Identification and biochemical characterization of novel putative substrates for the epidermal growth factor receptor kinase.

To gain insight into the mechanisms which control the mitogenic response to epidermal growth factor (EGF), we have partially purified and characterized several intracellular proteins which are phosphorylated on tyrosine residues following activation of the epidermal growth factor receptor (EGFR). Partial purification was achieved by immunoaffinity chromatography using immobilized anti-phosphotyrosine antibodies. Antisera generated against the partially purified proteins were used to identify at least five novel EGFR putative substrates, designated, on the basis of their apparent molecular weight, p97, p68, p61, p56, and p23. All of these proteins became specifically phosphorylated on tyrosine after EGF treatment of intact cells, as assessed by phosphoamino acid analysis, and none of them represented an EGFR degradation product. The phosphorylation of these proteins appeared to be relatively specific for the EGFR. In particular, an EGFR-related kinase, erbB-2 was much less efficient than EGFR at phosphorylating p97, p56, and p23 and incapable of phosphorylating p68. The identification of these novel EGFR putative substrates should lead to a better understanding of the mechanisms controlling the specificity of EGFR-mediated mitogenic signaling.     

A single amino acid substitution is sufficient to modify the mitogenic properties of the epidermal growth factor receptor to resemble that of gp185erbB-2.

The epidermal growth factor (EGF) receptor (EGFR) and the erbB-2 gene product, gp185erbB-2, exhibit distinct abilities to stimulate mitogenesis in different target cells. By using chimeric molecules between these two receptors, we have previously shown that their intracellular juxtamembrane regions are responsible for this specificity. Here we describe a genetically engineered EGFR mutant containing a threonine for arginine substitution at position 662 in the EGFR juxtamembrane domain, corresponding to threonine 694 in gp185erbB-2. This mutant, designated EGFRThr662, displayed affinity for EGF binding and catalytic properties that were indistinguishable from those of the wild type EGFR. However, EGFRThr662 behaved much as gp185erbB-2 in a number of bioassays which readily distinguish between the mitogenic effects of EGFR and gp185erbB-2. Moreover, significant differences were detected in the pattern of intracellular proteins phosphorylated on tyrosine in vivo by EGFR and EGFRThr662 in response to EGF. Thus, small differences in the primary sequence of two closely related receptors have dramatic effects on their ability to couple with mitogenic pathways.     

Rhinocerotidae (Mammalia,Perissodactyla) from the middle Pleistocene levels of Grotta Romanelli (Lecce,southern Italy)

The rhinoceros remains collected during the past century in the lower levels XII (= K) and XI (= I) of the famous Pleistocene locality of Grotta Romanelli (Lecce, southern Italy) are described and compared in detail for the first time. Some remains are referred to Stephanorhinus sp. and others are assigned here to the late early-middle Pleistocene European species Stephanorhinus hundsheimensis based on several morphological characters. Based on its olivine-bearing texture, the volcanoclastic ash sampled from some rhinoceros bones can be referred to the first phase of the Monte Vulture activity (around 630 ka). The results of the stable isotope analyses suggest that the climate in the lowest levels of Grotta Romanelli could have been more arid than it was at the time of the upper level IX, which is generally referred to the late Pleistocene. In addition, both recent day δ¹⁸O_ppt values and MAT are very similar to values calculated for levels X and XII, suggesting that the climate at those times may have been close to the Present one, whereas climate in level IX may have been somewhat cooler. The presence of Stephanorhinus hundsheimensis suggests a middle Pleistocene age for the lower levels of Grotta Romanelli, in agreement with the results obtained from the volcanoclastic material.     

Variation in wood density determines spatial patterns inAmazonian forest biomass

Uncertainty in biomass estimates is one of the greatest limitations to models of carbon flux in tropical forests. Previous comparisons of field‐based estimates of the aboveground biomass (AGB) of trees greater than 10 cm diameter within Amazonia have been limited by the paucity of data for western Amazon forests, and the use of site‐specific methods to estimate biomass from inventory data. In addition, the role of regional variation in stand‐level wood specific gravity has not previously been considered. Using data from 56 mature forest plots across Amazonia, we consider the relative roles of species composition (wood specific gravity) and forest structure (basal area) in determining variation in AGB. Mean stand‐level wood specific gravity, on a per stem basis, is 15.8% higher in forests in central and eastern, compared with northwestern Amazonia. This pattern is due to the higher diversity and abundance of taxa with high specific gravity values in central and eastern Amazonia, and the greater diversity and abundance of taxa with low specific gravity values in western Amazonia. For two estimates of AGB derived using different allometric equations, basal area explains 51.7% and 63.4%, and stand‐level specific gravity 45.4% and 29.7%, of the total variation in AGB. The variation in specific gravity is important because it determines the regional scale, spatial pattern of AGB. When weighting by specific gravity is included, central and eastern Amazon forests have significantly higher AGB than stands in northwest or southwest Amazonia. The regional‐scale pattern of species composition therefore defines a broad gradient of AGB across Amazonia.     

Phylogeny of the Highly Divergent Echinosteliales (Amoebozoa)

Myxomycetes or plasmodial slime molds are widespread and very common soil amoebae with the ability to form macroscopic fruiting bodies. Even if their phylogenetic position as a monophyletic group in Amoebozoa is well established, their internal relationships are still not entirely resolved. At the base of the most intensively studied dark‐spored clade lies the order Echinosteliales, whose highly divergent small subunit ribosomal (18S) RNA genes represent a challenge for phylogenetic reconstructions. This is because they are characterized by unusually long variable helices of unknown secondary structure and a high inter‐ and infraspecific divergence. Current classification recognizes two families: the monogeneric Echinosteliaceae and the Clastodermataceae with the genera Barbeyella and Clastoderma. To better resolve the phylogeny of the Echinosteliales, we obtained three new small subunit ribosomal (18S) RNA gene sequences of Clastoderma and Echinostelium corynophorum. Our phylogenetic analyses suggested the polyphyly of the family Clastodermataceae, as Barbeyella was more closely related to Echinostelium arboreum than to Clastoderma, while Clastoderma debaryanum was the earliest branching clade in Echinosteliales. We also found that E. corynophorum was the closest relative of the enigmatic Semimorula liquescens, a stalkless‐modified Echinosteliales. We discuss possible evolutionary pathways in dark‐spored Myxomycetes and propose a taxonomic update.