Heterochromatin differentiation and phylogenetic relationship of the A genomes in diploid and polyploid wheats

Summary Heterochromatin differentiation, including band size, sites, and Giemsa staining intensity, was analyzed by the HKG (HCl-KOH-Giemsa) banding technique in the A genomes of 21 diploid (Triticum urartu, T. boeoticum and T. monococcum), 13 tetraploid (T. araraticum, T. timopheevi, T. dicoccoides and T. turgidum var. Dicoccon, Polonicum), and 7 cultivars of hexaploid (T. aestivum) wheats from different germplasm collections. Among wild and cultivated diploid taxa, heterochromatin was located mainly at centromeric regions, but the size and staining intensity were distinct and some accessions' genomes had interstitial and telomeric bands. Among wild and cultivated polyploid wheats, heterochromatin exhibited bifurcated differentiation. Heterochromatinization occurred in chromosomes 4A^t and 7A^t and in smaller amounts in 2A^t, 3A^t, 5A^t, and 6A^t within the genomes of the tetraploid Timopheevi group (T. araraticum, and T. timopheevi) and vice versa within those of the Emmer group (T. dicoccoides and T. turgidum). Similar divergence patterns occurred among chromosome 4A^a and 7A^a of cultivars of hexaploid wheat (T. aestivum). These dynamic processes could be related to geographic distribution and to natural and artifical selection. Comparison of the A genomes of diploid wheats with those of polyploid wheats shows that the A genomes in existing diploid wheats could not be the direct donors of those in polyploid wheats, but that the extant taxa of diploids and polyploids probably have a common origin and share a common A-genomelike ancestor.Contribution of the College of Agricultural Sciences, Texas Tech Univ. Journal No. T-4-233.     

Activation of the human epsilon- and beta-globin promoters by SV40 T antigen.

We have studied the effect of the SV40 T antigen on expression from human globin promoters fused to the bacterial chloramphenicol acetyltransferase (CAT) gene and compared its effect with the SV40 enhancer and the adenovirus E1A protein. We have observed that expression of p epsilon GLCAT and p beta GLCAT (the epsilon-globin or beta-globin promoter linked to the CAT gene) was significantly stimulated when cotransfected with a cloned T antigen plasmid into CV-1 cells, indicating that trans-activation of the globin promoters was mediated by SV40 T antigen. Transfection of the p beta GLCAT-SV (p beta GLCAT containing the SV40 enhancer element) into CV-1 cells resulted in a 50-60-fold increase in CAT activity as compared to p beta GLCAT (no enhancer). However, cotransfection of the p beta GLCAT-SV with the cloned T antigen resulted in an additional increase of CAT expression, which suggests that T antigen and the SV40 enhancer activate globin gene expression independently. We found that T antigen but not E1A could further stimulate the expression of an enhancer-containing plasmid in CV-1 cells; whereas E1A but not T antigen could further stimulate p epsilon GLCAT expression in COS-1 cells which constitutively express the SV40 T antigen. These results suggest that T antigen and E1A also act independently. Deletion analysis showed that the minimum sequence required for a detectable level of stimulation of the epsilon-globin promoter by T antigen is 177 bp 5' to the cap site, suggesting that the target sequences for response to T antigen do not reside in the canonical 100 bp promoter region, but rather reside in sequences further upstream, and therefore the cellular factors interacting with T antigen are not the TATA or CAT box binding proteins, but the proteins interacting with upstream regulatory sequences.     

Identification of the ATP + Mg-dependent and polycation-stimulated protein phosphatases in the germinal vesicle of the Xenopus oocyte.

Two protein phosphatase activities were characterized in the germinal vesicle of Xenopus laevis oocytes after manual dissection of the nucleus. One enzyme can be classified as an active form of the ATP + Mg-dependent (AMD) phosphatase, the other as a polycation-stimulated (PCS) phosphatase. The activity of the PCS phosphatase is localized exclusively in the soluble compartment of the nucleus (nucleoplasm). The catalytic subunit of the AMD phosphatase activity is associated either with the nuclear particulate fraction or with an inhibitory subunit in the nucleoplasm.     

Microcalorimetric measurements of tissue cells in vitro

A culture flask was designed for the microcalorimetric measurements of tissue cells by an MS 80 standard calvet microcalorimeter. Tissue cells cultured in this flask behaved in the same manner as in the common culture flask used in cytobiological studies. The thermograms of human adenocarcinoma gastric cells (SGc 7901) and HeLa cells were obtained. The heat output power of SGc 7901 cells continuously increased for 70 h with an initial cell number of 3.0 X 10(5). The thermogram was reproducible under strictly controlled conditions. The relationship between the heat output power and the number of SGc 7901 cells within 48 h was obtained. The heat output power was 40 pW/cell to 49 pW/cell when the cell number was in the range 4.5 X 10(5) to 10.4 X 10(5). It was 62.3 +/- 2.9 pW/cell for HeLa cells when the cell number was 6 X 10(5).     

Scavenging effect of extracts of green tea and natural antioxidants on active oxygen radicals

With the use of the spin trapping methods, the scavenging effects of the extracts of green tea and other natural foods are studied. In stimulated polymorphonuclear leukocytes (PMN) system, water extract fraction 6 (F6) from green tea and green tea polyphenols (GTP) have the strongest scavenging effect on the active oxygen radicals, much stronger than vitamin C (Vc) and vitamin E (VE). Rosemary antioxidants (RA) and Curcumin (Cur) have weaker scavenging effects than Vc, but stronger than VE. In Fenton Reaction, Cur has the strongest scavenging effect (69%) on hydroxyl radicals. In irradiation, riboflavin system F6(74%) and GTP(72%) have very strong scavenging effects that are weaker than Vc, but much stronger than VE (23%). With the use of spin probe oxymetry, the oxygen consumption in respiratory burst of stimulated PMN were measured when the antioxidants existed in these systems. The results demonstrated that these antioxidants did not affect the respiratory burst of human polymorphonuclear leukocytes stimulated with PMA.     

Calcitonin gene-related peptide (CGRP) in normotensive and spontaneously hypertensive rats

With the techniques of specific radioimmunoassay and gel filtration it was found that CGRP was distributed in various tissues of normotensive (WKY) and spontaneously hypertensive rats (SHR) with the highest concentration in the lumbar spinal cord (1197 +/- 94.8 pg/mg tissue) and the lowest in the auricle (15.0 +/- 2.1 pg/mg tissue). In comparison with WKY, CGRP concentration in the plasma was decreased and in the abdominal aorta and hypothalamus was increased in SHR. Gel filtration revealed only one major CGRP molecular form in the tissues. In addition, CGRP reduced the mean arterial pressure (MAP) in SHR in a dose-dependent manner. These data suggest that CGRP may play an important role in the pathogenesis of hypertension and its possible therapy.     

A simple and inexpensive method for cellulase and β-glucosidase production by Neurospora crassa

Summary Elevated levels of cellobiohydrolase, carboxymethylcellulase (CMCase) and -glucosidase were produced by strain STA of Neurospora crassa, grown in solid-state fermentation on untreated wheat straw supplemented with simple mineral salts. Yields as high as 6.1 units of cellobiohydrolase, 969.2 units of CMCase and 169.4 units of -glucosidase per gram of straw were obtained at optimum growth and enzyme assay conditions.     

Solid-phase synthesis of oligoribonucleotides using 9-fluorenylmethoxycarbonyl (Fmoc) for 5''-hydroxyl protection.

Efficient solid-phase synthesis of a series of oligoribonucleotides of up to 20 residues is described that utilises the 9-fluorenylmethoxycarbonyl group (Fmoc) for 5'-protection and 4-methoxytetrahydropyran-4-yl (Mthp) for 2'-protection of ribonucleotide monomers and a phosphoramidite coupling procedure. The Fmoc group is removed after each coupling step by treatment with 0.1M DBU in acetonitrile. Oligoribonucleotides are isolated in 2'-protected form in good yield and shown to be readily and efficiently deprotected by mild acidic treatment.     

Modelling basic features of specificity in the binding of a dicationic steroid diamine to double-stranded oligonucleotides.

An investigation of the intrinsically preferred binding modes of a steroid diamine, dipyrandium, to the double-stranded hexanucleotides d(TATATA)2, d(ATATAT)2, and d(CGCGCG)2 is carried out by the energy minimization procedure JUMNA. Several alternative binding modes are compared: groove binding in which the conformation of the oligonucleotide remains close to that of B-DNA, intercalation between base-pairs and interaction with variously kinked structures in which base pairs of dinucleoside steps open towards the groove in which the binding occurs. The favored binding configuration occurs at the d(TpA) step of the AT kinked nucleotides in which the kink opens the base pairs towards the minor groove. Thus, for the d(T1A2T3A4T5A6)2 sequences the preferred complexation involves the kink at the T3A4 step facing the cyclohexane rings A, B, and C of the ligand. For the d(A1T2A3T4A5T6)2 sequence, the kink occurs at the T2A3 step facing the cationic pyrrolidine ring linked to ring A. The binding of dipyrandium to d(CGCGCG)2 is found to be considerably less favourable than for either of the two (AT) sequences.