Resistance ofSpodoptera frugiperda [Lep.: Noctuidae] to a nuclear polyhedrosis virus in the field and laboratory

Median lethal doses (LD₅₀s) of nuclear polyhedrosis virus (NPV) were determined in neonatal offspring ofSpodoptera frugiperda (J. E. Smith) (Sf) larvae captured in southeastern Louisiana in 1981, 1982, and 1984. These LD₅₀s ranged from 1.8 to 16.3 polyhedral inclusion bodies (PIB)/insect. The LD₅₀s significantly (P<0.05) increased during the season of 1982 but had no pattern in 1981 or 1984. However, the Sf populations increased in heterogeneity of response to the NPV during all 3 years. The LD₅₀ increased from 4.1 to 18.7 PIB/insect in a Sf laboratory colony exposed to the NPV LD₈₀ for 7 generations, whereas in a control colony not exposed to NPV the LD₅₀ was 5.9 PIB/insect after 7 generations.     

Autecological investigations on marine diatoms. 5:Coscinodiscus concinnus W. Smith andRhizosolenia setigera Brightwell

The cold oligo-eurytherm diatomsCoscinodiscus concinnus W. Smith andRhizosolenia setigera Brightwell were cultured to determine their best competitive position by growth. Comparison of their generation times with those of other diatoms indicate that they reach this position between 6°C and 12°C. Both species grew between –1.5°C and about 20°C. The experiments indicate thatC. concinnus flowerings are possible in a deep water column, during periods of high light intensities. The simultaneous death of species in the upper layer is also caused by high light intensities.C. concinnus appeared in two morphological forms; the normal voluminous form, and a flatter form with a few intercalary bands only, filled with large oil-droplets. The latter appeared at 0°C and below, and at the upper temperature limit for growth of about 19°C–20°C. The separation of nov. spec. fromC. concinnus based on the absence or presence of a hyaline area and intercalary bands as identification characteristics should be reconsidered.     

Structure of the Bombyx mori fibroin light-chain-encoding gene: upstream sequence elements common to the light and heavy chain.

Two overlapping genomic clones containing the fibroin light-chain (Fib-L)-encoding gene (Fib-L) were obtained from the cosmid library of the silkworm, Bombyx mori J-139, by hybridization with the Fib-L cDNA clone. Sequencing of the 14.6-kb region revealed that Fib-L was 13472 bp long containing seven exons, and that the gene contained a large first intron which occupied about 60% of the gene. Comparison of restriction patterns of the J-139 Fib-L with those of eight other B. mori breeds producing normal-level fibroin demonstrated that considerable restriction-fragment length polymorphisms were present in regions containing the first intron and the 3′-flanking sequence. However, sizes of the Fib-L mRNA and the Fib-L polypeptide were very similar among the nine breeds tested, suggesting that the exon sequences and the splice signals were all well conserved. 5′-Flanking regions of Fib-L and the fibroin heavy-chain (Fib-H)-encoding gene (Fib-H) compared in this study contained three 18-30-bp sequences of high similarity and many 8-10-bp common elements, six of which coincided with the binding sites of homeodomain proteins. Gel retardation assays with the nuclear extracts of the posterior and middle silk glands suggested that protein factors present in the posterior silk-gland nuclei could bind to a set of those common upstream elements.     

The effect of age on the response of the detrusor to intracellular mechanical stimulus: DNA replication and the cell actin matrix.

Benign prostatic hypertrophy and posterior urethral valves present at both extremes of the age spectrum. Both disease processes can obstruct the urinary stream and ultimately have pathophysiological effects on detrusor structure and function. The mechanisms regulating the structural reorganization of the detrusor to a mechanical outflow obstruction are not known. In an attempt to identify maturational differences in myocyte ultrastructure and consequent effects these might have in modifying the response of the detrusor to mechanical stimulus, we studied differences in dynamic nuclear-cytoskeletal interactions in detrusor tissue in an animal model. Using a drug which specifically severs actin, cytochalasin D (CD), as an intracellular mechanical stimulus, we measured changes in nuclear area and the rate of DNA synthesis in detrusor myocytes from young (2-3 week) and old (8-12 mon) guinea pigs. We found that there were age specific differences to intracellular mechanical stimuli in detrusor muscle. Nuclei of myocytes from young animals showed elastic recoil on severing the cell actin matrix and the tissue from young animals increased replicative DNA synthesis with an intracellular stimulus. In contrast, nuclear shape changes in myocytes from old animals suggested less elasticity, and there was no increase in DNA synthesis with disruption of the cell actin matrix. Anti-alpha-smooth muscle actin antibody and rhodamine phalloidin staining of actin in cytochalasin D treated primary explants of detrusor myocytes showed dose dependent disruption of the actin component of the cytoskeleton. These results suggest that there are fundamental modifications in detrusor myocyte ultrastructure with age. These maturational changes might result in differences in the pathophysiological and structural reorganization of the detrusor in response to outflow obstruction in infancy and adulthood. Furthermore, they suggest that 1) a tensile equilibrium exists between the myocyte nucleus and cytoskeleton; 2) there appears to be a decrease in myocyte nuclear elasticity with ageing; 3) release of nuclear template restrictions increases activity of DNA polymerase alpha in young, but not old, detrusor myocytes; and 4) mechanico-chemical signal transduction in detrusor myocytes may be mediated via the cytoskeleton. In addition, based on previous reports of actin within the nucleus, the results suggest that 1) nuclear actin may have a homeostatic structural role, maintaining the tensile equilibrium between nucleus and cytoskeleton, and 2) integrity of nuclear actin may function to maintain the spatial template restriction on DNA polymerase alpha activity.     

Interaction of MAR-sequences with nuclear matrix proteins.

The recent discovery of DNA sequences responsible for the specific attachment of chromosomal DNA to the nuclear skeleton (MARs/SARs) was an important step towards our understanding of the functional and structural organization of eukaryotic chromatin [Mirkovitch et al.: Cell 44:273-282, 1984; Cockerill and Garrard: Cell 44:273-282, 1986]. A most important question, however, remains the nature of the matrix proteins involved in the specific binding of the MARs. It has been shown that topoisomerase II and histone H1 were capable of a specific interaction with SARs by the formation of precipitable complexes [Adachi et al.: EMBO J8:3997-4006, 1989; Izaurralde et al.: J Mol Biol 210:573-585, 1989]. Here, applying a different approach, we were able to "visualize" some of the skeletal proteins recognizing and specifically binding MAR-sequences. It is shown that the major matrix proteins are practically the same in both salt- and LIS-extracted matrices. However, the relative MAR-binding activity of the individual protein components may be different, depending on the method of matrix preparation. The immunological approach applied here allowed us to identify some of the individual MAR-binding matrix proteins. Histone H1 and nuclear actin are shown to be not only important components of the matrix, but to be involved in a highly efficient interaction with MAR-sequences as well. Evidence is presented that proteins recognized by the anti-HMG antibodies also participate in MAR-interactions.     

Highly asymmetric intergeneric nuclear hybrids between Nicotiana and Petunia: evidence for recombinogenic and translocation events in somatic hybrid plants after “gamma”-fusion

Summary Extremely asymmetric nuclear hybrids have been obtained via protoplast fusion in an intergeneric combination. Irradiated (cobalt⁶⁰,100 krad) kanamycinresistant Petunia hybrida mesophyll protoplasts were chemically fused with wild-type mesophyll protoplasts of Nicotiana plumbaginifolia. Eighty-six hybrid colonies were selected on kanamycin-containing medium, and twenty-four of these could be induced to regenerate numerous shoots. Cytological analysis of the regenerants showed the presence of a few chromosome fragments in some lines, and even a metacentric chromosome in yet another line. Besides additional chromosome fragments some lines only possessed typical Nicotiana chromosomes, and this at the diploid (2n = 2X = 20) as well as the tetraploid (2n = 2X = 40) level. Biochemical analysis showed that all regenerants had neomycin phosphotransferase activity (NPTII), which suggests that intergenomic recombination and or translocation events took place at least in those lines where no additional chromosome fragments could be detected. The presence of the NPTII gene was shown by Southern hybridization. All regenerants tested were fertile, and the segregation ratios for the kanamycin gene (for self and backcross pollinations to the recipient partner) for some of the regenerants correspond with Mendelian rules for a monogenic dominant marker. Most of the regenerants showed abnormal segregation ratios; in this case, no correlation could be made between segregation ratio and chromosome composition.Our results demonstrate the existence of intergenomic recombination and translocations evens in nuclear somatic hybrid plants obtained via gamma-fusion.     

Targeting of T7 RNA polymerase to tobacco nuclei mediated by an SV40 nuclear location signal

We have expressed two T7 RNA polymerase genes by electroporation into tobacco protoplasts. One of the genes was modified by inserting nucleotides encoding a viral nuclear localization signal (NLS) from the large T antigen of SV40. Both T7 RNA polymerase genes directed synthesis of a ca. 100 kDa protein in the electroporated protoplasts. T7 RNA polymerase activity was detected in extracts of protoplasts electroporated with both genes. Immunofluorescence analysis of these protoplasts indicated that only the polymerase carrying the NLS accumulated in the cell nucleus. These experiments suggest that mechanisms involved in the transport from the cytoplasm to the nucleus are similar in plant and animal cells. This system demonstrates the feasibility of T7 RNA polymerase-based approaches for the high-level expression of introduced genes in plant cells.     

Nuclear genome characterization of the carrageenophyteAgardhiella subulata (Rhodophyta)

DNA reassociation kinetics were used to determine nuclear genome organization and complexity inAgardhiella subulata (Gigartinales, Rhodophyta). Results indicate the presence of three second-order components corresponding to fast (22%), intermediate (68%) and slow (10%) fractions. Thus, the genome consists of 90% repetitive sequences. Microspectrophotoometry with the DNA-localizing fluorochrome DAPI was used to confirm ploidy level differences in the gametophytic and tetrasporophytic phases. Results indicate that meiosis occurs during tetrasporogenesis. Comparison of mean nuclear DNA (I_f) values to chicken erythrocytes (RBC) resulted in an estimate of 0.9 pg/2C genome forAgardhiella. Karyological studies using aceto-orcein revealed a chromosome complement of 2N = 44 in carposporangia and the presence of 22 bivalents during diakinesis of tetraspore mother cells.