Retention of low-fitness genotypes over six decades of admixture between native and introduced tiger salamanders

Background  

Introductions of non-native tiger salamanders into the range of California tiger salamanders have provided a rare opportunity to study the early stages of secondary contact and hybridization. We produced first- and second-generation hybrid salamanders in the lab and measured viability among these early-generation hybrid crosses to determine the strength of the initial barrier to gene exchange. We also created contemporary-generation hybrids in the lab and evaluated the extent to which selection has affected fitness over approximately 20 generations of admixture. Additionally, we examined the inheritance of quantitative phenotypic variation to better understand how evolution has progressed since secondary contact.     

Reversible hyperphosphorylation and reorganization of vimentin intermediate filaments by okadaic acid in 9L rat brain tumor cells.

Okadaic acid (OA), a protein phosphatase inhibitor, was found to induce hyperphosphorylation and reorganization of vimentin intermediate filaments in 9L rat brain tumor cells. The process was dose dependent. Vimentin phosphorylation was initially enhanced by 400 nM OA in 30 min and reached maximal level (about 26-fold) when cells were treated with 400 nM OA for 90 min. Upon removal of OA, dephosphorylation of the hyperphosphorylated vimentin was observed and the levels of phosphorylation returned to that of the controls after the cells recovered under normal growing conditions for 11 h. The phosphorylation and dephosphorylation of vimentin induced by OA concomitantly resulted in reversible reorganization of vimentin filaments and alteration of cell morphology. Cells rounded up as they were entering mitosis in the presence of OA and returned to normal appearance after 11 h of recovery. Immuno-staining with anti-vimentin antibody revealed that vimentin filaments were disassembled and clustered around the nucleus when the cells were treated with OA but subsequently returned to the filamentous states when OA was removed. Two-dimensional electrophoresis analysis further revealed that hyperphosphorylation of vimentin generated at least seven isoforms having different isoelectric points. Furthermore, the enhanced vimentin phosphorylation was accompanied by changes in the detergent-solubility of the protein. In untreated cells, the detergent-soluble and -insoluble vimentins were of equal amounts but the solubility could be increased when vimentins were hyperphosphorylated in the presence of OA. Taken together, the results indicated that OA could be involved in reversible hyperphosphorylation and reorganization of vimentin intermediate filaments, which may play an important role in the structure-function regulation of cytoskeleton in the cell.     

Kinetic analysis of protonation of a specific site on a buffered surface of a macromolecular body

The kinetics of protonation of a specific site on a macromolecular structure (micelle) in buffered solution was studied with the purpose of evaluating the effect of buffer on the observed dynamics. The experimental system consisted of the following elements: Brij 58 micelles serving as homogeneous uncharged macromolecular bodies, bromocresol green, a well-adsorbed proton detector, and 2-naphthol-3,6-disulfonate as a proton emitter in the bulk. Imidazole was the mobile buffer while neutral red, which has a high affinity for the micellar surface, served as the immobile buffer. An intensive laser pulse ejects a proton from the proton emitter, and the subsequent proton-transfer reactions are measured by fast spectrophotometric methods. The dynamics of proton pulse in buffered solution are characterized by a very rapid trapping of the discharged protons by the abundant buffer molecules. This event has a major effect on the kinetic regime of the reaction. During the first 200 ns the proton flux is rate limited by free-proton diffusion. After this period, when the free-proton concentration decayed to the equilibrium level, the relaxation of the system is carried out by the diffusion of buffer. Thus in the buffered biochemical system, at neutral pH, most of proton flux between active sites and bulk is carried out by buffer molecules--not by diffusion of free protons. Surface groups on a high molecular weight body exchange protons among them at a very fast rate. This reaction has a major role on proton transfer from a specific site to the bulk.(ABSTRACT TRUNCATED AT 250 WORDS)     

Identification of a complex of the three forms of the rat liver asialoglycoprotein receptor

We have generated antibodies against synthetic peptides which represent the carboxyl terminus of either the major, or the two minor, forms of the rat hepatic lectin which recognizes galactose-terminated glycoproteins (asialoglycoproteins). The antibodies were shown to be specific for the form of the lectin containing the immunizing peptide sequence by the following: reaction with purified lectin after sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoprecipitation of sodium dodecyl sulfate-denatured lectin, immunoprecipitation of lectin synthesized in vitro. These antibodies, however, precipitated all three rat hepatic lectin forms from nonionic detergent extracts of hepatocytes labeled with 125I via the lactoperoxidase catalyzed technique. A similar result was obtained if antibody was bound to intact cells prior to solubilization with detergent and collection of the immune complexes. We conclude that at least the plasma membrane-associated fraction of the rat hepatic lectin forms exists as a heterotypic complex.     

In vitro assessment of the toxicity of metal compounds

A review of in vitro mutagenesis assessment of metal compounds in mammalian and nonmammalian test systems has been compiled. Prokaryotic assays are ineffective or inconsistent in their detection of most metals as mutagens, with the notable exception of hexavalent chromium. Mammalian assay systems appear to be similarly inappropriate for the screening of metal compounds based upon the limited number of studies that have employed those compounds having known carcinogenic activity. Although of limited value as screening tests for the detection of potentially carcinogenic metal compounds, the well-characterized in vitro mutagenesis systems may prove to be of significant value as a means to elucidate mechanisms of metal genotoxicity.     

ATP synthase from bovine mitochondria: sequences of imported precursors of oligomycin sensitivity conferral protein, factor 6, and adenosinetriphosphatase inhibitor protein

Oligomycin sensitivity conferral protein (OSCP), factor 6 (F6), and ATPase inhibitor protein are all components of the ATP synthase complex of bovine mitochondria. They are encoded in nuclear DNA. Complementary DNA clones encoding the precursors of these proteins have been isolated from a bovine library by using mixtures of synthetic oligonucleotides as hybridization probes, and their DNA sequences have been determined. The deduced protein sequences show that the OSCP, F6, and inhibitor proteins have N-terminal presequences of 23, 32, and 25 amino acids, respectively. These presequences are not present in the mature proteins. It is assumed that they serve to direct the proteins into the mitochondrial matrix. The cDNA clones have also been employed as hybridization probes to investigate the genetic complexity of the three proteins in cows and humans. These experiments indicate that the bovine and human inhibitor and bovine F6 proteins are encoded by single genes but suggest the possibility of the presence in both species of more than one gene (or pseudogenes) for the OSCP.     

Magnetic resonance imaging and electromyography as indexes of muscle function.

Electromyography (EMG) is commonly used to determine the electrical activity of skeletal muscle during contraction. To date, independent verification of the relationship between muscle use and EMG has not been provided. It has recently been shown that relaxation- (e.g., T2) weighted magnetic resonance images (MRI) of skeletal muscle demonstrate exercise-induced contrast enhancement that is graded with exercise intensity. This study was conducted to test the hypothesis that exercise-induced magnetic resonance (MR) contrast shifts would relate to EMG amplitude if both measures reflect muscle use during exercise. Both MRI and EMG data were collected for separate eccentric (ECC) and concentric (CON) exercise of increasing intensity to take advantage of the fact that the rate of increase and amplitude of EMG activity are markedly greater for CON muscle actions. Seven subjects 30 +/- 2 (SE) yr old performed five sets of 10 CON or ECC arm curls with each of four resistances representing 40, 60, 80, and 100% of their 10 repetition maximum for CON curls. There was 1.5 min between sets and 30 min between bouts (5 sets of 10 actions at each relative resistance). Multiple echo, transaxial T2-weighted MR images (1.5 T, TR/TE 2,000/30) were collected from a 7-cm region in the middle of the arm before exercise and immediately after each bout. Surface EMG signals were collected from both heads of the biceps brachii and the long head of the triceps brachii muscles. CON and ECC actions resulted in increased integrated EMG (IEMG) and T2 values that were strongly related (r = 0.99, P < 0.05) with relative resistance.(ABSTRACT TRUNCATED AT 250 WORDS)