Phytochrome and Seed Germination. IV. Action of Light Sources With Different Spectral Energy Distribution on the Germination of Tomato Seeds

Germination of tomato seeds exposed to a single, saturating irradiation from light sources of different spectral energy distribution seems to be dependent upon the photostationary P_FR/P_R ratio established by the irradiation. Germination of tomato seeds exposed to prolonged irradiations from the same light sources does not seem to be controlled solely by the P_FR/P_R ratio induced and maintained by the irradiation.     

Random location and absence of movement of the nucleosomes on SV 40 nucleoprotein complex isolated from infected cells.

SV 40 nucleoprotein complexes (NPC) containing viral DNA and cellular histones were extracted from nuclei of permissive cells and treated with either EcoR1 or BamI endonucleases. The fraction of SV 40 linear NPC, monitored by electron microscopy, reached a plateau value of about 15–20% after one hour and no further change occurred during further incubation for 2 hours. Free viral DNA added to the incubation mixture was totally cleaved. During the incubations of NPC and DNA, no redistribution of histones occurred, all the complexe still contained on average 21 nucleosomes and no nucleosomes were generated on the naked viral DNA. Our results suggest a random location and absence of movement of the nucleosomes in vitro on SV 40 nucleoprotein complex isolated from infected cells.     

Appressorium formation and nuclear division in Colletotrichum truncatum

Conidia of the soybean anthracnose fungus, Colletotrichum truncatum differentiate to form appressoria required for host invasion when the germ tube touches a hard surface. This thigmotrophic stimulus appears to be translated by the fungus during the second round of nuclear division. Inhibiting the second round of DNA synthesis by fluorodeoxyuridine or hydroxyurea blocked appearance of appressoria but not emergence of the germ tube. DNA synthesis and mitosis resumed upon removal of FUdR but only mycelia formed, and infection structures did not appear. In addition, actinomycin D reversibly blocked development of appressoria and synthesis of polyadenylate, but nuclear division was not affected. The data suggest that anthracnose conidia produce appressoria in response to germ tube contact by altering the messenger program of its germ tube nucleus. This study has also shown that mitochondrial DNA had an unusual bimodal distribution in CsCl at 1.690 and 1.719 g/cm³, respectively.Non-Standard Abbreviations FUdR 5-fluorodeoxyuridine - polyA polyadenylic acid     

INTERCELLULAR COMMUNICATION AND TISSUE GROWTH : II. Tissue Regeneration

Intercellular communication was examined in regenerating rat liver and urodele skin, two tissues of fast but normal growth. In both, cellular communication is in general as good as in their respective normal intact state. This stands in striking contrast to the lack of cellular communication in tissues with cancerous growth. Upon wounding of the urodele skin, the normally permeable junctional membranes of cells near the wound border seal themselves off, thereby insulating the interiors of the communicated cell systems from the exterior. When the cells of two opposing borders make mechanical contact in the course of wound closure, communication between them ensues within 30 min. Within this period all cell movement also ceases ("contact inhibition"). The possible implications of these findings in the control of tissue growth are discussed.     

ULTRASTRUCTURE AND PERMEABILITY OF NUCLEAR MEMBRANES

The fine structures of nuclear envelopes known to have different permeability properties were compared. Membranes of salivary gland cell nuclei of Drosophila (third instar) and Chironomus (prepupae), which are strong barriers to ion diffusion, and membranes of oocyte nuclei (germinal vesicle) of Xenopus and Triturus, which are much more ion-permeable, show no essential difference in size, frequency, and distribution of their membrane gaps ("pores") which could account for the marked disparities in membrane permeability. The gaps are occupied by diffuse electron-opaque material with occasional central regions of strong opacity. This material may possibly account for the high diffusion resistance of Drosophila and Chironomus nuclear envelopes, where the resistance is far too great to allow free diffusion through the gaps. But material of this kind is also present in the more permeable nuclear envelopes of Xenopus and Triturus oocytes, and there are no convincing structural differences discernible with the techniques employed.     

Sequential assembly of newly synthesized histone on replicating SV40 DNA

Studies on the assembly of histones with newly replicated SV40 DNA show that the four core histones do not associate simultaneously with the DNA. The arginine rich histones H3 and probably H4 associate first, followed by the association of H2a and H2b. Rapid exchange of histone H1 that occurs between cellular and viral chromatins during the extraction hampers studies on the specific association of H1 with newly replicated viral chromatin.     

Site-Directed Mutagenesis of Active Site Residues Reveals Plasticity of Human Butyrylcholinesterase in Substrate and Inhibitor Interactions

Abstract: In search of the molecular mechanisms underlying the broad substrate and inhibitor specificities of butyrylcholinesterase (BuChE), we employed site-directed mutagenesis to modify the catalytic triad residue Ser¹⁹⁸, the acyl pocket Leu²⁸⁶ and adjacent Phe³²⁹ residues, and Met⁴³⁷ and Tyr⁴⁴⁰ located near the choline binding site. Mutant proteins were produced in microinjected Xenopus oocytes, and K_m values towards butyrylthiocholine and IC₅₀ values for the organophosphates diisopropylfluorophosphonate (DFP), diethoxyphosphinylthiocholine iodide (echothiophate), and tetraisopropylpyrophosphoramide (iso-OMPA) were determined. Substitution of Ser¹⁹⁸ by cysteine and Met⁴³⁷ by aspartate nearly abolished activity, and other mutations of Ser¹⁹⁸ completely abolished it. Tyr⁴⁴⁰ and Leu²⁸⁶ mutants remained active, but with higher K_m and IC₅₀ values. Rates of inhibition by DFP were roughly parallel to IC₅₀ values for several Leu²⁸⁶ mutants. Both K_m and IC₅₀ values increased for Leu²⁸⁶ mutants in the order Asp < Gln < Lys. In contrast, cysteine, leucine, and glutamine mutants of Phe³²⁹ displayed unmodified K_m values toward butyrylthiocholine, but up to 10-fold decreased IC₅₀ values for DFP, iso-OMPA, and echothiophate. These findings add Tyr⁴⁴⁰ and Phe³²⁹ to the list of residues interacting with substrate and ligands, demonstrate plasticity in the active site region of BuChE, and foreshadow the design of recombinant BuChEs with tailored scavenging properties.     

Two RAREs and an overlapping CRE are involved in the hepatic transcriptional regulation of the Q10 MHC class I gene

The Q10 gene is a member of the major histocompatibility complex of the mouse that is expressed in the liver and kidney of the adult. Using transient expression assays, we found that the Q10 promoter was activated by retinoic acid (RA) and exogenous RARs and/or RXRs in a cell type-dependent manner. In addition, the basal activity of the Q10 promoter in HepG2 cells is lowered by expressing a dominant negative form of RARalpha. Incidentally, we have identified two cis-elements which consist of sequences related to retinoic acid response elements (RAREs) and a putative cAMP responsive element (CRE) the sequence of which overlaps one of the RAREs. RAR, RXR, CREB-ATF, and COUP-TF factors bind these elements and/or affect their activity. We also demonstrate that the CRE mediates part of the stimulation induced by activation of the cAMP pathway on the Q10 promoter, the residual activation being mediated by RARs. Our results suggest that Q10 expression in liver depends upon RA and the interaction between nuclear receptors that are expressed in this organ. The overlapping of the CRE with one of the RAREs together with the results of PKA activation also suggest that RA and cAMP signalling pathways are linked.     

Components of the Human SWI/SNF Complex Are Enriched in Active Chromatin and Are Associated with the Nuclear Matrix

Biochemical and genetic evidence suggest that the SWI/SNF complex is involved in the remodeling of chromatin during gene activation. We have used antibodies specific against three human subunits of this complex to study its subnuclear localization, as well as its potential association with active chromatin and the nuclear skeleton. Immunofluorescence studies revealed a punctate nuclear labeling pattern that was excluded from the nucleoli and from regions of condensed chromatin. Dual labeling failed to reveal significant colocalization of BRG1 or hBRM proteins with RNA polymerase II or with nuclear speckles involved in splicing. Chromatin fractionation experiments showed that both soluble and insoluble active chromatin are enriched in the hSWI/SNF proteins as compared with bulk chromatin. hSWI/SNF proteins were also found to be associated with the nuclear matrix or nuclear scaffold, suggesting that a fraction of the hSWI/SNF complex could be involved in the chromatin organization properties associated with matrix attachment regions.