Inhibition or down-regulation of protein kinase C attenuates late phase p70s6k activation induced by epidermal growth factor but not by platelet-derived growth factor or insulin.

The late phase of the time-dependent epidermal growth factor (EGF)-induced biphasic activation of the p70s6k is selectively attenuated by the specific PKC inhibitor, CGP 41,251, a staurosporine derivative. At a 40-fold lower concentration than CGP 41,251, staurosporine inhibits both phases of S6 kinase activation to the same extent, whereas the inactive staurosporine derivative CGP 42,700 shows no effect on either phase. Platelet-derived growth factor (PDGF) and insulin also induce biphasic S6 kinase activation, but in neither case is either phase of activation affected by the presence of CGP 41,251. This finding was unexpected in the case of PDGF, which is a potent activator of PKC and whose receptor directly interacts with phospholipase C gamma 1. However, similar results were obtained following down-regulation of PKC by prolonged 12-O-tetradecanoylphorbol-13-acetate treatment. Therefore, even though EGF and PDGF induce PKC activation, PDGF, unlike EGF, does not appear to use this signaling pathway for late phase p70s6k activation.     

The new higher level classification of eukaryotes with emphasis on the taxonomy of protists

This revision of the classification of unicellular eukaryotes updates that of Levine et al. (1980) for the protozoa and expands it to include other protists. Whereas the previous revision was primarily to incorporate the results of ultrastructural studies, this revision incorporates results from both ultrastructural research since 1980 and molecular phylogenetic studies. We propose a scheme that is based on nameless ranked systematics. The vocabulary of the taxonomy is updated, particularly to clarify the naming of groups that have been repositioned. We recognize six clusters of eukaryotes that may represent the basic groupings similar to traditional "kingdoms." The multicellular lineages emerged from within monophyletic protist lineages: animals and fungi from Opisthokonta, plants from Archaeplastida, and brown algae from Stramenopiles.     

A model of motor and sensory axon activation in the median nerve using surface electrical stimulation

Surface electrical stimulation has the potential to be a powerful and non-invasive treatment for a variety of medical conditions but currently it is difficult to obtain consistent evoked responses. A viable clinical system must be able to adapt to variations in individuals to produce repeatable results. To more fully study the effect of these variations without performing exhaustive testing on human subjects, a system of computer models was created to predict motor and sensory axon activation in the median nerve due to surface electrical stimulation at the elbow. An anatomically-based finite element model of the arm was built to accurately predict voltages resulting from surface electrical stimulation. In addition, two axon models were developed based on previously published models to incorporate physiological differences between sensory and motor axons. This resulted in axon models that could reproduce experimental results for conduction velocity, strength-duration curves and activation threshold. Differences in experimentally obtained action potential shape between the motor and sensory axons were reflected in the models. The models predicted a lower threshold for sensory axons than motor axons of the same diameter, allowing a range of sensory axons to be activated before any motor axons. This system of models will be a useful tool for development of surface electrical stimulation as a method to target specific neural functions.     

Analysis of the Pan Genome of <Emphasis Type="Italic">Campylobacter jejuni</Emphasis> Isolates Recovered from Poultry by Pulsed-Field Gel Electrophoresis,Multilocus Sequence Typing (MLST), and Repetitive Sequence Polymerase Chain Reaction (rep-PCR) Reveals Different Discriminatory Capabilities

Campylobacter jejuni is one of the leading bacterial causes of food-borne illness in the USA. Molecular typing methods are often used in food safety for identifying sources of infection and pathways of transmission. Moreover, the identification of genetically related isolates (i.e., clades) may facilitate the development of intervention strategies for control and prevention of food-borne diseases. We analyzed the pan genome (i.e., core and variable genes) of 63 C. jejuni isolates recovered from chickens raised in conventional, organic, and free-range poultry flocks to gain insight into the genetic diversity of C. jejuni isolates recovered from different environments. We assessed the discriminatory power of three genotyping methods [i.e., pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and repetitive extragenic palindromic polymerase chain reaction (rep-PCR)]. The rep-PCR fingerprint was generated by determining the presence of repetitive sequences that are interspersed throughout the genome via repetitive extragenic palindromic PCR, enterobacterial repetitive intergenic consensus sequence PCR (ERIC-PCR), and BOX element PCR (BOX-PCR) and combining the data to form a composite fingerprint. The genetic fingerprints were subjected to computer-assisted pattern analysis. Comparison of the three genotypic methods revealed that repREB-PCR showed greater discriminatory power than PFGE and MLST. ERIC-PCR and BOX-PCR yielded the highest number of PCR products and greatest reproducibility. Regardless of the genotyping method, C. jejuni isolates recovered from chickens reared in conventional, organic, and free-range environments all exhibit a high level of genotypic diversity.     

Role of subunit crosslinking in fibrin solubility.

¹²⁵I-labelled fibrinogen was clotted by thrombin in the presence of activated Factor XIII and the rates of formation of γ dimers and α polymers were measured. These changes in fibrin subunits were correlated with the solubility of fibrin in 1% monochloroacetic acid. In the presence of the factor XIIIa inhibitor, glycine methyl ester, fibrin solubility was found to depend on the level of α polymers formed. A preferential inhibition of α polymer formation rather than γ dimer was observed in the presence of glycine methyl ester.     

Biomechanical analysis of the Rolled (RLD) leaf phenotype of maize

The pleiotropic effects of the Rld1-O/+ mutation of Zea mays (Poaceae) on leaf phenotype include a suppression of normal transverse unrolling, a reversed top/bottom epidermal polarity, and an apparently straighter longitudinal shape. According to engineering shell theory, there might be mechanical coupling between transverse and longitudinal habit, i.e., the leaf rolling itself might produce the longitudinal straightening. We tested this possibility with quantitative curvature measurements and mechanical uncoupling experiments. The contributions of elastic bending under self weight, mechanical coupling, and rest state of leaf parts to the longitudinal and transverse habit were assessed in Rld1-O/+ mutants and a population of sibling +/+ segregants. Elastic bending and curvature coupling are shown to be relatively unimportant. The Rld1-O/+ mutation is shown to alter not only the unrolling process, but also the developmental longitudinal curving in the growing leaf, leading to a straighter midrib and a rolled lamina. The Rld1-O/+ mutant is thus a suitable model to study the relation between tissue polarity and differential curvature development in the maize leaf. Since on the abaxial side of the leaf, more abundant sclerenchyma is found in +/+ than in Rld1-O/+, a gradient in sclerification may contribute to the development of midrib curvature.     

Trapping of a novel coenzyme A containing intermediate of 3-hydroxy-3-methylglutaryl-CoA synthase.

Carboxyamidomethylated J chain was shown to be an excellent substrate for the enzyme, pyrollidone carboxylyl peptidase, which specifically removed the cyclized amino terminal glutamyl residue. J chain lacking the “blocked” PCA group was subjected to automated Edman degradation and the amino terminal amino acid sequence determined as: PCA-Glu-Asp-Glu-Arg-Ile-Val-Leu-Val-Asp-Asn-Lys-CMCys-Lys-CMCys-Ala-Arg. Previous studies by others have identified a disulfide bridge between the heavy chain of immunoglobulins and a tripeptide identical in composition with the sequence at positions 15–17 in the J chain. These two sets of data locate the linkage of immunoglobulin heavy chain with Cys 15 of the J chain.     

The construction and replication properties of hybrid plasmids composed of the r-determinant of R100.1 and the plasmids pCRI or pSC201

Summary We have cloned the entire r-determinant of the antibiotic resistance plasmid R100.1 on the plasmid vectors pCR1 and pSC201. We find that the hybrid plasmids segregate from cultures in which replication of the vector is blocked. This suggests that the r-det is not capable of autonomous replication.