Cloning and restriction fragment length polymorphism analysis of a cDNA for swine PIT-1, a gene controlling growth hormone expression*

A partial swine cDNA which encodes the functional domain of PIT-1 was isolated by the polymerse chain reaction (PCR). The swine PIT-1 cDNA clone is 95% identical at the protein level to the rat Pit-1 gene. Thus, Pit-l's known function in control of rat growth hormone and prolactin expression is likely to be conserved in swine. This swine cDNA clone was used to investigate genetic variability at PIT-1 in several American and Chinese breeds. Polymorphic BamIII fragments were found in pure-bred Meishan animals (n= 13), but only monomorphic fragments in five American breeds (n= 36).     

Whole-plant ABA flux and the regulation of water loss in Cedrella odorata

Three-month-old Cedrella odorata seedlings were exposed to a soil-drying treatment. During this period, xylem sap was periodically collected from the plant by applying pneumatic pressure to the roots. This also allowed whole-plant water status to be measured by recording the balancing pressure applied. The concentration of ABA in xylem sap (C) was related to the whole-plant transpiration rate (V) which was measured with a sap flow gauge. The analysis of these paired measurements centred on how the reciprocal of C (R) varied with respect to V. This revealed that (1) the observed increases in C could not be explained by the reductions in V alone, (2) initially, decreases in V were associated with proportional increases in the whole-plant ABA flux (M), and (3) this relationship broke down at low values of V since zero flow was associated with a finite value for C estimated to be 41 pmol ABA mmol^?1 H₂O. A simple static model is developed from the observations that is able to explain the data well, and the results are discussed in terms of the effects of ABA on stomatal conductance (g_sw).     

Endothelin-1 stimulates catalase activity through the PKCδ-mediated phosphorylation of serine 167

Our previous studies have shown that endothelin-1 (ET-1) stimulates catalase activity in endothelial cells and in lambs with acute increases in pulmonary blood flow (PBF), without altering gene expression. The purpose of this study was to investigate the molecular mechanism by which this occurs. Exposing pulmonary arterial endothelial cells to ET-1 increased catalase activity and decreased cellular hydrogen peroxide (H₂O₂) levels. These changes correlated with an increase in serine-phosphorylated catalase. Using the inhibitory peptide δV1.1, this phosphorylation was shown to be protein kinase Cδ (PKCδ) dependent. Mass spectrometry identified serine 167 as the phosphorylation site. Site-directed mutagenesis was used to generate a phospho-mimic (S167D) catalase. Activity assays using recombinant protein purified from Escherichia coli or transiently transfected COS-7 cells demonstrated that S167D catalase had an increased ability to degrade H₂O₂ compared to the wild-type enzyme. Using a phospho-specific antibody, we were able to verify that pS167 catalase levels are modulated in lambs with acute increases in PBF in the presence and absence of the ET receptor antagonist tezosentan. S167 is located on the dimeric interface, suggesting it could be involved in regulating the formation of catalase tetramers. To evaluate this possibility we utilized analytical gel filtration to examine the multimeric structure of recombinant wild-type and S167D catalase. We found that recombinant wild-type catalase was present as a mixture of monomers and dimers, whereas S167D catalase was primarily tetrameric. Further, the incubation of wild-type catalase with PKCδ was sufficient to convert wild-type catalase into a tetrameric structure. In conclusion, this is the first report indicating that the phosphorylation of catalase regulates its multimeric structure and activity.     

Effects of starvation and sucrose feeding on corpora allata of last instar larvae of Manduca sexta: in vitro activity, size, and cell number

Abstract Larvae of the tobacco hornworm moth Manduca sexta starved for the first 3 days of the last (fifth) stadium undergo a supernumerary moult. If they are provided with sucrose during the starvation period, they develop into normal pupae although pupation is delayed. The activities of the corpora allata (CA) from normal, starved, and sucrose fed larvae were followed through the fifth stadium with a radiochemical assay for Juvenile Hormone (JH) biosynthesis. An attempt was made to correlate CA-activity with CA cell number, size, and protein content.
In CA of normally fed larvae the rate of JH synthesis declined to undetectable levels by day 4 which was also the time of exposure of the dorsal vessel. In CA of starved larvae, the rate of JH synthesis at first decreased but began to increase on day 3 and reached a peak value by day 7 , at which time head capsule slippage occurred. In CA of sucrose fed larvae, the rate of biosynthesis declined as in normal larvae but the decline was extended over a longer period. Exposure of the dorsal vessel was delayed in the same manner and occurred on days 7–9. The major JH in all cases was JH-II.
The CA comprise c. 150 cells in the early fifth stadium, and this number remained constant during the fifth stadium in all three feeding regimens. In normal larvae, CA size and protein content increased several-fold during the stadium whereas in starved and sucrose-fed larvae they increased slowly and in agreement with the altered timing of developmental events. In none of the groups was the CA activity pattern correlated with morphometric changes of the CA. The rates of JH biosynthesis were not closely correlated with published JH titre curves. The in vivo mechanisms for regulation of JH production remain to be elucidated.     

Hydraulic conductance, stomatal conductance, and maximal photosynthetic rate in bean leaves

A positive correlation was found between steady state values of hydraulic (L _pA) and stomatal conductance (g _s) of French bean leaves: both were lower in the dark than in the light and lower in water-deficient plants than in the well-watered ones. The relative rate of stomatal opening after a pressure rise in the xylem was also positively related to L _pA. The L _pA and g _s were both related to the maximal photosynthetic rate at saturating CO2 concentrations. This revised version was published online in June 2006 with corrections to the Cover Date.     

From the beginning to the present state of molecular microbial pathogenesis—A tribute to Pascale Cossart

The universe of Molecular Microbial Pathogenesis is filled with many female and male stars. But there are two particularly bright shining supernovae-like stars: the late Stanley Falkow and the very lively and creative Pascale Cossart. These two outstanding luminaries, surrounded by numerous planets, do not only belong to different scientific generations but their splendor also comes from very different scientific concepts. Stanley Falkow, often referred to as the ‘Father of Molecular Microbial Pathogenesis’, made many groundbreaking contributions to this field by addressing almost all important bacterial pathogens. Pascale Cossart, who could be called in analogy the ‘Queen of Modern Molecular Microbial Pathogenesis’ by combining the Microbiology and Cell Biology, concentrates in her similarly impressive scientific work essentially on a single bacterial species which she studied and still studies in great depth: the facultative intracellular bacterial pathogen Listeria monocytogenes—and the vast majority of her most prominent publications deals with this pathogen in almost all facets. It is certainly not an exaggeration to say that she together with her co-workers and collaborators developed this model bacterium into a paradigm among the intracellular bacterial pathogens.     

Field Hypothesis on the Self-regulation of Gene Expression

The mechanism of the self-regulation of gene expression in living cells is generally explained by considering complicated networks of key-lock relationships, and in fact there is a large body of evidence on a hugenumber of key-lock relationships. However, in the present article we stress that with the network hypothesis alone it is impossible to fully explain the mechanism of self-regulation in life. Recently, it has been established that individual giant DNA molecules, larger than several tens of kilo base pairs, undergo a large discrete transition in their higher-order structure. It has become clear that nonspecific weak interactions with various chemicals, suchas polyamines, small salts, ATP and RNA, cause on/off switching in the higher-order structure of DNA. Thus, the field parameters of the cellular environment should play important roles in the mechanism of self-regulation, in addition to networks of key and locks. This conformational transition induced by field parameters may be related to rigid on/off regulation, whereas key-lock relationships may be involved in a more flexible control of gene expression.