Climate change and its impact on the forests of Kilimanjaro

Cloud forests are of great importance in the hydrological functioning of watersheds in subhumid East Africa. However, the montane forests of Mt. Kilimanjaro are heavily threatened by global change impacts. Based on an evaluation of over 1500 vegetation plots and interpretation of satellite imagery from 1976 and 2000, land-cover changes on Kilimanjaro were evaluated and their impact on the water balance estimated. While the vanishing glaciers of Kilimanjaro attract broad interest, the associated increase of frequency and intensity of fires on the slopes of Kilimanjaro is less conspicuous but ecologically far more significant. These climate change-induced fires have lead to changes in species composition and structure of the forests and to a downward shift of the upper forest line by several hundred metres. During the last 70 years, Kilimanjaro has lost nearly one-third of its forest cover, in the upper areas caused by fire, on the lower forest border mainly caused by clearing. The loss of 150 km² of cloud forest – the most effective source in the upper montane and subalpine fog interception zone – caused by fire during the last three decades means a considerable reduction in water yield. In contrast to common belief, global warming does not necessarily cause upward migration of plants and animals. On Kilimanjaro the opposite trend is under way, with consequences more harmful than those due to the loss of the showy ice cap of Africa's highest mountain.     

Stimulation of r- vs. K-selected microorganisms by elevated atmospheric CO2 depends on soil aggregate size

Increased root exudation under elevated atmospheric CO₂ and the contrasting environments in soil macro- and microaggregates could affect microbial growth strategies. We investigated the effect of elevated CO₂ on the contribution of fast- ( r -strategists) and slow-growing ( K -strategists) microorganisms in soil macro- and microaggregates. We fractionated the bulk soil from the ambient and elevated (for 5 years) CO₂ treatments of FACE-Hohenheim (Stuttgart) into large macro- (>2 mm), small macro- (0.25–2.00 mm), and microaggregates (<0.25 mm) using 'optimal moist' sieving. Microbial biomass (C_mic), the maximum specific growth rate (μ), growing microbial biomass (GMB) and lag-period ( t _lag) were estimated by the kinetics of CO₂ emission from bulk soil and aggregates amended with glucose and nutrients. Although C_org and C_mic were unaffected by elevated CO₂, μ values were significantly higher under elevated than ambient CO₂ for bulk soil, small macroaggregates, and microaggregates. Substrate-induced respiratory response increased with decreasing aggregate size under both CO₂ treatments. Based on changes in μ, GMB and lag period, we conclude that elevated atmospheric CO₂ stimulated the r- selected microorganisms, especially in soil microaggregates. Such an increase in r -selected microorganisms indicates acceleration of available C mineralization in soil, which may counterbalance the additional C input by roots in soils in a future elevated atmospheric CO₂ environment.     

Vegetation of Kilimanjaro: hidden endemics and missing bamboo

Kilimanjaro has a large variety of forest types over an altitudinal range of 3000 m containing over 1200 vascular plant species. Montane Ocotea forests occur on the wet southern slope. Cassipourea and Juniperus forests grow on the dry northern slope. Subalpine Erica forests at 4100 m represent the highest elevation cloud forests in Africa. In contrast to this enormous biodiversity, the degree of endemism is low. However, forest relicts in the deepest valleys of the cultivated lower areas suggest that a rich forest flora inhabited Mt Kilimanjaro in the past, with restricted‐range species otherwise only known from the Eastern Arc mountains. The low degree of endemism on Kilimanjaro may result from destruction of lower altitude forest rather than the relatively young age of the mountain. Another feature of the forests of Kilimanjaro is the absence of a bamboo zone, which occurs on all other tall mountains in East Africa with a similarly high rainfall. Sinarundinaria alpina stands are favoured by elephants and buffaloes. On Kilimanjaro these megaherbivores occur on the northern slopes, where it is too dry for a large bamboo zone to develop. They are excluded from the wet southern slope forests by topography and humans, who have cultivated the foothills for at least 2000 years. This interplay of biotic and abiotic factors could explain not only the lack of a bamboo zone on Kilimanjaro but also offers possible explanations for the patterns of diversity and endemism. Kilimanjaro's forests can therefore serve as a striking example of the large and long‐lasting influence of both animals and humans on the African landscape.     

Escherichia coli relA strains as hosts for amplification of pBR322 plasmid DNA

Abstract We have proposed that guanosine tetraphosphate produced in Escherichia coli cells subjected to an isoleucine limitation inhibits pBR322 DNA replication [1]. In E. coli relA which cannot synthesize guanosine tetraphosphate (ppGpp) upon amino acid limitation pBR322 DNA is amplified after arginine starvation. The yield of plasmid DNA amplified either by chloramphenicol (Cm) or by arginine limitation is compared. The plasmid yield per cell is equal in amino acid-starved cells and in cells treated with Cm. To increase the plasmid content per ml of cell suspension the growth medium was supplemented with increasing amounts of nutrients. Plasmid DNA can be isolated in large quantities by this procedure. This simple method can be used for the enrichment of pBR325 DNA which cannot be amplified by Cm treatment. Our results indicate that E. coli relA strains might be suitable hosts for the amplification of pBR322 and related plasmids in E. coli .     

Biochemical Characterization of Different Molecular Forms of the Neural Cell Adhesion Molecule L1

The neural cell adhesion molecule L1 is a phosphorylated, integral membrane glycoprotein that is recovered from adult mouse brain tissue by immunoaffinity chromatography as a set of polypeptides with apparent molecular masses of 200, 180, 140, and 80 kilodaltons (L1–200, L1–180, L1–140, and L1–80, respectively). It has been shown that L1–140 and the phosphorylated L1–80 is generated from L1–200 by mild proteolytic treatment of intact cells. In the present study we have investigated the structural relationships between the different molecular forms of L1 and their location with regard to the surface membrane. We could show that L1–200 has two preferred cleavage sites, one that generates the amino terminal, extracellularly exposed L1–140 and the carboxy terminal L1–80 that spans the membrane. Cleavage at the other site leads to the generation of the amino terminally located L1–180 and the membrane-attached, phosphorylated carboxy terminal L1–30. This site is cleaved during treatment of live cultured cells with broad-spectrum, protease-free phospholipase C (but not phosphatidylinositol-specific phospholipase C) or exposure to sodium azide or cyanogen bromide. Other conditions that cause damage to cells do not lead to the generation of L1–180 and L1–30, suggesting a particular cell-intrinsic cleavage mechanism. L1–180 is truly soluble in aqueous solutions, since it can be recovered from culture supernatants and in the supernatant of a crude membrane fraction after incubation for 2 h at 37°C. Although trypsin treatment alone does not release L1–140 into the supernatant, combination of phospholipase C and mild tryptic treatment leads to the release of L1–140 and L1–50, the latter being most likely the extracellularly exposed domain of L1–80 that is complementary to the membrane-integrated phosphorylated L1–30. Phase separation experiments with Triton X-114 show that the released forms of L1–180 and L1–140 distribute into the aqueous phase, whereas they distribute into the detergent phase when in association with L1–200 or L1–80. However, when L1–80 is cleaved to yield the soluble L1–50 and membrane-anchored L1–30, L1–140 is released into the supernatant together with L1–50. A strong affinity of L1–200, L1–140, and L1–80 to each other is also indicated by the fact that they incorporate together into liposomes and separate only under strong detergent conditions. Also, a strong tendency to aggregate is observed for L1-containing liposomes, but not for those containing the adhesion molecules neural cell adhesion molecule and myelin-associated glycoprotein. Although the physiological roles of the soluble L1 forms, their mode of generation, and the strong affinity for each other remain to be investigated, the availability of soluble forms of L1 opens the possibility to use them as probes for the functional properties of L1 in assay systems involving live cells in vitro.     

The murine homeo box gene product, Hox 1.1 protein, is growth-controlled and associated with chromatin

In order to gain insight into the function of the Hox 1.1 gene, we studied the expression of the murine homeo box gene product, the Hox 1.1 protein. Monoclonal antibodies were raised against synthetic peptides of the Hox 1.1 protein to study the localization and expression pattern of this protein under various culture conditions. By means of indirect immunofluorescence we localized the Hox 1.1 protein to the nucleus in differentiated F9 and NIH 3T3 cells. During mitosis the protein was found to be associated with chromatin. Confluent NIH 3T3 cells harbored little if any Hox 1.1 protein. After "wounding" the cells in this confluent monolayer, we observed an induction of the expression of the Hox 1.1 protein. However, addition of insulin to F9 and contact-inhibited NIH 3T3 cells led to an increase of the Hox 1.1 RNA and protein expression. Thus, the induction of the Hox 1.1 protein is associated not only with the differentiation of embryonal carcinoma (EC) cells, but may also correlate with stages of cell growth.     

Purification and Characterization of a Soluble and a Particulate Glutamate Dehydrogenase from Rat Brain

Glutamate dehydrogenase (GDH) activity was determined in high-speed fractions (100,000 g for 60 min) obtained from whole rat brain homogenates after removal of a low-speed pellet (480 g for 10 min). Approximately 60% of the high-speed GDH activity was particulate (associated with membrane) and the remaining was soluble (probably of mitochondrial matrix origin). Most of the particulate GDH activity resisted extraction by several commonly used detergents, high concentration of salt, and sonication; however, it was largely extractable with the cationic detergent cetyltrimethylammonium bromide (CTAB) in hypotonic buffer solution. The two GDH activities were purified using a combination of hydrophobic interaction, ion exchange, and hydroxyapatite chromatography. Throughout these purification steps the two activities showed similar behavior. Kinetic studies indicated similar Km values for the two GDH fractions for the substrates alpha-ketoglutarate, ammonia, and glutamate; however, there were small but significant differences in Km values for NADH and NADPH. Although the allosteric stimulation by ADP and L-leucine and inhibition by diethylstilbestrol was comparable, the two GDH components differed significantly in their susceptibility to GTP inhibition in the presence of 1 mM ADP, with apparent Ki values of 18.5 and 9.0 microM GTP for the soluble and particulate fractions, respectively. The Hill plot coefficient, binding constant, and cooperativity index for the GTP inhibition were also significantly different, indicating that the two GDH activities differ in their allosteric sites. In addition, enzyme activities of the two purified proteins exhibited a significant difference in thermal stability when inactivated at 45 degrees C and pH 7.4 in 50 mM phosphate buffer.     

Depolarizing Agents Regulate the Phosphorylation of Myelin Basic Protein in Rat Optic Nerves

The regulation of the state of phosphorylation of myelin basic protein has been studied in intact rat optic nerves incubated in vitro. For this purpose the endogenous state of phosphorylation was preserved and the "back-phosphorylation" technique was used to determine the amount of dephosphorylated protein present in extracts of the nerves. Our results indicate that when nerves were incubated in the presence of depolarizing agents, the state of phosphorylation of myelin basic protein was increased. This effect was calcium-dependent and was partly inhibited by chlorpromazine.     

The CCK-B Antagonist CI-988 Increases Dopamine Levels in Microdialysate from the Rat Nucleus Accumbens via a Tetrodotoxin- and Calcium-Independent Mechanism

Abstract: CI-988, a water-soluble, selective cholecystokinin-B antagonist, was perfused through a microdialysis probe into the anterior nucleus accumbens, posterior nucleus accumbens, or caudate nucleus of anesthetized rats. High concentrations of CI-988 produced three- to fivefold increases in dopamine overflow, at all three sites, that were temporally correlated with the CI-988 perfusion and returned to baseline levels upon cessation of CI-988 perfusion. However, the cholecystokinin-A antagonist CAM-1481, and the relatively inactive enantiomer of CI-988, CAM-1241, also increased dopamine overflow in the nucleus accumbens. Furthermore, the ability of CI-988 to increase dopamine overflow persisted in the absence of calcium in the perfusate and was not sensitive to tetrodotoxin treatment. The mechanism by which locally administered CI-988 increases dopamine overflow appears not to be anatomically specific, not selective for one cholecystokinin receptor subtype, and may be nonvesicular.