Estimating dataset size requirements for classifying DNA microarray data.

A statistical methodology for estimating dataset size requirements for classifying microarray data using learning curves is introduced. The goal is to use existing classification results to estimate dataset size requirements for future classification experiments and to evaluate the gain in accuracy and significance of classifiers built with additional data. The method is based on fitting inverse power-law models to construct empirical learning curves. It also includes a permutation test procedure to assess the statistical significance of classification performance for a given dataset size. This procedure is applied to several molecular classification problems representing a broad spectrum of levels of complexity.     

On the analytic solution of electrotonic spread in branched passive dendritic trees

From the classical work of Rall it is known that the spread of electric potential in a passive dendritic tree may be obtained by expressing the initial conditions as a linear combination of a set of trigonometric eigenfunctions, each decaying with the associated time constant. It is shown here that in order to evaluate the permissible parameters in these eigenfunctions one may formulate the boundary conditions at all the junctions and endings of the dendritic tree as a set of homogeneous linear equations in which the parameters in the eigenfunctions are the unknowns. These equations have a nontrivial solution if the relevant determinant vanishes, a condition that permits the evaluation of the various parameters, thus providing an analytic approach to the expression of the eigenfunctions as well as the decay time constants. The above approach is illustrated by application to a dendritic tree that has a parent segments and two generations of offspring segments, without any restrictions as to the relative diameters or lengths of the various segments in the tree. General properties of the tree may be readily derived, like the variation of the eigenvalues on scaling of the lengths or diameters of all the segments. A few special cases with specified dimensions of the various segments are derived from the general case. In the case of a dendritic tree that fulfills the equivalent cylinder conditions, all of the eigenvalues and eigefunctions of the tree may be determined by the proposed method, including those that do not apply to the equivalent cylinder. The orthogonality properties of the eigenfunctions are discussed.     

An easy method for the removal of Epon resin from semi-thin sections. Application of the avidin-biotin technique

Summary A simple procedure is described for removing Epon resin from semi-thin 1 m sections, which permits excellent postembedding immunohistochemical staining (avidin-biotin complex technique). The procedure was developed for the detection of growth hormone and prolactin in bovine adenohypophysis fixed with 2% paraformaldehyde and 0.5% glutaraldehyde in 0.1 m sodium cacodylate buffer pH 7.4–7.6. The results indicate that the removal of the epoxy embedding medium prior to the application of the immunohistochemical reagents was essential for the successful localization of the antigenic determinants of the two hormones. The immunocytochemical reactivity was obtained only after treating the sections with a solution of potassium hydroxide in a mixture of absolute methyl alcohol and propylene oxide (Maxwell's solution). An enhanced immunoreactivity was obtained when this treatment was followed by an additional treatment with either 4% hydrogen peroxide or a saturated aqueous solution of sodium metaperiodate. Because of the easy preparation of the Epon removal solution and the good structural preservation without damage to the antigenic determinants, Maxwell's solution is suggested as a good etching agent which can be used in immunohistochemical studies on semi-thin sections with excellent results.     

Diversity, abundance or rare species as a target for the conservation of mammalian carnivores: a case study in Southern Spain

Management goals in protected areas and/or communities usually include diversity as one of the most valuable and confident criteria. Nevertheless, the use of diversity and related indices as a means of evaluating successful management practices could produce conflicting results. Here we report a case study in one of the most important European protected areas. After 6 years of intensive conservation management of the Don~ana National Park, the general abundance and numbers of the target single-species conservation plan (the Iberian lynx) increased, although carnivore community diversity and evenness decreased. This was a result of a disproportionate increase of an oportunistic native species, the red fox. We propose the combined use of diversity, richness and evenness indices when monitoring management practices such as those reported here.     

Differential effects of five types of antipathogenic plant peptides on model membranes

The effects of five antipathogenic plant peptides, wheat α-thionin, potato PTH1 defensin, barley LTP2 lipid transfer protein, and potato tuber DL1 and DL2 defensins, have been tested against phospholipid vesicles (liposomes). Wheat thionin very actively induces aggregation and leakage of negatively charged vesicles. LTP2 displays the same activities, although to a limited extent. Under certain conditions PTH1 and DL2 induce vesicle aggregation, but not leakage. Potato defensin DL1 failed to show any effect on liposomes. The same peptides have been assayed against a plant pathogenic bacterium, both the membrane-active and -inactive compounds having efficient antibacterial action.     

Connexin46 Is Retained as Monomers in a trans-Golgi Compartment of Osteoblastic Cells

Connexins are gap junction proteins that form aqueous channels to interconnect adjacent cells. Rat osteoblasts express connexin43 (Cx43), which forms functional gap junctions at the cell surface. We have found that ROS 17/2.8 osteosarcoma cells, UMR 106-01 osteosarcoma cells, and primary rat calvarial osteoblastic cells also express another gap junction protein, Cx46. Cx46 is a major component of plasma membrane gap junctions in lens. In contrast, Cx46 expressed by osteoblastic cells was predominantly localized to an intracellular perinuclear compartment, which appeared to be an aspect of the TGN as determined by immunofluorescence colocalization. Hela cells transfected with rat Cx46 cDNA (Hela/Cx46) assembled Cx46 into functional gap junction channels at the cell surface. Both rat lens and Hela/Cx46 cells expressed 53-kD (nonphosphorylated) and 68-kD (phosphorylated) forms of Cx46; however, only the 53-kD form was produced by osteoblasts. To examine connexin assembly, monomers were resolved from oligomers by sucrose gradient velocity sedimentation analysis of 1% Triton X-100–solubilized extracts. While Cx43 was assembled into multimeric complexes, ROS cells contained only the monomer form of Cx46. In contrast, Cx46 expressed by rat lens and Hela/Cx46 cells was assembled into multimers. These studies suggest that assembly and cell surface expression of two closely related connexins were differentially regulated in the same cell. Furthermore, oligomerization may be required for connexin transport from the TGN to the cell surface.     

The racemization of free and peptide-bound serine and aspartic acid at 100°C as a function of pH: Implications for in vivo racemization

The relative racemization rates of free and peptide-bound serine and aspartic acid have been studied as a function of pH at 100°C. These results have been used to explain the observed relative in vivo racemization rates of serine and aspartic acid in human tooth dentin and human ocular lens proteins.     

Immunological comparison of desmosomal components from several bovine tissues

A panel of monoclonal antibodies and conventional antisera directed against desmosomal proteins from bovine muzzle epidermis was used to identify immunologically related proteins from two other bovine stratified squamous epithelia, cornea and esophagus. Desmosome-enriched tissue fractions were prepared from epidermis, cornea, and esophagus. These tissue extracts were electrophoresed on sodium dodecyl sulfate (SDS)-polyacrylamide gels, blotted onto nitrocellulose paper, and labeled using an indirect immunoperoxidase technique. Labeling with the conventional antisera demonstrates that each of the previously characterized epidermal desmosomal proteins or protein families has an immunologically cross-reacting counterpart in cornea and esophagus. However, chemical differences between homologous desmosomal proteins in these three tissues have also been detected. The corresponding proteins in the different tissues have similar but not always identical apparent molecular weights. Moreover, tissue-restricted antigenic determinants were detected in two of the desmosomal proteins families using four monoclonal antibodies, each of which recognizes a distinct antigenic determinant.     

Self vs nonself in tissue assembly: Correlated changes in recognition behavior and tissue cohesiveness

Vertebrate embryonic cell populations of unlike kind, when combined in vitro, typically spread around and sort out from one another in combination-specific patterns, whereas like cell populations merely coalesce. These differing responses to self and nonself constitute one form of morphogenetic self-recognition behavior. Prolonged shaker-flask culturing and dissociation and reaggregation of embryonic chick heart tissue were both previously shown to reverse the tissue's spreading behavior with liver. Here, we show that these treatments simultaneously initiate, in heart tissue, a “foreign” spreading reaction toward untreated heart. Moreover, the direction of this heart-heart spreading can be deduced from the change in direction of heart-liver spreading. This suggests that certain properties of heart tissue participate in the determination of both the foreign- and the self-recognition behaviors studied here. The differential adhesion hypothesis postulates that these properties are the intensities of tissue cohesion, with less cohesive tissues enveloping more cohesive ones. If so, our observations imply that heart fragments precultured $\text{1}\text{2}$ day should be more cohesive than $\text{1}\text{2}\text{-}\text{day}$ precultured heart reaggregates, but less cohesive than heart fragments precultured $\text{2}\text{1}\text{2}$ days. With our centrifugation assay, in which relative tissue cohesiveness is assessed by the relative roundness of centrifuged aggregates at shape equilibrium, we confirm this prediction.     

Escherichia coli mutants deficient in the aspartate and aromatic amino acid aminotransferases.

Two new mutations are described which, together, eliminate essentially all the aminotransferase activity required for de novo biosynthesis of tyrosine, phenylalanine, and aspartic acid in a K-12 strain of Escherichia coli. One mutation, designated tyrB, lies at about 80 min on the E. coli map and inactivates the "tyrosine-repressible" tyrosine/phenylalanine aminotransferase. The second mutation, aspC, maps at about 20 min and inactivates a nonrespressible aspartate aminotransferase that also has activity on the aromatic amino acids. In ilvE- strains, which lack the branched-chain amino acid aminotransferase, the presence of either the tyrosine-repressible aminotransferase or the aspartate aminotransferase is sufficient for growth in the absence of exogenous tyrosine, phenylalanine, or aspartate; the tyrosine-repressible enzyme is also active in leucine biosynthesis. The ilvE gene product alone can reverse a phenylalanine requirement. Biochemical studies on extracts of strains carrying combinations of these aminotransferase mutations confirm the existence of two distinct enzymes with overlapping specificities for the alpha-keto acid analogues of tyrosine, phenylalanine, and aspartate. These enzymes can be distinguished by electrophoretic mobilities, by kinetic parameters using various substrates, and by a difference in tyrosine repressibility. In extracts of an ilvE- tyrB- aspC- triple mutant, no aminotransferase activity for the alpha-keto acids of tyrosine, phenylalanine, or aspartate could be detected.