Physical and Functional Interactions between the Herpes Simplex Virus UL15 and UL28 DNA Cleavage and Packaging Proteins

Herpes simplex virus (HSV) DNA is cleaved from concatemers and packaged into capsids in infected cell nuclei. This process requires seven viral proteins, including UL15 and UL28. UL15 expressed alone displays a nuclear localization, while UL28 remains cytoplasmic. Coexpression with UL15 enables UL28 to enter nuclei, suggesting an interaction between the two proteins. Additionally, UL28 copurified with UL15 from HSV-infected cells after ion-exchange and DNA affinity chromatography, and the complex sedimented as a 1:1 heterodimer upon sucrose gradient centrifugation. These findings are evidence of a physical interaction of UL15 and UL28 and a functional role for UL15 in directing UL28 to the nucleus.     

Akinete Formation in Tribonema bombycinum Derbes et Solier (Xanthophyceae) in Relation to Freezing Tolerance

Tribonema bombycinum (Xanthophyceae), was examined. T. bombycinum shifted from vegetative cells to akinetes with starving by a prolonged batch culture, by culture with a diluted medium, or by culture with a single nutrient-deficient medium. In addition, akinetes developed by desiccation, but cold treatment at 4 C did not facilitate akinete formation. During starving, the vegetative cells, which had a large central vacuole in the protoplasm and thin cell walls, finally changed to akinetes, which had many small vacuoles and oil droplets in the protoplasm and thick cell walls. During akinete formation by starving, the freezing tolerance (LT₅₀) increased gradually from −3 C in vegetative cells to far below −30 C in akinetes. When vegetative cells were subjected to equilibrium freezing, their size shrank greatly and aparticulate domains accompanied by fracture-jump lesions developed in the plasma membranes. Akinetes subjected to equilibrium freezing showed little shrinkage, and freezing-induced ultrastructural changes did not occur in the plasma membranes. The morphological changes in the process of akinete formation and the responses to equilibrium freezing resembled those of cold-acclimated terrestrial plants. Received 24 November 1998/ Accepted in revised form 1 February 1999     

Tissue-Specific Somatic Mosaicism in Spinal and Bulbar Muscular Atrophy Is Dependent on CAG-Repeat Length and Androgen Receptor–Gene Expression Level

The factors influencing the tissue-specific pattern of somatic mosaicism in CAG-repeat diseases have not yet been fully resolved. We performed a detailed analysis of the degree of somatic mosaicism in various tissues from 20 patients with spinal and bulbar muscular atrophy (SBMA), including 4 who were deceased. The most outstanding feature was the prominent somatic mosaicism observed in the cardiac and skeletal muscles, composed predominantly of postmitotic cells, and in the skin, prostate, and testis. The CNS tissues, liver, and spleen showed the least mosaicism. The tissue distribution of somatic mosaicism in patients with SBMA was markedly different from that in patients with Huntington disease (HD) and from that in patients with dentatorubral-pallidoluysian atrophy (DRPLA). The degree of somatic mosaicism correlated with the CAG-repeat number but not with age at examination. Furthermore, tissues with a higher mosaicism level corresponded well to those with a higher expression level of androgen receptor protein. The tissue-specific pattern of somatic mosaicism related not only to cell composition with different cell turnover rates but to repeat size and gene expression levels, and postnatal cell division is unlikely to be a major cause of somatic mosaicism probably because of the relative stability of CAG repeat in SBMA.     

Conformational behaviour of Cα,α-diphenylglycine: folded vs. extended structures in DϕG-containing tripeptides

The crystal structures of three fully protected tripeptides containing the Dϕg residue (C^α,α-diphenylglycine) in the central position are reported, namely Z-Gly-Dϕg-Gly-OMe ( a ), Z-Gly-Dϕg-Aib-OMe ( b ) and Z-Aib-Dϕg-Aib-OMe ( c ). The molecular conformations are quite unusual because the Dϕg residue adopts a folded conformation in the 3₁₀-helical region when the following residue adopts a folded conformation of opposite handedness (peptides b and c ). In contrast, the Dϕg residue adopts the more frequently observed fully extended conformation when the following residue adopts a semi-extended conformation (peptide a ). These findings are in agreement with the theoretical calculations on Ac-Dϕg-Aib-NHCH₃ and Ac-Aib-Dϕg-NHCH₃ also reported in this work. © 1998 European Peptide Society and John Wiley & Sons, Ltd.     

Rapid isolation and lipid characterization of plasma membranes from normal and malignant lymphoid cells of mouse

A rapid isolation method was developed for plasma membranes from mouse lymphoid cells such as lymph node lymphocytes, thymocytes, radiation-induced thymoma cells and L1210 cells. Lysates of these lymphoid cells were prepared by Dounce homogenization under hypotonic conditions and directly layered on sucrose step density gradients containing 2 mM CaCl₂ and 5 mM MgCl₂, and centrifuged at $\text{52 000 ×}\text{g}$ for 1 h. Plasma membrane fractions appeared at the interface between 20 and 42% sucrose in the gradients. The procedure permitted purified membranes from cells to be obtained within 3 h, and the preparations appeared to be uniform by electron microscopy. Specific activities of ($\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATpase}$, Mg²⁺-ATPase and 5′-nucleotidase of the isolated plasma membranes were enriched 23- to 61-fold, 12- to 15-fold and 18- to 34-fold, respectively, in comparison with those of the corresponding cell homogenates. Cholesterol content of the malignant cell membranes was lower than that of the normal membranes and the molar ratio of cholesterol to phospholipid of the malignant cell membranes was also lower than that of the normal membranes. A decreased plasmalogen content was observed in the malignant plasma membranes, together with a higher percentage of phosphatidylethanolamine and a lower percentage of phosphatidylserine. In the normal cell membranes, thymocytes contained a higher percentage of phosphatidylcholine and a lower percentage of sphingomyelin than those of the lymph node lymphocytes. At all temperature ranges (5 to 40°C) the plasma membranes of the malignant cells had lower microviscosity than those of the normal cells.     

Evolutionary Relationship and Structural Characterization of the EPF/EPFL Gene Family

EPF1-EPF2 and EPFL9/Stomagen act antagonistically in regulating leaf stomatal density. The aim of this study was to elucidate the evolutionary functional divergence of EPF/EPFL family genes. Phylogenetic analyses showed that AtEPFL9/Stomagen-like genes are conserved only in vascular plants and are closely related to AtEPF1/EPF2-like genes. Modeling showed that EPF/EPFL peptides share a common 3D structure that is constituted of a scaffold and loop. Molecular dynamics simulation suggested that AtEPF1/EPF2-like peptides form an additional disulfide bond in their loop regions and show greater flexibility in these regions than AtEPFL9/Stomagen-like peptides. This study uncovered the evolutionary relationship and the conformational divergence of proteins encoded by the EPF/EPFL family genes.