Role of the 6-20 disulfide bridge in the structure and activity of epidermal growth factor

Two synthetic analogues of murine epidermal. growth factor, [Abu6, 20] mEGF4-48 (where Abu denotes amino-butyric acid) and [G1, M3, K21, H40] mEGF1-48, have been investigated by NMR spectroscopy. [Abu6, 20] mEGF4-48 was designed to determine the contribution of the 6-20 disulfide bridge to the structure and function of mEGF The overall structure of this analogue was similar to that of native mEGF, indicating that the loss of the 6-20 disulfide bridge did not affect the global fold of the molecule. Significant structural differences were observed near the N-terminus, however, with the direction of the polypeptide chain between residues four and nine being altered such that these residues were now located on the opposite face of the main beta-sheet from their position in native mEGF Thermal denaturation experiments also showed that the structure of [Abu6, 20] mEGF4-48 was less stable than that of mEGF. Removal of this disulfide bridge resulted in a significant loss of both mitogenic activity in Balb/c 3T3 cells and receptor binding on A431 cells compared with native mEGF and mEGF4-48, implying that the structural changes in [Abu6, 20] mEGF4-48, although limited to the N-terminus, were sufficient to interfere with receptor binding. The loss of binding affinity probably arose mainly from steric interactions of the dislocated N-terminal region with part of the receptor binding surface of EGF [G1, M3, K21, H40] mEGF1-48 was also synthesized in order to compare the synthetic polypeptide with the corresponding product of recombinant expression. Its mitogenic activity in Balb/c 3T3 cells was similar to that of native mEGF and analysis of its H-1 chemical shifts suggested that its structure was also very similar to native.

Antipronation taping and temporary orthoses - Effects on tibial rotation position after exercise

Abnormal lower-limb biomechanics-in particular, abnormal pronation of the subtalar joint with concomitant increased internal rotation of the tibia-is one of the major causes of overuse injuries of the lower limb. A randomized, controlled, within-subjects research design (N = 14) was used to investigate the effect of a temporary felt orthosis and an antipronation taping technique to control the transverse tibial rotation position immediately after application and after each of two 10-minute periods of exercise. The results showed that the taping technique was superior to both the orthosis and no intervention in controlling tibial rotation position immediately after application and after 10 minutes of exercise. After 20 minutes of exercise, neither the tape nor the orthosis was significantly superior to the control; however, the trends suggested that some residual control was maintained. Future studies are needed to determine the amount of foot pronation control required to relieve symptoms in a symptomatic population in order to determine the clinical effectiveness of these treatment methods.

The influence of Lys68 in decapeptide agonists of C5a on C5a receptor binding, activation and selectivity

The potent, conformationally biased C5a agonist peptide YSFKPMPLaR (C5a(65-74), Y65, F67, P69, P71, D-Ala73) was used as a template to gain insight into the nature and importance of lysine at position 68 in the peptide-receptor interaction. A panel of YSFKPMPLaR analogs with systematic substitutions for Lys68 was evaluated for C5a receptor (C5aR) binding affinity and activation in two well-characterized assay systems: human polymorphonuclear leukocytes (PMNs) and human fetal artery. In addition, we determined the activity of these new analogs in transfected rat basophilic leukemia (RBL) cells in which the Glu at position 199 of the C5aR (wtGlu199) was replaced by a Gin (C5aR-Gln199) or a Lys (C5aR-Lys199). Our results indicated that Lys68 in YSFKPMPLaR plays an important role in binding the C5aR expressed on PMNs and RBL cells. Furthermore, the data indicated that Lys68 interacted with Glu199 of the C5aR in PMNs and RBL cells. In human fetal artery, however, Lys68 substitutions had little or no effect on activity, which suggested that the receptor conformation may be different in this tissue. Thus, the interaction between Lys68 of the decapeptide agonist and Glu199 of the C5aR may be cell type-specific and may form the molecular basis for tissue-specific responses to C5a agonists.

A cell type-specific constitutive point mutant of the common beta-subunit of the human granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin (IL)-3, and IL-5 receptors requires the GM-CSF receptor alpha-subunit for activation

The high affinity receptor for human granulocyte-macrophage colony-stimulating factor (GM-CSF) consists of a cytokine-specific alpha-subunit (hGMR alpha) and a common signal-transducing beta-subunit (hpc) that is shared with the interleukin-3 and -5 receptors, We have previously identified a constitutively active extracellular point mutant of hpc, I374N, that can confer factor independence on murine FDC-P1 cells but not BAF-B03 or CTLL-2 cells (Jenkins, B. J., D'Andrea, R. J., and Gonda, T. J. (1995) EMBO J. 14, 4276-4287), This restricted activity suggested the involvement of cell type-specific signaling molecules in the activation of this mutant. We report here that one such molecule is the mouse GMR alpha (mGMR alpha) subunit, since introduction of mGMR alpha, but not hGMR alpha, into BAF-B03 or CTLL-2 cells expressing the I374N mutant conferred factor independence, Experiments utilizing mouse/human chimeric GMR alpha subunits indicated that the species specificity lies in the extracellular domain of GMRa. Importantly, the requirement for mGMR alpha correlated with the ability of I374N (but not wild-type hpc) to constitutively associate with mGMRa. Expression of I374N in human factor-dependent UT7 cells also led to factor-independent proliferation, with concomitant up-regulation of hGMR alpha surface expression. Taken together, these findings suggest a critical role for association with GMR alpha in the constitutive activity of I374N.

Relation of TNF-related apoptosis-inducing ligand (TRAIL) receptor and FLICE-inhibitory protein expression to TRAIL-induced apoptosis of melanoma

Past studies have shown that apoptosis mediated by TNF-related apoptosis-inducing ligand (TRAIL) is regulated by the expression of two death receptors [TRAIL receptor 1 (TRAIL-RI) and TRAIL-R2] and two decoy receptors (TRAIL-R3 and TRAIL-R4) that inhibit apoptosis, In previous studies, me have shown that TRAIL but not other members of the tumor necrosis factor family induce apoptosis in approximately two-thirds of melanoma cell lines. Here, we examined whether the expression of TRAIL-R at the mRNA and protein level in a panel of 28 melanoma cell lines and melanocytes correlated with their sensitivity to TRAIL-induced apoptosis, We report that at least three factors appear to underlie the variability in TRAIL-induced apoptosis. (a) Pour of nine cell lines that were insensitive to TRAIL-induced apoptosis failed to express death receptors, and in two instances, lines were devoid of all TRAIL-Rs. Southern analysis suggested this was due to loss of the genes for the death receptors, (b) Despite the presence of mRNA for the TRAIL-R, some of the lines failed to express TRAIL-R protein on their surface. This was evident for TRAIL-RI and more so for the TRAIL decoy receptors TRAIL-R3 and -R4, Studies on permeabilized cells revealed that the receptors were located within the cytoplasm and redistribution from the cytoplasm may represent a posttranslational control mechanism. (c) Surface expression of TRAIL-RI and -R2 (but not TRAIL-R3 and -R4) showed an overall correlation with TRAIL-induced apoptosis. However, certain melanoma cell lines and clones were relatively resistant to TRAIL-induced apoptosis despite the absence of decoy receptors and moderate levels of TRAIL-RI and -R2 expression. This may indicate the presence of inhibitors within the cells, but resistance to apoptosis could not be correlated with expression of the caspase inhibitor FLICE-inhibitory protein. mRNA for another TRAIL receptor, osteoprotegerin, was expressed in 22 of the melanoma lines but not on melanocytes. Its role in induction of apoptosis remains to be studied. These results appear to have important implications for future clinical studies on TRAIL.

Is the Australian wool futures market efficient as a predictor of spot prices?

Giles and Goss (1980) have suggested that, if a futures market provides a forward pricing function, then it is an efficient market. In this article a simple test for whether the Australian Wool Futures market is efficient is proposed. The test is based on applying cointegration techniques to test the Law of One Price over a three, six, nine, and twelve month spread of futures prices. We found that the futures market is efficient for up to a six-month spread, but no further into the future. Because futures market prices can be used to predict spot prices up to six months in advance, woolgrowers can use the futures price to assess when they market their clip, but not for longer-term production planning decisions. (C) 1999 John Wiley & Sons, Inc.

Leaking urine in Australian women: Prevalence and associated conditions

The paper aims to (1) assess the prevalence of leaking urine and to (2) explore associations between leaking urine and a variety of other symptoms, conditions, surgical procedures and life events in three large cohorts of Australian women, who are participants in the Australian Longitudinal Study on Women's Health. Young women aged 18-23 (N = 14,000), mid-age women, 45-50 (N 13,738) and older women, 70-75 (N = 12,417), were recruited randomly from the national HIC/Medicare database. Leaking urine was reported by approximately one in eight young women [estimated prevalence 12.8% (95% CI: 12.2-13.3)] and one in three mid-age women [36.1% (CI: 35.2-37.0)] and older women [35.0% (CI: 34.1-35.9)]. Leaking urine was significantly associated with parity, conditions which increase the pressure on the pelvic floor such as constipation and obesity, past gynecological surgery and conditions which can impact on bladder control. The study showed that fewer than half the women had sought help for the problem and that younger women were less likely to be satisfied with the help available for this problem. Strategies for continence promotion, including opportunistic raising of the issue at the time of cervical screening and pregnancy care are suggested, so that the health and social outcomes of untreated chronic incontinence in women might be improved.

Relationships between movement initiation times and movement-related cortical potentials in Parkinson's disease

Movement-related cortical potentials recorded from the scalp reveal increasing cortical activity occurring prior to voluntary movement. Studies of set-related cortical activity recorded from single neurones within premotor and supplementary motor areas in monkeys suggest that such premovement activity may act to prime activity of appropriate motor units in readiness to move, thereby facilitating the movement response. Such a role of early stage premovement activity in movement-related cortical potentials was investigated by examining the relationship between premovement cortical activity and movement initiation or reaction times. Parkinson's disease and control subjects performed a simple button-pressing reaction time task and individual movement-related potentials were averaged for responses with short compared with long reaction times. For Parkinson's disease subjects but not for the control subjects, early stage premovement cortical activity was significantly increased in amplitude for faster reaction times, indicating that there is indeed a relationship between premovement cortical activity amplitude and movement initiation or reaction times. In support of studies of set-related cortical activity in monkeys, it is therefore suggested that early stage premovement activity reflects the priming of appropriate motor units of primary motor cortex, thereby reducing movement initiation or reaction times. (C) 1999 Elsevier Science B.V. All rights reserved.

Phylogeographic differentiation in the mitochondrial control region in the koala, Phascolarctos cinereus (Goldfuss 1817)

The koala, Phascolarctos cinereus, is a geographically widespread species endemic to Australia, with three currently recognized subspecies: P.c. adustus, P.c. cinereus, and P.c. victor. Intraspecific variation in the mitochondrial DNA (mtDNA) control region was examined in over 200 animals from 16 representative populations throughout the species' range. Eighteen different haplotypes were defined in the approximate to 860 bp mtDNA control region as determined by heteroduplex analysis/temperature gradient gel electrophoresis (HDA/TGGE). Any single population typically possessed only one or two haplotypes yielding an average within-population haplotypic diversity of 0.180 +/- 0.003, and nucleotide diversity of 0.16%. Overall, mtDNA control region sequence diversity between populations averaged 0.67%, and ranged from 0% to 1.56%. Nucleotide divergence between populations averaged 0.51%, and ranged from 0% to 1.53%. Neighbour-joining methods revealed limited phylogenetic distinction between geographically distant populations of koalas, and tentative support for a single evolutionarily significant unit (ESU). This is consistent with previous suggestions that the morphological differences formalized by subspecific taxonomy may be interpreted as clinal variation. Significant differentiation in mtDNA-haplotype frequencies between localities suggested that little gene now currently exists among populations. When combined with microsatellite analysis, which has revealed substantial differentiation among koala populations, we conclude that the appropriate short-term management unit (MU) for koalas is the local population.

Late Quaternary cycles of mangrove development and decline on the north Australian continental shelf

Mangrove communities in the Australian tropics presently occur as narrow belts of vegetation in estuaries and on sheltered, muddy coasts. Palynological data from continental shelf and deep-sea cores indicate a long-term cyclical component of mangrove development and decline at a regional scale, which can be linked to specific phases of late Quaternary sealevel change. Extensive mangrove development, relative to today, occurs during periods of marine transgression, whereas very diminished mangrove occurs during marine regressions and during rarer periods of relative sea-level stability. Episodes of flourishing mangrove cannot be linked to phases of humid climate, as has been suggested in studies elsewhere. Rather, the cycle of expansion and decline of mangrove communities on a grand scale is explained in terms of contrasting physiographic settings characteristic of continental-shelf coasts during transgressive and regressive phases, in particular by the existence, or lack, of well-developed tidal estuaries. Copyright (C) 1999 John Wiley & Sons, Ltd.

Movement-related potentials in Parkinson's disease: External cues and attentional strategies

Hypokinetic movement can be greatly improved in Parkinson's disease patients by the provision of external cues to guide movement. It has recently been reported, however, that movement performance in parkinsonian patients can be similarly improved in the absence of external cues by using attentional strategies, whereby patients are instructed to consciously attend to particular aspects of the movement which would normally be controlled automatically. To study the neurophysiological basis of such improvements in performance associated with the use of attentional strategies, movement-related cortical potentials were examined in Parkinson's disease and control subjects using a reaction time paradigm. One group of subjects were explicitly instructed to concentrate on internally timed responses to anticipate the presentation of a predictably timed go signal. Other subjects were given no such instruction regarding attentional strategies. Early-stage premovement activity of movement-related potentials was significantly increased in amplitude and reaction times were significantly faster for Parkinson's disease subjects when instructed to direct their attention toward internally generating responses rather than relying on external cues. It is therefore suggested that the use of attentional strategies may allow movement to be mediated by less automatic and more conscious attentional motor control processes which may be less impaired by basal ganglia dysfunction, and thereby improve movement performance in Parkinson's disease.

Evolution of the genetic covariance between male and female components of mate recognition: An experimental test

The evolution of a positive genetic correlation between male and female components of mate recognition systems will result as a consequence of assortative mating and, in particular, is central to a number of theories of sexual selection. Although the existence of such genetic correlations has been investigated in a number of taxa, it has yet to be shown that such correlations evolve and whether they may evolve as rapidly as suggested by sexual selection models. In this study, I used a hybridization experiment to disrupt natural mate recognition systems and then observed the subsequent evolutionary dynamics of the genetic correlation between male and female components for 56 generations in hybrids between Drosophila serrata and Drosophila birchii. The genetic correlation between male and female components evolved from 0.388 at generation 5 to 1.017 at generation 37 and then declined to -0.040 after a further 19 generations. These results indicated that the genetic basis of the mate recognition system in the hybrid populations evolved rapidly. The initial rapid increase in the genetic correlation was consistent with the classic assumption that male and female components will coevolve under sexual selection. The subsequent decline in genetic correlation may be attributable to the fixation of major genes or, alternatively, may be a result of a cyclic evolutionary change in mate recognition.

Information internalization and hurdle rates in small and medium enterprise internationalization

We investigate the role of information in the internationalization of small and medium enterprises (SMEs). Information internalization is fundamentally antecedent to SME internationalization and is being facilitated increasingly by recent important trends. We offer a conceptual explanation and related propositions on information internalization, emphasizing hurdle rate theory for ascertaining the acceptability of firms' internationalization projects.

Participation in physical activities by older Australians: a review of the social psychological benefits and constraints

Participation in physical activities has been found to be an important factor in contributing to a healthy lifestyle. Research has found strong relationships between participation in regular physical activity and the prevention of disease, while its relationship to the psychological and social dimensions have been neglected. Recently however, several studies have found causal relationships between physical activity and improved mood state, reduced anxiety, reduced depression, and increased social support. Despite this, surveys indicate that participation levels in physical activities are declining among older Australians, with the exceptions of walking and gardening. This paper also examines constraints to participation in leisure programs, such as lack of time, poor health, fear of crime, the financial cost and the lack of a partner to participate with. A number of strategies have been suggested to overcome these constraints.

Novel vascular graft grown within recipient's own peritoneal cavity

A method by which to overcome the clinical symptoms of atherosclerosis is the insertion of a graft to bypass an artery blocked or impeded by plaque. However, there may be insufficient autologous mammary artery for multiple or repeat bypass, saphenous vein may have varicose degenerative alterations that can lead to aneurysm in high-pressure sites, and small-caliber synthetic grafts are prone to thrombus induction and occlusion. Therefore, the aim of the present study was to develop an artificial blood conduit of any required length and diameter from the cells of the host for autologous transplantation. Silastic tubing, of variable length and diameter, was inserted into the peritoneal cavity of rats or rabbits. By 2 weeks, it had become covered by several layers of myofibroblasts, collagen matrix, and a single layer of mesothelium. The Silastic tubing was removed from the harvested implants, and the tube of living tissue was everted such that it now resembled a blood vessel with an inner lining of nonthrombotic mesothelial cells (the intima), with a media of smooth muscle-like cells (myofibroblasts), collagen, and elastin, and with an outer collagenous adventitia. The tube of tissue (10 to 20 mm long) was successfully grafted by end-to-end anastomoses into the severed carotid artery or abdominal aorta of the same animal in which they were grown. The transplant remained patent for at least 4 months and developed structures resembling elastic lamellae. The myofibroblasts gained a higher volume fraction of myofilaments and became responsive to contractile agonists, similar to the vessel into which they had been grafted. It is suggested that these nonthrombogenic tubes of living tissue, grown in the peritoneal cavity of the host, may be developed as autologous coronary artery bypass grafts or as arteriovenous access fistulae for hemodialysis patients.