Selection for components related to body composition in mice: direct responses

Summary Replicated within full-sib family single-trait selection was conducted for 10 generations in mice for (1) high or low 12-week epididymal fat pad percentage (100 x epididymal fat pad weight/body weight) or (2) high or low 12-week hind carcass percentage (100 x hind carcass weight/body weight). Pooled realized heritabilities based on high, low and divergent selection were 0.66±0.09, 0.65±0.13 and 0.66±0.05 for epididymal fat pad percentage and 0.48±0.08, 0.33±0.08 and 0.40±0.04 for hind carcass percentage. The pooled realized genetic correlation (r_{G R}) between epididymal fat pad percentage and hind carcass percentage based on divergence was –0.67±0.04. Other estimates of (r_{G R}) were: epididymal fat pad percentage with body weight (0.57±0.05); epididymal fat pad percentage with epididymal fat pad weight (1.17±0.05); hind carcass percentage with body weight (–0.61±0.09); hind carcass percentage with hind carcass weight (–0.05±0.11). Indirect measures of fat and lean tissue percentages were highly heritable, and (r_{G R}) between them would be desirable from the standpoint of analogous types of traits in livestock. In the same context, undesirable (r_{G R})'s were found between epididymal fat pad percentage and body weight and between hind carcass percentage and body weight.Paper No. 10957 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh, North Carolina 27695-7601, USA. The use of trade names in this publication does not imply endorsement by the North Carolina Agricultural Research Service of the products named, nor criticism of similar ones not mentioned     

Amphomycin: A tool to study protein N-glycosylation

Radio-labelled amphomycin (³H-amphomycin) forms a complex with dolichylmonophosphate in presence of Ca²⁺. Complex formation has also been documented with retinylmonophosphate and perhydromonoeneretinylmonophosphate. Analysis of the space-filling model suggested both fatty acylated aspartic acid residue at the N-terminus of the lipopeptide and phosphate head group of dolichylmonophosphate are necessary for the complex formation. The binding ability of amphomycin is then utilized to localize dolichylmonophosphate in the microsomal membrane. Studies with microsomal membranes from hen oviduct suggested that dolichylmonophosphate is located in the cytoplasmic side of the membrane.     

Calcium-activated neutral protease and its endogenous inhibitor Activation at the cell membrane and biological function

The structures of calcium-activated neutral protease (CANP) and its endogenous inhibitor elucidated recently have revealed novel features with respect to their structure-function relationship and enzyme activity regulation. The protease is regarded as a proenzyme which can be activated at the cell membrane in the presence of Ca²⁺ and phospholipid, and presumably regulates the functions of proteins, especially membrane-associated proteins, by limited proteolysis. Protein kinase C is hydrolysed and activated by CANP at the cell membrane to a cofactor-independent form. These results are reviewed and the possible involvement of CANP in signal transduction is discussed.     

Subcellular Location and Neuronal Release of Diazepam Binding Inhibitor

Diazepam binding inhibitor (DBI), a peptide located in CNS neurons, blocks the binding of benzodiazepines and beta-carbolines to the allosteric modulatory sites of gamma-aminobutyric acid (GABAA) receptors. Subcellular fractionation studies of rat brain indicate that DBI is compartmentalized. DBI-like immunoreactivity is highly enriched in synaptosomes obtained by differential centrifugation in isotonic sucrose followed by a Percoll gradient. In synaptosomal lysate, DBI-like immunoreactivity is primarily associated with synaptic vesicles partially purified by differential centrifugation and continuous sucrose gradient. Depolarization induced by high K+ levels (50 mM) or veratridine (50 microM) released DBI stored in neurons of superfused slices of hypothalamus, hippocampus, striatum, and cerebral cortex. The high K+ level-induced release is Ca2+ dependent, and the release induced by veratridine is blocked by 1.7 microM tetrodotoxin. Depolarization released GABA and Met5-enkephalin-Arg6-Phe7 together with DBI. DBI is also released by veratridine depolarization, in a tetrodotoxin-sensitive fashion, from primary cultures of cerebral cortical neurons, but not from cortical astrocytes. Depolarization fails to release DBI from slices of liver and other peripheral organs. These data support the view that DBI may be released as a putative neuromodulatory substance from rat brain neurons.     

冬小麦春化过程中可溶性蛋白质组成的变化与形态发生的关系

冬小麦“农大139”经40天左右的春化处理才能迅速而整齐地抽穗,但经14—21天低温处理,已经具有在夏季抽穗的可能性,虽然抽穗推迟且极不整齐;再将春化时间延长,则抽穗百分比增加,且从播种到抽穗的时间缩短。这表明,春化过程中低温对发育的作用有两种效应:前期低温是诱发生理状态的转变,后期低温则只具有加速发育的作用,两个时期的转变是在春化的中期。蛋白质合成抑制剂乙基硫氨酸和对-氟苯丙氨酸能抑制冬小麦的春化,抑制时期也是在春化过程的中期。不同时间低温处理后冬小麦幼芽中可溶性蛋白质含量及组成发生了变化,春化过程中期(低温处理14天之后)不仅含量比对照增加了一倍,而且有新的蛋白质谱带出现。春麦中无类似现象,未经低温处理的春麦已含有冬麦中新出现的谱带。说明冬小麦春化过程的第14—21天左右是与春化过程有关的蛋白质合成的关键时期,该时期新合成的蛋白质与植株的发育状态之间存在着密切的相关关系。     

Purification of an acid protease from Mucor rouxii that inactivates chitin synthetase

An acid protease was purified from the mycelial form of Mucor rouxii by a method which involved salt and acid precipitation, gel filtration and anion-exchange chromatography. The enzyme had a molecular mass of 16,000 Da. Its optimum pH was 4.0, maximal activity was obtained at 50°C, and it was inactivated at 70°C. It was not affected by leupeptin or N -p-tosyl-L-lysine chloromethyl ketone (TLCK) but diazoacetyl-DL-norleucine methyl ester (DNME) in the presence of Cu²⁺ and more noticeably pepstatin A, strongly inhibited the activity. This acid protease did not activate zymogenic chitin synthetase from the fungus, but brought about its inactivation even at low concentrations and after short periods of incubation time.Abbreviations TLCK N -p-tosyl-L-lysine chloromethyl ketone - DNME diazoacetyl-DL-norleucine methyl ester - TCA trichloroacetic acid - SDS sodium dodecyl sulfate     

The highly modified membrane of cornified cells in stratified squamous epithelia: A comparison of heterogenous deposits in keratinized and nonkeratinized epithelia

Summary The chemical nature of the thickened plasma membrane of cornified cells in stratified squamous epithelium was investigated in comparison with that in noncornified epithelium. Localizations of transglutaminase, molecular weight 92000 daltons, and detection of epidermal cysteine proteinase inhibitor were effected with a monoclonal antibody and a monospecific rabbit anti-inhibitor immunoglobulin, respectively, directed to the antigens. N-(7-dimethylamino-4-methylcoumarinyl) maleimide was used to demonstrate S-S cross-linking. In all keratinizing epithelia, the enzyme and inhibitor were deposited on membranes of granular cells. S-S bonds were formed in cornification with the appearance of electron-dense material by the inner leaflet. Both enzyme and inhibitors occurred on the corneal epithelium, but S-S linkage and the thickened plasma membrane did not form even at the last stage of maturation. On the other hand, the internal vaginal epithelium in the proestrous stage without keratinization contained the enzyme, but neither inhibitor nor S-S linkage. Both antigens and S-S bonds were detected when keratinization proceeded during estrus. The staining patterns in the epithelium near the vaginal introitus were identical to those in the skin. Cuboidal and simple epithelia exhibited none of those constituents. The findings indicated that heterogenous components contribute to modification of the plasma membrane of cornified cells, but S-S cross-linkages are associated exclusively with formation of the ultrastructurally unique membrane structure. In addition, findings suggested hormonal regulation in the chemical modification of the membrane in estrogen-sensitive internal vaginal epithelium.     

光质对一品红叶离体培养中形态发生及生化变化的影响

本文讨论了将光质应用于一品红(Euphorbia pulcherrima)大规模细胞繁殖及形态发生调控的可行性,并测定了一些形态及生化指标作为这种调控的参数。 以一品红试管苗的叶片作为外植体,置于MS培养基中并分别培养于白(W)、红(R)、黄(Y)、蓝(B)、绿(G)色光及黑暗(D)下,一个月后进行测定。蓝光和黄光明显促进出苗。而愈伤组织形成的情况则正相反,红光利于愈伤组织的形成,黄光效果最差。过氧化物酶、过氧化氢酶活力及蛋白质、RNA、碳水化合物的含量与愈伤组织形成有较为一致的关系,但与分化不直接相关。 实验中还测定了叶绿素a、叶绿素b及类胡萝卜素的含量。白光与蓝光均能促进叶绿素及类胡萝卜素的合成,但蓝光中chl b/chl a的比率较高。此外,本文尚涉及暗处理中的晶质化现象(vitrification)及其成因。     

The gene for trypsin inhibitor CMe is regulated in trans by the lys 3a locus in the endosperm of barley (Hordeum vulgare L.)

Summary A cDNA encoding trypsin inhibitor CMe from barley endosperm has been cloned and characterized. The longest open reading frame of the cloned cDNA codes for a typical signal peptide of 24 residues followed by a sequence which is identical to the known amino acid sequence of the inhibitor, except for an Ile/Leu substitution at position 59. Southern blot analysis of wheat-barley addition lines has shown that chromosome 3H of barley carries the gene for CMe. This protein is present at less than 2%–3% of the wild-type amount in the mature endosperm of the mutant Risø 1508 with respect to Bomi barley, from which it has been derived, and the corresponding steady state levels of the CMe mRNA are about I%. One or two copies of the CMe gene (synonym Itc1) per haploid genome have been estimated both in the wild type and in the mutant, and DNA restriction patterns are identical in both stocks, so neither a change in copy number nor a major rearrangement of the structural gene account for the markedly decreased expression. The mutation at the lys 3a locus in Risø 1508 has been previously mapped in chromosome 7 (synonym 5H). A single dose of the wild-type allele at this locus (Lys 3a) restores the expression of gene CMe (allele CMe-1) in chromosome 3H to normal levels.     

Voltage clamping with single microelectrodes: Comparison of the discontinuous mode and continuous mode using the Axoclamp 2A amplifier

Summary The voltage clamp technique is a powerful method for studying the physiology of excitable membrane. This technique has made possible the determination of ionic responses generated by activation of either receptor-mediated or voltage-dependent processes. The development of the whole-cell, tight-seal voltage clamp method has allowed the analysis and examination of membrane physiology at the single cell level. The method allows the characterization of voltage-dependent ionic conductances both at the macroscopic (whole-cell) and at the microscopic (unitary conductance or single channel) level in cells less than 10 µm in diameter, a feat difficult to achieve with conventional fine-tipped micropipettes.In this paper, several methologies used for culturing neuronal and non-neuronal cells in the laboratory are described. A comparison between the two modes of voltage clamp using blunt-tipped patch-microelectrodes, the switching (discontinuous) and the non-switching (continuous) modes, of the Axoclamp-2A amplifier is made. Some results on membrane currents obtained from neuronal and non-neuronal cells using the single electrode whole-cell tight-seal voltage clamp is illustrated. The possible existence of two inactivating K⁺ currents, one dependent on Ca⁺⁺ the other is not, is discussed.