Mitotic arrest and enhanced nuclear protein phosphorylation in human leukemia K562 cells by okadaic acid, a potent protein phosphatase inhibitor and tumor promoter.

We investigated the effects of the non-phorbol tumor promoter okadaic acid on human leukemia K562 cells. It was found that okadaic acid potently and reversibly inhibited cell growth, with a nearly complete inhibition of thymidine uptake seen at about 10 nM. The cytotoxicity of okadaic acid was characterized by a marked mitotic arrest of the cells exhibiting scattered chromosomes and abnormal anaphase-like structures, a phenomenon distinct from the typical metaphase arrest caused by colchicine. Okadaic acid (10-1,000 nM) greatly stimulated phosphorylation of a number of nuclear proteins in K562 cells. Phosphorylation of many of the same proteins was also stimulated by 12-O-tetradecanoylphorbol-13-O-acetate, a protein kinase C activator. The present findings, consistent with recent reports that okadaic acid is a potent inhibitor of protein phosphatases 1 and 2A (PP1 and PP2A) shown to be essential for normal mitosis, provided evidence for the first time that okadaic acid inhibition of PP1/PP2A resulted in enhanced nuclear protein phosphorylation and subsequent mitotic arrest.     

Tyr60 variants of Flp recombinase generate conformationally altered protein-DNA complexes. Differential activity in full-site and half-site recombinations

The tyrosine at position 60 of the Flp recombinase of the Saccharomyces cerevisiae plasmid, 2 mu circle, is invariant among site-specific recombinases of the "yeast plasmid family". Alterations of this residue give rise to Flp variants that show no recombination activity when assayed in vivo in Escherichia coli. Upon purification, they bind substrate, execute DNA cleavage and catalyze recombination. The efficiency of strand cleavage follows the order: Flp(Y60F) greater than Flp greater than Flp(Y60S) greater than Flp(Y60D); efficiency of recombination between Flp sites on a linear substrate and a circular one follows the order: Flp greater than Flp(Y60F) greater than Flp(Y60S) greater than Flp(Y60D). Methylation footprints of the DNA-protein complexes formed by two of the Flp variants, Flp(Y60S) and Flp(Y60D), do not show hypermethylation of the G residues within the substrate core that is characteristic of complexes formed by wild-type Flp. The third variant, Flp(Y60F), causes significant distortion (although less than wild-type Flp) of the substrate core, as indicated by enhanced G-methylation. Binding profiles with circularly permuted substrates indicate that Flp(Y60S) and Flp(Y60D), but not Flp(Y60F), are defective in bending substrate DNA. In recombination between two Flp half-sites, the variant proteins are significantly more active than in normal full-site recombination.     

蜀龙头骨及脑颅的解剖

本文对蜥脚类蜀龙(Shunosaurus)的脑颅分别从其头骨特征、脑垂体、耳区、视觉系统、脑神经等诸方面进行了较为详细的解剖研究,结果表明 Shunosaurus 内颈动脉不分叉;基蝶骨突强烈收缩;外淋巴囊出现在中耳及 IX、X、XI 孔与颈静脉孔内分外合等现象十分独特.因此,Shunosaurus 应视为一种特化的原始蜥脚类.     

甲壳动物的生殖量与环境关系：Ⅱ.桡足类

本文是《甲壳动物的生殖量与环境关系》系列论文的第二篇,专述桡足类。它和枝角类一样,是一类小型低等甲壳动物,也是浮游生物群落的一个重要组成部分。种类多、数量大、分布广是它的生态特点。由于生活习性不     

人源性流行性出血热病毒的形态特征

本文报道流行性出血热病毒(汉坦病毒)H-114株的电镜形态。发现形态发生以内质网膜和胞浆膜芽生为主。病毒颗粒为圆形或卵圆形。具有双层膜结构,大小为90～120nm。提出了汉坦病毒形态发生的理论观点。     

甲肝病毒在人血白细胞中体外连续传代和在分类白细胞中增殖的研究

1985年我们采用间接免疫荧光法(IF法)检测出甲肝患者外周血白细胞中有甲肝抗原(HAAg)存在,继而又将HAAg阳性白细胞直接种入PLC/PRF/5细胞,分离到两株甲肝病毒(HAV)NJ—3株和H—1株。为了弄清白细胞所携带的病毒究竟仅为吸附吞饮,抑或能在其中复制增殖,我们将分离到的HAV用正常人血白细胞进行体外增殖试验,现将结果报告如下。     

Membrane interactions of mastoparan analogues related to their differential effects on protein kinase C, Na, K-ATPase and HL60 cells.

Membrane interactions of tetradecapeptide toxin mastoparan (MP) and analogues (MP-3, MP-X and polistes MP), as indicated by inhibition of various enzymatic and cellular activities, were investigated. MP-3 was found to be the least active in inhibiting protein kinase C (PKC; activated by phosphatidylserine vesicles, synaptosomal membranes or phorbol ester), synaptosomal membrane Na,K-ATPase and proliferation and viability of leukemia HL60 cells. MP-3, however, was as active as others in inhibiting PKC activated by arachidonate monomers and phorbol ester binding. The unique properties of MP-3, the [des-Ile1-Asn2]-analogue of MP, might be related to its low functional amphiphilicity compared to others and useful in further delineating biological activities associated with or regulated by membranes.     

Aberrant splicing caused by the insertion of the B2 sequence into an intron of the complement C4 gene is the basis for low C4 production in H-2k mice.

The serum level of the fourth component of complement (C4) in mice bearing the H-2k haplotype is only 1/10 to 1/20 of that of non-H-2k mice. We have analyzed C4 cDNA clones from B10.BR(H-2k) mouse liver and found aberrant C4 cDNA which contained a 200-base pair (bp) insertion between the exon 13 and exon 14 encoded sequences in addition to the normal C4 cDNA. The 5' 148 bp and the 3' 52 bp of this insert were derived from the B2 sequence, the short interspersed repeats of mouse genome, and the central part of intron 13, respectively. Sequence analysis of intron 13 of the C4k gene showed the presence of a complete copy of a B2 consensus sequence. The structure of aberrant C4 mRNA indicated that the possible 3' splice site in the B2 sequence and the cryptic 5' splice site in intron 13 were used. Both the insertion of the B2 sequence into intron 13 and the presence of aberrant mRNA in the liver were specific to H-2k-bearing mice, suggesting that the aberrant splicing due to the B2 insertion is the basis for low C4 expression in H-2k mice.     

Strong electrostatic loop-helix interactions in bundle motif protein structures.

Based on CHARMM potential (Brooks et al., 1983) an energetic analysis has been carried out for four typical 4-alpha-helix bundle proteins, i.e., methemerythrin, cytochrome b-562, cytochrome c', and bovine somatotropin. The bovine somatotropin possesses long loops, but all the other three proteins have short loops. It was found that in all these four 4-alpha-helix bundle motif structures the interaction between loops and helices was much stronger than the interaction among the four helices themselves. Particularly for the electrostatic interaction energy, the loop-helix interaction is overwhelmingly stronger than the interhelix interaction although the latter involves the favorable helix dipole interaction due to the antiparallel arrangement of neighboring alpha-helices. The present study indicates that such a conclusion holds true regardless of what loops, long or short, are in the 4-alpha-helix bundle protein, and also regardless of which empirical potential, ECEPP or CHARMM, is used for calculations although in CHARMM the electrostatic energy is much more heavily emphasized than in ECEPP. Therefore, no appropriate conclusion can be drawn in arguing whether the dipole interaction among the four alpha-helices play a stabilizing role or destabilizing role for a 4-alpha-helix bundle protein without taking into consideration the effect of interaction between helices and loops. The calculated results reported here provide, from a different point of view, insights that might be useful for revealing the essence of the driving forces during the folding of proteins.     

The use of ejaculated boar semen after freezing in 2 or 6% glucerol for in vitro fertilization of porcine oocytes matured in vitro

Two concentrations of glycerol in a freezing diluent were tested with respect to the in vitro fertilizing capacity of frozen-thawed boar spermatozoa which, before exposure to oocytes, were subjected to 3 methods of fractionation. These were 1) the upper fraction, 2) the swim-up and 3) percoll gradinet-centrifugation. The highest proportions of motile spermatozoa were obtained by the swim-up procedure, while acrosomal integrity was best preserved by the upper fraction procedure. Raising the glycerol concentration from 2 to 6% (v/v) during freezing decreased the proportion of spermatozoa with a normal apical ridge. Spermatozoa separated by the upper fraction method showed the greatest penetration of oocytes and produced the highest indidence of polyspermy. The glycerol level affected penetration and polyspermy only with spermatozoa separated in a percoll gradient, where the higher level of glycerol increased oocytes penetration and polyspermy. Pronuclei formation was influenced by the separation procedure and by the glycerol concentration in the freezing diluent. The results indicate that frozen boar semen can be used for in vitro fertilization more successfully than fresh semen since penetration by frozen upper fraction spermatozoa was similar to, the degree of polyspermy was lower, and the formation of two pronuclei was greater (P<0.01) than in oocytes exposed to fresh semen.