachment, the retina bordering the retinotomy was coagulated with endolaser photocoagulation. Silicone oil was used as temporary tamponade. RESULTS: In all patients the subfoveal CNV was removed and the macula was translocated by a 15 degrees-45 degrees rotation onto functional RPE. The mean duration of follow-up was 11 +/- 3 months. Initial visual acuity ranged from 20/80 to hand movements. Final visual acuity was 20/100 to 20/400. Initially all patients complained of tilted vision. During follow-up the rotation of the image regressed and was well tolerated by all patients. Complications included the development of retinal detachment in three patients after silicone oil removal, development of a macula pucker, and a significant increase of lens opacity in the phakic eyes. CONCLUSION: In our series rapid improvement of visual function was observed in one patient only, even if the macula appeared ophthalmoscopically and angiographically normal. Vitreoretinal complications occurred frequently during follow-up.     

A reporter mutation approach shows incorporation of the "orphan" subunit beta3 into a functional nicotinic receptor

We have investigated whether the neuronal nicotinic subunit beta3 can participate in the assembly of functional recombinant receptors. Although beta3 is expressed in several areas of the central nervous system, it does not form functional receptors when expressed heterologously together with an alpha or another beta nicotinic subunit. We inserted into the human beta3 subunit a reporter mutation (V273T), which, if incorporated into a functional receptor, would be expected to increase its agonist sensitivity and maximum response to partial agonists. Expressing the mutant beta3(V273T) in Xenopus oocytes together with both the alpha3 and the beta4 subunits resulted in the predicted changes in the properties of the resulting nicotinic receptor when compared with those of alpha3 beta4 receptors. This indicated that some of the receptors incorporated the mutant beta3 subunit, as part of a "triplet" alpha3 beta4 beta3 receptor. The proportion of triplet receptors was dependent on the ratios of the alpha3:beta4:beta3 cRNA injected. We conclude that, like the related alpha5 subunit, the beta3 subunit can form functional receptors only if expressed together with both alpha and beta subunits.