Optimizing purebred selection for crossbred performance using QTL with different degrees of dominance

A method was developed to optimize simultaneous selection for a quantitative trait with a known QTL within a male and a female line to maximize crossbred performance from a two-way cross. Strategies to maximize cumulative discounted response in crossbred performance over ten generations were derived by optimizing weights in an index of a QTL and phenotype. Strategies were compared to selection on purebred phenotype. Extra responses were limited for QTL with additive and partial dominance effects, but substantial for QTL with over-dominance, for which optimal QTL selection resulted in differential selection in male and female lines to increase the frequency of heterozygotes and polygenic responses. For over-dominant QTL, maximization of crossbred performance one generation at a time resulted in similar responses as optimization across all generations and simultaneous optimal selection in a male and female line resulted in greater response than optimal selection within a single line without crossbreeding. Results show that strategic use of information on over-dominant QTL can enhance crossbred performance without crossbred testing.     

Specific enzymatic dephosphorylation of the retinoblastoma protein.

The retinoblastoma gene product (RB) undergoes cell cycle-dependent phosphorylation and dephosphorylation. Pulse-chase experiments revealed that the change in RB gel electrophoretic migration which occurs near mitosis is due to enzymatic dephosphorylation (J. W. Ludlow, J. Shon, J. M. Pipas, D. M. Livingston, and J. A. DeCaprio, Cell 60:387-396, 1990). To determine the precise timing of RB dephosphorylation and whether a specific phosphatase is active in this process, we have utilized a nocodazole block and release protocol which allows a large population of cells to progress synchronously through mitosis. In such experiments, RB dephosphorylation began during anaphase and continued until complete dephosphorylation was apparent in the ensuing G1 period. In addition, late mitotic cell extracts were capable of dephosphorylating RB in vitro. This RB-specific mitotic phosphatase activity was more active in anaphase extracts than in pro- or metaphase extracts, which is consistent with the results obtained in vivo. Okadaic acid and protein phosphatase inhibitors 1 and 2 inhibited this specific RB phosphatase activity. These results suggest a role for serine and threonine phosphoprotein phosphatase type 1 in the late mitotic dephosphorylation of RB.     

Termite Prey Specialization in the Pitcher Plant Nepenthes albomarginata--Evidence from Stable Isotope Analysis

Old World pitcher plants (Nepenthes spp., Nepenthaceae) trapand digest invertebrate prey to derive nutrients, primarilynitrogen (N). In the majority of lowland Nepenthes species studiedto date, ants (Hymenoptera, Formicidae) are numerically thedominant prey taxon. Nepenthes albomarginata is unusual in showingan apparent bias towards the capture of termites (Isoptera).We tested the hypothesis that N. albomarginata derives N fromtermite capture, by comparison of foliar stable N isotope abundance(     

8-Hydroxy-5-deazaflavin-reducing hydrogenase from Methanobacterium thermoautotrophicum: 2. Kinetic and hydrogen-transfer studies

Steady-state kinetic parameters have been obtained for the pure 8-hydroxy-5-deazaflavin-reducing hydrogenase. With H2 and 8-hydroxy-5-deazariboflavin (F0) as substrates, Km (H2) = 12 microM, Km (F0) = 26 microM, and Kcat = 225 s-1. In the back-direction, F0H2 is reoxidized (anaerobically) at 225 s-1. Initial velocity patterns, product inhibition patterns, dead-end inhibition by carbon monoxide, and transhydrogenation to Procion Red HE-3B suggest a two-site hybrid ping-pong mechanism. A kinetic derivation for the rate equation is provided in the Appendix. Studies with D2 and with D2O reveal that no steps involving D transfer are substantially rate determining. Further, D2 yields F0H2 with no deuterium at C5 while in D2O a 5-monodeuterio F0H2 product is formed, indicating complete exchange of hydrogens from H2 with solvent before final transfer of a hydride ion out from reduced enzyme to C5 of F0.     

Expression of {beta}-galactoside {alpha}2,6 sialyltransferase blocks synthesis of polysialic acid in Xenopus embryos

Polysialic acid is a developmentally regulated carbohydratestructure found on neural cell adhesion molecules (NCAM). Expressionof ß-galactoside 2,6-sialyltransferase in Xenopusembryos, by injection of mRNA, prevents the polysialylationof NCAM, presumably by introducing a different type of sugarlinkage that terminates chain elongation. Abnormalities in neuraldevelopment result from this treatment, but in general the bodyplan of the injected embryos is not severely affected. The resultsprovide evidence that the mis-expression of glycosyltransferasescan be used to interfere with the normal pattern of glycosylationin whole organisms. glycosylation NCAM polysialic acid Xenopus development     

Effect of limited homology on gene conversion in a Saccharomyces cerevisiae plasmid recombination system.

Plasmids containing heteroallelic copies of the Saccharomyces cerevisiae HIS3 gene undergo intramolecular gene conversion in mitotically dividing S. cerevisiae cells. We have used this plasmid system to determine the minimum amount of homology required for gene conversion, to examine how conversion tract lengths are affected by limited homology, and to analyze the role of flanking DNA sequences on the pattern of exchange. Plasmids with homologous sequences greater than 2 kilobases have mitotic exchange rates as high as 2 x 10(-3) events per cell per generation. As the homology is reduced, the exchange rate decreases dramatically. A plasmid with 26 base pairs (bp) of homology undergoes gene conversion at a rate of approximately 1 x 10(-10) events per cell per generation. These studies have also shown that an 8-bp insertion mutation 13 bp from a border between homologous and nonhomologous sequences undergoes conversion, but that a similar 8-bp insertion 5 bp from a border does not. Examination of independent conversion events which occurred in plasmids with heteroallelic copies of the HIS3 gene shows that markers within 280 bp of a border between homologous and nonhomologous sequences undergo conversion less frequently than the same markers within a more extensive homologous sequence. Thus, proximity to a border between homologous and nonhomologous sequences shortens the conversion tract length.     

Effect of postnatal development on calcium currents and slow charge movement in mammalian skeletal muscle

Single- (whole-cell patch) and two-electrode voltage-clamp techniques were used to measure transient (Ifast) and sustained (Islow) calcium currents, linear capacitance, and slow, voltage-dependent charge movements in freshly dissociated fibers of the flexor digitorum brevis (FDB) muscle of rats of various postnatal ages. Peak Ifast was largest in FDB fibers of neonatal (1-5 d) rats, having a magnitude in 10 mM external Ca of 1.4 +/- 0.9 pA/pF (mean +/- SD; current normalized by linear fiber capacitance). Peak Ifast was smaller in FDB fibers of older animals, and by approximately 3 wk postnatal, it was so small as to be unmeasurable. By contrast, the magnitudes of Islow and charge movement increased substantially during postnatal development. Peak Islow was 3.6 +/- 2.5 pA/pF in FDB fibers of 1-5-d rats and increased to 16.4 +/- 6.5 pA/pF in 45-50-d-old rats; for these same two age groups, Qmax, the total mobile charge measurable as charge movement, was 6.0 +/- 1.7 and 23.8 +/- 4.0 nC/microF, respectively. As both Islow and charge movement are thought to arise in the transverse-tubular system, linear capacitance normalized by the area of fiber surface was determined as an indirect measure of the membrane area of the t-system relative to that of the fiber surface. This parameter increased from 1.5 +/- 0.2 microF/cm2 in 2-d fibers to 2.9 +/- 0.4 microF/cm2 in 44-d fibers. The increases in peak Islow, Qmax, and normalized linear capacitance all had similar time courses. Although the function of Islow is unknown, the substantial postnatal increase in its magnitude suggests that it plays an important role in the physiology of skeletal muscle.     

Myoglobin: cytochrome b5 interactions and the kinetic mechanism of metmyoglobin reductase

Metmyoglobin (metMb) reduction by metMb reductase from heart muscle requires cytochrome b5 as electron-transfer mediator. The existence of a metMb-ferrous cytochrome b5 complex is demonstrated by mutual perturbation of the proteins' respective electrophoretic titration curves between pH 4 and 7. The same technique shows a preferential binding of cytochrome b5 over metMb by the enzyme. The paramagnetic hyperfine shifts in the cytochrome b5 1H NMR spectrum are perturbed by metMb, indicating the formation of a specific bimolecular complex with a 1:1 stoichiometry and a binding constant estimated to be less than 10 microM. The resonances assigned to the cytochrome b5 heme 6-propionate methylene group exhibit the largest complexation shifts. Computer modeling implicates lysines 47, 50, and 98 of metMb as contact points with cytochrome b5 carboxylate residues 43, 44, 60, and heme 6-propionate. The mechanism of the enzymatic reduction establishes metMb reductase as an NADH-cytochrome b5 oxidoreductase. Cytochrome b5 is reduced at near diffusion-controlled rates by the enzyme with a turnover number of 1000 min-1 X Km for the cytochrome is 0.9 microM versus 100 microM reported for the erythrocyte enzyme. Ferrous cytochrome b5 then reduces metMb nonenzymatically with an apparent rate constant of 4.9 X 10(4) M-1 min-1 X Acetylation of metMb, which does not affect its oxygen affinity or chemical reduction, renders it a poor substrate for enzymatic reduction. This study suggests a function for the three exterior lysine residues conserved in all mammalian myoglobin sequences: they are contact points for complexation with cytochrome b5.