Structure and regulation of the envoplakin gene

Envoplakin, a member of the plakin family of proteins, is a component of desmosomes and the epidermal cornified envelope. To understand how envoplakin expression is regulated, we have analyzed the structure of the mouse envoplakin gene and characterized the promoters of both the human and mouse genes. The mouse gene consists of 22 exons and maps to chromosome 11E1, syntenic to the location of the human gene on 17q25. The exon-intron structure of the mouse envoplakin gene is common to all members of the plakin family: the N-terminal protein domain is encoded by 21 small exons, and the central rod domain and the C-terminal globular domain are coded by a single large exon. The C terminus shows the highest sequence conservation between mouse and human envoplakins and between envoplakin and the other family members. The N terminus is also conserved, with sequence homology extending to Drosophila Kakapo. A region between nucleotides -101 and 288 was necessary for promoter activity in transiently transfected primary keratinocytes. This region is highly conserved between the human and mouse genes and contains at least two different positively acting elements identified by site-directed mutagenesis and electrophoretic mobility shift assays. Mutation of a GC box binding Sp1 and Sp3 proteins or a combined E box and Krüppel-like element interacting with unidentified nuclear proteins virtually abolished promoter activity. 600 base pairs of the mouse upstream sequence was sufficient to drive expression of a beta-galactosidase reporter gene in the suprabasal layers of epidermis, esophagus, and forestomach of transgenic mice. Thus, we have identified a regulatory region in the envoplakin gene that can account for the expression pattern of the endogenous protein in stratified squamous epithelia.     

Oligosaccharides related to xyloglucan: synthesis and X-ray crystal structure of methyl alpha-L-fucopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1-->2)-alpha-D-x ylopyranoside and the synthesis of methyl alpha-L-fucopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1-->2)-beta-D-xy lopyranoside

Trisaccharides, methyl alpha-L-fucopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1-->2)-alpha-D-xy lopyranoside and methyl alpha-L-fucopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1-->2)-beta-D-xyl opyranoside, which are related to the side chain of xyloglucan have been synthesised. The beta-galactopyranosyl linkage of each was constructed using silver trifluoromethanesulfonate-promoted glycosylations of 2-O-acetyl-3,4,6-tri-O-benzyl-beta-D-galactopyranosyl chloride and the corresponding anomer of methyl 3,4-tri-O-benzyl-D-xylopyranoside. The resulting disaccharides were deacetylated and fucosylated using assisted halide reactions with tri-O-benzyl-alpha-L-fucopyranosyl bromide. Hydrogenolytic debenzylation of the resulting protected trisaccharides gave the methyl glycosides of the fucose-containing xyloglucan side chain. The structure of methyl alpha-L-fucopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1-->2)-alpha-D-xy lopyranoside as the monohydrate was confirmed by an X-ray crystallographic study.     

Contribution of Midparental BMI and other determinants of obesity in adult offspring

The aim of this study was to evaluate midparental BMI among intergenerational factors associated with obesity in adult offspring. The data was from an unusual two-generational observational design of 1,477 married couples from Renfrew and Paisley in Scotland who were aged 45-64 years when screened in 1972-1976, and 1,040 sons and 1,298 daughters aged 30-59 years when screened in 1996. BMI was categorized as normal (< 25 kg/m(2)), overweight (25-29.9 kg/m(2)), and obese (> or = 30 kg/m(2)) in offspring and parents. Midparental BMI was defined as the mean of the mother's and father's BMI. Low physical activity, nonsmoking status, higher cholesterol level, and manual social class were all associated with increased BMI in offspring. The effect of reported dietary intake was less clear. Offspring of obese parents (defined by midparental BMI) were over four times more likely to be obese than offspring of normal weight parents. Midparental BMI had a strong effect on offspring BMI, independent of social class, smoking habit, physical activity, and reported dietary intake. Adding midparental BMI to the regression model more than doubled the explained variation of offspring BMI from 7.7 to 17%. Every 1 kg/m(2) increment in midparental BMI was associated with a BMI greater by 0.51 kg/m(2) in offspring. We conclude that midparental BMI is a useful simple tool to predict offspring BMI. Whether it represents genetic or environmental family effects, it is easily ascertained by the individual and could be used in health promotion and clinical settings to target individuals who are at increased risk of becoming obese.     

The prion protein and neuronal zinc homeostasis

Although the prion protein (PrP) is known to be the causative agent of the neurodegenerative transmissible spongiform encephalopathies, its normal cellular function remains elusive. Octapeptide repeats in the N terminus of PrP bind metal ions and are required for the endocytosis of PrP upon exposure of cells to copper or zinc. As the concentration of zinc in the extracellular spaces of the brain is higher than that for copper, we put forward the hypothesis that PrP is involved in neuronal zinc homeostasis; PrP might be involved in transport of zinc into the cell or might act as a zinc sensor. In prion disease, when the protein undergoes a conformational change to the infectious form, this function of PrP in zinc homeostasis might be compromised.     

The basement membrane of hair follicle stem cells is a muscle cell niche

The hair follicle bulge in the epidermis associates with the arrector pili muscle (APM) that is responsible for piloerection ("goosebumps"). We show that stem cells in the bulge deposit nephronectin into the underlying basement membrane, thus regulating the adhesion of mesenchymal cells expressing the nephronectin receptor, α8β1 integrin, to the bulge. Nephronectin induces α8 integrin-positive mesenchymal cells to upregulate smooth muscle markers. In nephronectin knockout mice, fewer arrector pili muscles form in the skin, and they attach to the follicle above the bulge, where there is compensatory upregulation of the nephronectin family member EGFL6. Deletion of α8 integrin also abolishes selective APM anchorage to the bulge. Nephronectin is a Wnt target; epidermal β-catenin activation upregulates epidermal nephronectin and dermal α8 integrin expression. Thus, bulge stem cells, via nephronectin expression, create a smooth muscle cell niche and act as tendon cells for the APM. Our results reveal a functional role for basement membrane heterogeneity in tissue patterning. PAPERCLIP:     

Effect of epinephrine on glucose disposal during exercise in humans: role of muscle glycogen

This study examined the effect of epinephrine on glucose disposal during moderate exercise when glycogenolytic flux was limited by low preexercise skeletal muscle glycogen availability. Six male subjects cycled for 40 min at 59 +/- 1% peak pulmonary O2 uptake on two occasions, either without (CON) or with (EPI) epinephrine infusion starting after 20 min of exercise. On the day before each experimental trial, subjects completed fatiguing exercise and then maintained a low carbohydrate diet to lower muscle glycogen. Muscle samples were obtained after 20 and 40 min of exercise, and glucose kinetics were measured using [6,6-2H]glucose. Exercise increased plasma epinephrine above resting concentrations in both trials, and plasma epinephrine was higher (P < 0.05) during the final 20 min in EPI compared with CON. Muscle glycogen levels were low after 20 min of exercise (CON, 117 +/- 25; EPI, 122 +/- 20 mmol/kg dry matter), and net muscle glycogen breakdown and muscle glucose 6-phosphate levels during the subsequent 20 min of exercise were unaffected by epinephrine infusion. Plasma glucose increased with epinephrine infusion (i.e., 20-40 min), and this was due to a decrease in glucose disposal (R(d)) (40 min: CON, 33.8 +/- 3; EPI, 20.9 +/- 4.9 micromol. kg(-1). min(-1), P < 0.05), because the exercise-induced rise in glucose rate of appearance was similar in the trials. These results show that glucose R(d) during exercise is reduced by elevated plasma epinephrine, even when muscle glycogen availability and utilization are low. This suggests that the effect of epinephrine does not appear to be mediated by increased glucose 6-phosphate, secondary to enhanced muscle glycogenolysis, but may be linked to a direct effect of epinephrine on sarcolemmal glucose transport.