Effect of hepatocyte growth factor/scatter factor and other growth factors on motility and morphology of non-tumorigenic and tumor cells

Summary Using an automated cell analyzer system, the effect of hepatocyte growth factor/scatter factor (HGF/SF), epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), endothelial acidic fibroblast growth factor (a-FGF), platelet derived growth factor (PDGF), and recombinant human insulinlike growth factor (IGF) on the motility and morphology of Madin-Darby canine kidney (MDCK), rat hepatomas, C₂, and H_5–6 and murine mammary carcinoma (EMT-6) cells was investigated. Treatment of MDCK cells with HGF/SF, bFGF, EGF, and a-FGF resulted in an increase in average cell velocity and in the fraction of moving cells. Cells treated with the PDGF and IGF did not show significant alterations in velocity. MDCK cells treated with each growth factor were classified into groups of “fast” and “slow” moving cells based on their average velocities, and the average morphologic features of the two groups were quantitated. Fast-moving cells had larger average area, circularity, and flatness as compared to slow-moving cells. Factors that stimulated cell movement also induced alterations in cell morphologic parameters including spreading, flatness, area, and circularity. HGF/SF also scattered and stimulated motility of C₂ and H_5–6 hepatoma cells. In contrast to MDCK cells, there was no significant difference between the morphology of the fast moving and slow moving C₂ and H_5–6 cells. These studies suggest that growth factor cytokines have specific effects on motility of normal and tumor cells.     

Thermal denaturation and loss of viability in Escherichia coli and Bacillus stearothermophilus

When Escherichia coli was heated at 10°C/min in a differential scanning calorimeter, the onset of irreversible thermal denaturation occurred at 51°C, about 5°C above the maximum growth temperature. The temperature at which death rate was maximal (63°C) coincided with the thermogram peak caused by denaturation of the 30S ribosomal subunit. The maximum death rate in vegetative cells of Bacillus stearothermophilus occurred at the higher temperature of 71°C which also coincided with the leading edge of the main thermogram peak.     

谷类作物β-葡聚糖合成酶基因家族的研究进展

Beta-葡聚糖是由β-(1,3)和β-(1,4)糖苷键连接的非纤维素多糖,主要分布在谷类作物籽粒胚乳及糊粉层中,在高尔基体合成,经由囊泡运输到质膜,最终在细胞壁上沉积。通过增加胆汁酸排泄,延迟葡萄糖吸收,β-葡聚糖可有效降低胆固醇及血糖水平。Beta-葡聚糖合成酶基因家族成员最早在水稻(Oryza sativa)中得到鉴定,后在其他作物中陆续被发现。该基因家族包括3个主要成员：CslF、CslH和CslJ亚基因家族,起源于不同分支,经过趋同演化,执行合成β-葡聚糖的功能。Beta-葡聚糖基因家族成员均受到负选择压力,演化过程中序列高度保守。CslF亚家族基因成员相对较多,常在染色体上形成基因簇,CslF6是介导β-葡聚糖合成的主效基因。CslF亚家族在叶基部等幼嫩组织中表达水平相对较高,且明显受到光照强度的影响;CslH和CslJ亚家族成员较少,其中CslH亚家族在叶尖等成熟组织中的表达水平高,而CslJ亚家族在籽粒中有较高的表达水平。该文综述了β-葡聚糖合成酶基因家族成员的系统发育关系、表达模式,β-葡聚糖合成酶的亚细胞定位,以及作物中的定向育种研究进展,提出β-葡聚糖合成酶基因家...     

Some results in graph theory and their application to abstract relational biology

It has been shown in earlier work that one approach to what Rashevsky has called “abstract biology” is through the study of the class of ( )-systems that can be formed in an arbitrary subcategory of the category of sets. The concept of the ( )-system, however, depends on the availability of mappings that contain other mappings in their range. It is shown that, by introducing an appropriate measure for this property, the problem of characterizing those categories suitable for a rich theory of ( )-systems reduces to a problem familiar from the general theory of graphs. Some new results in these directions are obtained, and it is then shown that any category with mappings that possess properties we might expect to hold in the physical world will also admit a rich theory of ( )-systems. In particular, it is shown that a sufficiently large family of mappings drawn at random from such a category will with overwhelming probability contain an ( )-system. This research was supported by the United States Air Force through the Air Force Office of Scientific Research of the Air Research and Development Command, under Grant No. AF-AFOSR-9-63.     

Tomato exo-(1-->4)-beta-D-galactanase. Isolation, changes during ripening in normal and mutant tomato fruit, and characterization of a related cDNA clone.

An exo-(1-->4)-beta-D-galactanase was isolated from ripe tomato fruit (Lycopersicon esculentum Mill. cv Ailsa Craig and cv Better Boy) using anion-exchange, gel filtration, and cation-exchange chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the most active fraction revealed a predominant protein band at 75 kD and several minor bands. A 30-amino acid N-terminal sequence from this 75-kD protein showed a high degree of homology with other recently identified beta-galactosidase/ galactanase proteins from persimmon and apple fruits (I.-K. Kang, S.-G. Suh, K.C. Gross, J.-K. Byun [1994] Plant Physiol 105: 975-979; G.S. Ross, T. Wegrzyn, E.A. MacRae, R.J. Redgwell [1994] Plant Physiol 106: 521-528) and with the predicted polypeptide sequence encoded by the ethylene-regulated SR12 gene in carnation (K.G. Raghothama, K.A. Lawton, P.B. Goldsbrough, W.R. Woodson [1991] Plant Mol Biol 17: 61-71). The enzyme focused to a single band of beta-galactosidase activity on an isoelectrofocusing gel at pH 9.8. The enzyme was specific for (1-->4)-beta-D-galactan substrates with a pH optimum of 4.5. The only reaction product detected was monomeric galactose, indicating that the enzyme was an exo (1-->4)-beta-D-galactanase. beta-Galactanase activity increased at the onset of ripening in normal fruit, but no similar increase was detected in the nonripening mutants nor and rin. A tomato homolog (pTombetagal1) was isolated using the SR12 cDNA clone from carnation as a probe. This clone showed 73% identify at the amino acid level with beta-galactosidase-related sequences from apple and asparagus and 66% identity with SR12. pTombetagal1 is a member of a gene family. Northern analysis demonstrated that pTombetagal1 expression was ripening related in normal fruits, with lower levels apparent in the nonsoftening mutants.     

Dual mode of energy coupling by the oxyanion-translocating ArsB protein.

The arsA and arsB genes of the ars operon of R-factor R773 confer arsenite resistance in Escherichia coli by coding for an anion-translocating ATPase. Arsenite resistance and the in vivo energetics of arsenite transport were compared in cells expressing the arsA and arsB genes and those expressing just the arsB gene. Cells expressing the arsB gene exhibited intermediate arsenite resistance compared with cells expressing both the arsA and arsB genes. Both types of cells exhibited energy-dependent arsenite exclusion. Exclusion of 73AsO2- from cells expressing only the arsB gene was coupled to electrochemical energy, while in cells expressing both genes, transport was coupled to chemical energy, most likely ATP. These results suggest that the Ars anion transport system can be either an obligatory ATP-coupled primary pump or a secondary carrier coupled to the proton motive force, depending on the subunit composition of the transport complex.     

The ars operon of Escherichia coli confers arsenical and antimonial resistance.

The chromosomally encoded arsenical resistance (ars) operon subcloned into a multicopy plasmid was found to confer a moderate level of resistance to arsenite and antimonite in Escherichia coli. When the operon was deleted from the chromosome, the cells exhibited hypersensitivity to arsenite, antimonite, and arsenate. Expression of the ars genes was inducible by arsenite. By Southern hybridization, the operon was found in all strains of E. coli examined but not in Salmonella typhimurium, Pseudomonas aeruginosa, or Bacillus subtilis.     

Independent changes in type I and type II receptors for transforming growth factor beta induced by bone morphogenetic protein 2 parallel expression of the osteoblast phenotype.

Transforming growth factor beta (TGF-beta), a potent regulator of bone formation, has bifunctional effects on osteoblast replication and biochemical activity that appear differentiation dependent. We now show that cell surface binding sites for TGF-beta vary markedly among fibroblasts, bone-derived cells, and highly differentiated osteosarcoma cultures from fetal rats. Expression of betaglycan and type II receptors decline relative to type I receptor expression in parallel with an increase in osteoblast-like activity, predicting that the ratio among various TGF-beta binding sites could influence how its signals are perceived. Bone morphogenetic protein 2 (BMP-2), which induces osteoblast function, does not alter TGF-beta binding or biochemical activity in fibroblasts and has only small effects in less differentiated bone cells. In contrast, BMP-2 rapidly reduces TGF-beta binding to betaglycan and type II receptors in osteoblast-enriched primary cell cultures and increases its relative binding to type I receptors in these cells and in ROS 17/2.8 cultures. Pretreatment with BMP-2 diminishes TGF-beta-induced DNA synthesis in osteoblast-enriched cultures but synergistically enhances its stimulatory effects on either collagen synthesis or alkaline phosphatase activity, depending on the present state of bone cell differentiation. Therefore, BMP-2 shifts the TGF-beta binding profile on bone cells in ways that are consistent with progressive expression of osteoblast phenotype, and these changes distinguish the biochemical effects mediated by each receptor. Our observations indicate specific stepwise actions by TGF-beta family members during osteoblast differentiation, developing in part from changes imprinted by BMP-2 on TGF-beta receptor stoichiometry.     

Secretion of unprocessed human surfactant protein B in milk of transgenic mice

Because of the apparent clinical importance of human pulmonary surfactant B (SP-B), the expression of SP-B was directed to the mammary gland of transgenic mice using previously characterized rat whey acidic protein (WAP) regulatory sequences. rWAP/SP-B mRNA was expressed specifically in the mammary gland, and ranged from 1 to 5% of the endogenous WAP mRNA levels. SP-B was detected immunologically in both tissue and milk. The transgene product had an apparent molecular weight of 40--45 kDa, corresponding to the predicted size of the SP-B proprotein. Incubation of an SP-B-enriched fraction of milk with cathepsin D in vitro produced 20--25 kDa species, consistent with cleavage of the amino terminal domain by cathepsin D. This was confirmed using antibodies specific to the carboxy-terminal domain of SP-B. However, the appearance of only the SP-B proprotein in milk suggests that cathepsin D is not involved in the in vivo processing of SP-B. The SP-B proprotein can be expressed in milk of transgenic mice without any observed effects on mammary gland morphology or lactation