Mechanisms of heat protection by cycloheximide or puromycin: Involvement of polyamines?

1. 1.|When Chinese hamster ovary cells are treated with cycloheximide (10 μg/ml) or puromycin (100 μg/ml) for 2 h before and during heating at 43°C for 3 h, there is protection from hyperthermic killing; i.e. the plating efficiency increases 2000-fold from 3.7 × 10⁻⁵ to (6–9) × 10⁻².

2. 2.|The total intracellular levels of spermidine and spermine are not altered by the hyperthermic or drug treatments.

3. 3.|The small 30% decrease in intracellular putrescine observed after heating is not altered by drug treatment.

4. 4.|Heat protection by treatment with cycloheximide or puromycin cannot be attributed to changes in levels of total intracellular polyamines.

Nuclease hypersensitivity in the beta-globin gene region of K562 cells

We have investigated chromatin structure in the beta-globin gene region of the K562 human erythroleukemic cell line by using S1 and DNase I nuclease sensitivity assays. Despite the lack of beta-globin gene expression in these cells, we find nuclease-hypersensitive sites to these enzymes in its 5' and 3' flanking regions in K562 chromatin. This result is in contrast to previous reports in which no hypersensitive sites were found in the immediate vicinity of this gene. In the 3' region, one major hypersensitive site at 0.9 kpb 3' and three minor hypersensitive sites at 0.7 kbp, 0.5 kbp 3' and 0.2 kbp 5' of the polyadenylation site were observed; these sites are very similar to those found in fetal liver and adult bone marrow cells in which the beta-globin gene is expressed. We find hypersensitive sites to both enzymes in the 5' region of the beta-globin gene: a major site 0.8 kbp 5' to the cap site, and two minor sites 1.2 and 1.5 kbp 5' to the cap site. The -0.8 kbp site is also present in plasmids containing the beta-globin gene. Our results suggest that the lack of beta-globin gene expression may be related to the lack of hypersensitivity sites in the immediate (150 bp) 5' flanking region of the beta-globin gene, as occurs in other active globin genes.     

Cell-surface expression of a membrane-anchored form of the human chorionic gonadotropin alpha subunit

We carried out experiments designed to generate a novel cell-surface protein from a small glycosylated secretory protein. DNA encoding the entire precursor of human chorionic gonadotropin (hCG, alpha subunit) was fused precisely to DNA encoding the transmembrane and cytoplasmic domains of the vesicular stomatitis virus glycoprotein. When expressed in animal cells this DNA encoded the 92-amino acid hCG-alpha subunit anchored in cellular membranes by an extension composed of the 49 carboxyl-terminal amino acids of vesicular stomatitis virus glycoprotein. This hybrid protein was transported efficiently to the plasma membrane of animal cells. The two asparagine-linked glycans on the anchored form of hCG-alpha were large and heterogeneous when compared to those on the secretory form. Experiments employing in vitro mutagenesis and the glycosylation inhibitor tunicamycin established that the presence of at least one of the two asparagine-linked glycans was required for expression of the anchored molecule on the cell surface. However, as reported previously, secretion of hCG-alpha occurred in the absence of glycosylation. Also, mutations eliminating the second glycosylation site (at amino acid 78) in both the anchored or secreted forms apparently led to partial denaturation or a conformational change interfering with transport of the protein.     

Some observations on the localization of mitomycin C-induced aberrations in human lymphocytes

Actively cycling human lymphocytes were treated with mitomycin C for 1 h (1.4 μg/ml) and then grown in medium containing 10 μg/ml bromodeoxyuridine. Serial 5-h colcemid accumulation samples were taken up to 35 h and the air-dried methaphase spreads stained for replication banding. A complete cell-cycle subphasing analysis was made, and classified cells scored for all categories of chromatid-type aberrations and their location.

Inspite of the high dose which produced massive delay and cycle perturbation, there was no evidence for selectively lethality of early-S cells, in fact such cells were in excess. Extreme localization of aberrations to late-replicating (mostly centromeric) regions was found at all subphases and in pre-S cells. This rules out ‘localization by default’ as an explanation for the observed preferential occurence of ‘break points’ in these regions.

The frequency of incomplete intrachanges, low in late S, rises dramaticallyin early S to become maximal in pre-S cells.     

Stimulation of proliferation ofTetrahymena pyriformis by trace rare earths

Using two Chinese strains ofTetrahymena pyriformis, S1 and BJ4, as the biological models, the effects of lighter rare earths (lanthanum, cerium, praseodymium, neodymium, samarium, and europium), representatives of heavier rare earths (yttrium and thulium), and mixed rare earths were studied. The stimulation of population growth ofTetrahymena in peptone-glucose media containing trace amounts of these elements have been observed. The mechanisms for the beneficial effects of rare earth elements in low concentrations remains to be discovered.     

肝素修饰尿激酶某些性质的研究

本文对比研究了溴化氰活化及高碘酸活化肝素修饰的两种修饰尿激酶的性质。结果表明尿激酶在溴化氰活化肝素(肝素CN),高碘酸钠活化肝素(肝素I_4)的共价修饰后,其残余自由氨基分别是64％和52％;酶活性分别保留94％和90％;抗胃蛋白酶水解以及抗冻融变性的能力均高于天然酶;在离体血浆中的失活速变低于天然酶。本文还对修饰酶进行了萤光及紫外差光谱的分析,讨论了修饰过程对构象的影响。     

四川成都市郊正常成年男性血清生殖激素水平及其频数分布

本文用放射免疫测定法(RIA),测定了200～290例中国四川成都市郊30～73岁的正常成年男性血清睾酮(T)促黄体激素(LH).卵泡刺激素(FSH)及泌乳激素(PRL)的水平及其分布,进行了不同年龄水平差异显著性的观察,见到睾酮与泌乳激素水平无显著差异,促黄体激素及卵泡刺激素水平,随年龄之增加而升高,差异有显著性,并测得它们在血清中水平的分布:LH、FSH及PRL均为对数正态公布,T既非正态又非对数正态,但其频率分布较对称,接近正态分布。     

Biological activity in vivo of insulin analogues modified in the N-terminal region of the B-chain

[AlaB5]Insulin as well as a hybrid analogue of insulin and "insulin-like growth factor" (IGF-I), in which the N-terminal amino-acid sequence H-Phe-Val-Asn-Gln- of the B-chain has been replaced by the tripeptide H-Gly-Pro-Glu-of IGF-I, have been prepared by the partial-synthetic route. Their biological activity in vivo has been compared with that of other analogues in rabbits, mice and rats as far as data are available. These rodents respond differently, rats being less sensitive to modifications than rabbits and mice. The results explain unexpected discrepancies discussed in previous papers.