Plant intraspecific functional trait variation is related to within‐habitat heterogeneity and genetic diversity in Trifolium montanum L.

Intraspecific trait variation (ITV), based on available genetic diversity, is one of the major means plant populations can respond to environmental variability. The study of functional trait variation and diversity has become popular in ecological research, for example, as a proxy for plant performance influencing fitness. Up to now, it is unclear which aspects of intraspecific functional trait variation (iFD_CV) can be attributed to the environment or genetics under natural conditions. Here, we examined 260 individuals from 13 locations of the rare (semi‐)dry calcareous grassland species Trifolium montanum L. in terms of iFD_CV, within‐habitat heterogeneity, and genetic diversity. The iFD_CV was assessed by measuring functional traits (releasing height, biomass, leaf area, specific leaf area, leaf dry matter content, F_v/F_m, performance index, stomatal pore surface, and stomatal pore area index). Abiotic within‐habitat heterogeneity was derived from altitude, slope exposure, slope, leaf area index, soil depth, and further soil factors. Based on microsatellites, we calculated expected heterozygosity (H_e) because it best‐explained, among other indices, iFD_CV. We performed multiple linear regression models quantifying relationships among iFD_CV, abiotic within‐habitat heterogeneity and genetic diversity, and also between separate functional traits and abiotic within‐habitat heterogeneity or genetic diversity. We found that abiotic within‐habitat heterogeneity influenced iFD_CV twice as strong compared to genetic diversity. Both aspects together explained 77% of variation in iFD_CV ( = .77, F_{2, 10} = 21.66, p < .001). The majority of functional traits (releasing height, biomass, specific leaf area, leaf dry matter content, F_v/F_m, and performance index) were related to abiotic habitat conditions indicating responses to environmental heterogeneity. In contrast, only morphology‐related functional traits (releasing height, biomass, and leaf area) were related to genetics. Our results suggest that both within‐habitat heterogeneity and genetic diversity affect iFD_CV and are thus crucial to consider when aiming to understand or predict changes of plant species performance under changing environmental conditions.     

Assessment of histogenesis and proliferative potential in cytologic specimens of human brain tumors. Value of immunocytochemistry and nucleolar organizer regions.

The value of immunocytochemistry and nucleolar organizer regions (NORs) for the histogenetic identification and the estimation of the proliferative potential of brain tumors was assessed by the investigation of imprint smears of 51 neurosurgical tumor specimens. A panel of five monoclonal antibodies was used to cover a broad range of immunohistochemical markers. For the assessment of NORs, a silver staining technique (AgNOR) was used. NORs were enumerated and measured by means of an interactive image analysis system. The immunocytochemical results were similar for the smears and paraffin-embedded sections for 95.6% of the investigations performed and for 76.2% of the cases. Glial fibrillary acidic protein (GFAP) was positive in 9 of 17 tumors of glial origin, but was negative in 9 metastatic tumors. Vimentin was positive in 10 of 10 and fibronectin in 9 of 10 meningiomas investigated. The number of NORs increased steadily with the increasing grade of malignancy. Especially in glioblastomas, the number of NORs per cell exhibited a wide range, which might reflect the heterogeneity of these neoplasms. Metastases revealed a higher number of NORs per cell than did glioblastomas. In the cytologic differential diagnosis of these tumors, an absence of GFAP expression combined with a high NOR count is suggestive of a metastatic tumor.     

Evolution of a B2 tagged sequence from a long-range repeat family in the genus Mus

A long-range repeat family of more than 50 kb repeat size is clustered in Chromosomes (Chr) 1 of Mus musculus and M. spretus. In M. musculus this long-range repeat family shows considerable variation of copy-number frequency and contains coding regions for at least two genes. In an intron of a gene, which is part of the repeat, a B2 small interspersed repetitive element (SINE) is inserted at identical positions. The B2 element is present in all copies of the long-range repeat family; it was presumably a component of the ancestral single-copy precursor sequence that gave rise by amplification to the repeat family. Copies of the long-range repeat family vary with respect to the number of TAAA tandem repeats in the A-rich 3 end region of the B2 element. As inferred from polymerase chain reaction (PCR) data, presence and frequency of repeat number variants in the (TAAA)_n block are strain and species specific. The B2 element and its flanking regions were sequenced from two copies of the long-range repeat family. Sequence divergence between the two copies (only non-CG base substitutions and deletions/insertions) was determined to be 2.6%. Based on the drift rate in human Alu elements and a correction for the higher drift rates in rodents, and estimate for the divergence time of 1.7 million years was calculated. Since the long-range repeat family is present in M. musculus and M. spretus, it must have evolved by amplification before the separation of the two species about 1–4 million years ago.     

Identification of two types of keratin polypeptides within the acidic cytokeratin subfamily I

Cytoskeletal filaments of the α-keratin type (cytokeratins) are a characteristic of epithelial cells. In diverse mammals (man, cow and rodents) these cytokeratins consist of a family of approximately 20 polypeptides, which may be divided into the more acidic (I) and the more basic (II) subfamilies. These two subfamilies show only limited amino acid sequence homology. In contrast, nucleic acid hybridization experiments and peptide maps have been interpreted to show that polypeptides of the same subfamily share extended sequence homology.We compare two polypeptides of the acidic cytokeratin subfamily, VIb (M_r 54,000) and VII (M_r 50,000), which are co-expressed in large amounts in bovine epidermal keratinocytes. These two epidermal keratins can be distinguished by specific antibodies and show different patterns of expression among several bovine tissues and cultured cells. In addition, they differ in the stability of their complexes with basic keratin polypeptides and in their tryptic peptide maps. The amino acid sequences deduced from the nucleotide sequences of complementary DNA clones containing the 3′ ends of the messenger RNAs for these keratins are compared with each other and with available amino acid sequences of human, murine and amphibian epidermal keratins. Bovine keratins VIb and VII share considerable sequence homology in the α-helical portion (68% residues identical) but lack significant homology in the extrahelical portion. Bovine keratin VIb shows, in its α-helical region, a pronounced sequence homology (88% identity) to the murine epidermal keratin of M_r 59,000. In addition, the non-helical carboxy-terminal regions of both proteins are glycinerich and contain a canonic sequence GGG^S_GYGG, which may be repeated several times. Moreover, their mRNAs present a highly conserved stretch of 236 nucleotides containing, in the murine sequence, the end of the coding and all of the non-coding region (81% identical nucleotides). Bovine keratin VII is considerably different from the murine M_r 59,000 keratin but is almost identical to the human cytokeratin number 14 of M_r 50,000, both in the α-helical and in the non-α-helical regions of the proteins, and the mRNAs of the human and the bovine keratins also display a high homology in their 3′ non-coding ends.The results show that in the same species keratins of the same subfamily can differ considerably, whereas equivalent keratin polypeptides of different species are readily identified by characteristic sequence homologies in the α-helical and the non-helical regions as well as in the 3′ non-coding portions of their mRNAs. Among the members of the acidic subfamily I of cytokeratin polypeptides that are co-expressed in bovine epidermis, at least two types can be distinguished by their carboxy-terminal sequences. One type is characterized by its abundance of glycine residues, a consensus GGG^S_GYGG heptapeptide sequence, which may be repeated several times, and an extended stretch of high RNA sequence homology in the 3′ non-coding part. The other type shows a predominance of serine and valine residues, a subterminal GGG^S_GYGG sequence (which has been maintained in Xenopus, cow and man) and also a high level of homology in the 3′ non-coding part of the mRNA. The data indicate that individual keratin type specificity overrides species diversity, both at the protein and the mRNA level. We discuss the evolutionary conservation and the tissue distribution of these two types of acidic keratin polypeptides as well as their possible biological functions.     

Isolation and characterization of a 3-chlorobenzoate degrading pseudomonad

A pseudomonad has been isolated from sewage, which can utilize 3-chlorobenzoic acid as a sole carbon source. In cells grown on benzoate the enzymes of the -ketoadipic acid pathway are present. Considerable enzymic activities for chlorinated substrates were found in benzoate grown cells only for the oxygenation of 3-chlorobenzoate and the dehydrogenation of 3- and 5-chloro-3,5-cyclohexadiene-1,2-diol-1-carboxylic acid. 3-Chlorobenzoate grown cells show additional high activities for the turnover of 3- and 4-chlorocatechols and chloromuconic acids.Abbreviations Used DHB (-)-3,5-cyclohexadiene-1,2-diol-1-carboxylic acid (derived from the trivial name, dihydrodihydroxybenzoate) - 3- and 5-Cl-DHB correspondingly 3- and 5-chloro-3,5-cyclohexadiene-1,2-diol-1-carboxylic acid     

An Integrated Map with Cosmid/PAC Contigs of a 4-Mb Down Syndrome Critical Region

The major phenotypic features of Down syndrome have been correlated with partial trisomies of chromosome 21, allowing us to define the candidate gene region to a 4-Mb segment on the 21q22.2 band. We present here a high-resolution physical map with megabase-sized cosmid/PAC contigs. This ordered clone library has provided unique material for the integration of a variety of mappable objects, including exons, cDNAs, restriction sites, etc. Furthermore, our results have exemplified a strategy for the completion of the chromosome 21 map to sequencing.     

The intrinsic electrophysiological characteristics of fly lobula plate tangential cells: I. Passive membrane properties

The passive membrane properties of the tangential cells in the fly lobula plate (CH, HS, and VS cells, Fig. 1) were determined by combining compartmental modeling and current injection experiments. As a prerequisite, we built a digital base of the cells by 3D-reconstructing individual tangential cells from cobalt-stained material including both CH cells (VCH and DCH cells), all three HS cells (HSN, HSE, and HSS cells) and most members of the VS cell family (Figs. 2, 3). In a first series of experiments, hyperpolarizing and depolarizing currents were injected to determine steady-state I-V curves (Fig. 4). At potentials more negative than resting, a linear relationship holds, whereas at potentials more positive than resting, an outward rectification is observed. Therefore, in all subsequent experiments, when a sinusoidal current of variable frequency was injected, a negative DC current was superimposed to keep the neurons in a hyperpolarized state. The resulting amplitude and phase spectra revealed an average steady-state input resistance of 4 to 5 M and a cut-off frequency between 40 and 80 Hz (Fig. 5). To determine the passive membrane parameters R _m (specific membrane resistance), R _i (specific internal resistivity), and C _m (specific membrane capacitance), the experiments were repeated in computer simulations on compartmental models of the cells (Fig. 6). Good fits between experimental and simulation data were obtained for the following values: R _m = 2.5 kcm², R _i = 60 cm, and C _m = 1.5 F/cm² for CH cells; R _m = 2.0 kcm², R _i = 40 cm, and C _m = 0.9 F/cm² for HS cells; R _m = 2.0 kcm², R _i = 40 cm, and C _m = 0.8 F/cm² for VS cells. An error analysis of the fitting procedure revealed an area of confidence in the R _m -R _i plane within which the R _m -R _i value pairs are still compatible with the experimental data given the statistical fluctuations inherent in the experiments (Figs. 7, 8). We also investigated whether there exist characteristic differences between different members of the same cell class and how much the exact placement of the electrode (within ±100 m along the axon) influences the result of the simulation (Fig. 9). The membrane parameters were further examined by injection of a hyperpolarizing current pulse (Fig. 10). The resulting compartmental models (Fig. 11) based on the passive membrane parameters determined in this way form the basis of forthcoming studies on dendritic integration and signal propagation in the fly tangential cells (Haag et al., 1997; Haag and Borst, 1997).     

Critical Reactions in Fluorobenzoic Acid Degradation by Pseudomonas sp. B13

3-Chlorobenzoate-grown cells of Pseudomonas sp. B13 readily cometabolized monofluorobenzoates. A catabolic pathway for the isomeric fluorobenzoates is proposed on the basis of key metabolites isolated. Only 4-fluorobenzoate was utilized and totally degraded after a short period of adaptation. The isoenzymes for total degradation of chlorocatechols, being found during growth with 3-chlorobenzoate or 4-chlorophenol, were not induced in the presence of fluorobenzoates. Correspondingly, only the ordinary enzymes of the benzoate pathway were detected in 4-fluorobenzoate-grown cells. Ring cleavage of 3-fluorocatechol was recognized as a critical step in 3-fluorobenzoate degradation. 2-Fluoro-cis,cis-muconic acid was identified as a dead-end metabolite from 2- and 3-fluorobenzoate catabolism. During 2-fluorobenzoate cometabolism, fluoride is eliminated by the initial dioxygenation.     

Degradation of chlorophenols by a defined mixed microbial community

Synthetic sewage containing phenol, acetone, and alkanols plus 4-chlorophenol or a mixture of isomeric chlorophenols is completely degraded by a defined mixed culture with Pseudomonas sp. strain B13 as a chlorocatechol-dissimilating member of the community. Total degradation of the organic carbon was indicated by release of stoichiometric amounts of chloride and low content of dissolved organic carbon in the cell-free effluents. During adaptation to high loads of chlorophenols the initial meta-cleavage activity was completely replaced by ortho-cleavage activity of type I and II. In the fully acclimated culture, hybrid strains such as Alcaligenes sp. strain A7-2 were detected, which are more competitive than Pseudomonas sp. strain B13 with respect to chlorophenol degradation.