Ectodomain shedding and remnant peptide signalling of EGFRs and their ligands

Both receptor tyrosine kinases epidermal growth factor receptors (EGFRs) and their ligands are transmembrane proteins. It has been known that ligand binding activates cytoplasmic tyrosine kinase domains of EGFRs, resulting in the transduction of signals for cell proliferation, migration, differentiation or survival. In an EGFRs-ligands system, however, signal transduction occurs not only unidirectionally but also bidirectionally, which is regulated by cell-cell contact and proteolytic cleavage. Recent studies of proteolytic cleavage 'ectodomain shedding' of EGFRs and their ligands mediated by membrane-type metalloproteinases, a disintegrin and metalloproteinases have been unveiling novel functions and molecular mechanism of their remnant peptides. In addition, the study of the remnant peptide signalling would be essential for understanding the physiological and pathological relevance of anti-shedding therapeutic strategies for diseases such as cancer.     

Reduced hyperthermia-induced cutaneous vasodilation and enhanced exercise-induced plasma water loss at simulated high altitude (3,200 m) in humans

We examined whether less convective heat loss during exercise at high altitude than at sea level was partially caused by reduced cutaneous vasodilation due to enhanced plasma water loss into contracting muscles and whether it was caused by hypoxia rather than by hypobaria. Seven young men performed cycling exercise for 40 min at 50% peak aerobic power in normoxia at (710 mmHg) 610 m, determined before the experiments, in three trials: 1) normobaric normoxia at 610 m (CNT), 2) hypobaric hypoxia [low pressure and low oxygen (LPLO)] at 3,200 m (510 mmHg), 3) normobaric hypoxia [normal pressure and low oxygen (NPLO)] at 610 m, in an artificial climate chamber where atmospheric temperature and relative humidity were maintained at 30°C and 50%, respectively. Subjects in CNT and LPLO breathed room air, whereas those in NPLO breathed a mixed gas of 14% O? balanced N?, equivalent to the gas composition in LPLO. We measured change in PV (ΔPV), oxygen consumption rate (Vo?), mean arterial blood pressure (MBP), esophageal temperature (T(es)), mean skin temperature (T(sk)), forearm skin blood flow (FBF), and sweat rate (SR) during exercise. Although Vo?, MBP, T(sk), and SR responses during exercise were similar between trials (P > 0.05), the sensitivity of forearm vascular conductance (FBF/MBP) in response to increased T(es) was lower in LPLO and NPLO than in CNT (P < 0.05), whereas that of SR was not, resulting in a greater increase in T(es) from minute 5 to 40 of exercise in LPLO and NPLO than in CNT (P = 0.026 and P = 0.011, respectively). ΔPV during exercise was twofold greater in LPLO and NPLO than in CNT. These variables were not significantly different between LPLO and NPLO. Thus reduced convective heat loss during exercise at 3,200 m was partially caused by reduced cutaneous vasodilation due to enhanced PV loss. Moreover, this may be caused by hypoxia rather than by hypobaria.     

Remarkable acceleration of a DNA/RNA inter-strand functionality transfer reaction to modify a cytosine residue: the proximity effect via complexation with a metal cation

Modified nucleosides in natural RNA molecules are essential for their functions. Non-natural nucleoside analogues have been introduced into RNA to manipulate its structure and function. We have recently developed a new strategy for the in situ modification of RNA based on the functionality transfer reaction between an oligodeoxynucleotide probe and an RNA substrate. 2′-Deoxy-6-thioguanosine (6-thio-dG) was used as the platform to anchor the transfer group. In this study, a pyridinyl vinyl ketone moiety was newly designed as the transfer group with the expectation that a metal cation would form a chelate complex with the pyridinyl-2-keto group. It was demonstrated that the (E)-pyridinyl vinyl keto group was efficiently and specifically transferred to the 4-amino group of the opposing cytosine in RNA in the presence of NiCl₂ with more than 200-fold accelerated rate compared with the previous system with the use of the diketo transfer group. Detailed mechanistic studies suggested that NiCl₂ forms a bridging complex between the pyridinyl keto moiety and the N7 of the purine residue neighboring the cytosine residue of the RNA substrate to bring the groups in close proximity.     

Bee pollination of the endangered orchid Calanthe discolor through a generalized food-deceptive system

Calanthe discolor is a Japanese terrestrial orchid that is cultivated for its beautiful flowers arranged in racemose inflorescences. Although its propagation for horticultural purposes has been studied extensively resulting in the successful production of seedlings, little is known about the pollinators and breeding system of C. discolor in its natural habitat. The current study, which combined field observations and pollination experiments, was conducted to gain further insight into the reproduction of this important orchid species. Three bee species: Eucera nipponensis, Osmia cornifrons and Apis cerana japonica, were found to be effective pollinators, transferring the pollinaria on their heads. Pollination experiments also revealed that this species was self-compatible, although it was neither autogamous nor apogamous. The fruit set for the open-pollinated flowers was less than 10 %, suggesting a high degree of pollinator limitation, possibly as a result of the deceptive nature of this species. These results provide evidence that pollinator specificity is the primary mechanism of reproductive isolation between C. discolor and its close relative Calanthe striata, because the latter species is known to be exclusively pollinated by carpenter bee.     

Catalytic implications of the higher plant ADP-glucose pyrophosphorylase large subunit

ADP-glucose pyrophosphorylase, a key regulatory enzyme of starch biosynthesis, is composed of a pair of catalytic small subunits (SSs) and a pair of catalytically disabled large subunits (LSs). The N-terminal region of the LS has been known to be essential for the allosteric regulatory properties of the heterotetrameric enzyme. To gain further insight on the role of this region and the LS itself in enzyme function, the six proline residues found in the N-terminal region of the potato tuber AGPase were subjected to scanning mutagenesis. The wildtype and various mutant heterotetramers were expressed using our newly developed host-vector system, purified, and their kinetic parameters assessed. While P(17)L, P(26)L, and P(55)L mutations only moderately affected the kinetic properties, P(52)L and P(66)L gave rise to significant and contrasting changes in allosteric properties: P(66)L enzyme displayed up-regulatory properties toward 3-PGA while the P(52)L enzyme had down-regulatory properties. Unlike the other mutants, however, various mutations at P(44) led to only moderate changes in regulatory properties, but had severely impaired catalytic rates, apparent substrate affinities, and responsiveness to metabolic effectors, indicating Pro-44 or the LS is essential for optimal catalysis and activation of the AGPase heterotetramer. The catalytic importance of the LS is further supported by photoaffinity labeling studies, which revealed that the LS binds ATP at the same efficiency as the SS. These results indicate that the LS, although considered having no catalytic activity, may mimic many of the catalytic events undertaken by the SS and, thereby, influences net catalysis of the heterotetrameric enzyme.     

Redox potential of plastoquinone A in spinach chloroplasts

The redox potential of plastoquinone A in spinach chloroplasts was determined. The midpoint potential of the quinone is about +80 mV at pH 7.0 with an n value of 2. The pH-dependence of the potential is ?30 mV per pH between pH 4.0 and 5.7, and ?60 mV per pH between pH 5.7 and 8.0. The change of the slope at pH 5.7 is interpreted as the protonation of the oxidized plastoquinone A.     

Genetic evidence for key roles of decorin and biglycan in dentin mineralization

Targeted disruption of the dentin sialophosphoprotein (DSPP) gene in the mice (Dspp^?/?) results in dentin mineralization defects with enlarged predentin phenotype similar to human dentinogenesis imperfecta type III. Using DSPP/biglycan (Dspp^?/?Bgn^?/0) and DSPP/decorin (Dspp^?/?Dcn^?/?) double knockout mice, here we determined that the enlarged predentin layer in Dspp^?/? teeth is rescued in the absence of decorin, but not in the absence of biglycan. However, Fourier transform infrared (FTIR) spectroscopy analysis reveals similar hypomineralization of dentin in both Dspp^?/?Bgn^?/0 and Dspp^?/?Dcn^?/? teeth. Atomic force microscopy (AFM) analysis of collagen fibrils in dentin shows subtle differences in the collagen fibril morphology in these genotypes. The reduction of enlarged predentin in Dspp^?/?Dcn^?/? mice suggests that the elevated level of decorin in Dspp^?/? predentin interferes with the mineralization process at the dentin mineralization front. On the other hand, the lack of DSPP and biglycan leads to the increased number of calcospherites in Dspp^?/?Bgn^?/0 predentin, suggesting that a failure in coalescence of calcospherites was augmented in Dspp^?/?Bgn^?/0 teeth as compared to Dspp^?/? teeth. These findings indicate that normal expression of small leucine rich proteoglycans, such as biglycan and decorin, plays an important role in the highly orchestrated process of dentin mineralization.