The higher order repeat structure of chromatin is built up of globular particles containing eight nucleosomes

Mild nuclease digestion of rat liver chromatin generates particles with sedimentation coefficients of about 33S, 60S, and 90S (in 50 mM NaCl). The kinetics of appearance and disappearance of these particles with progressive digestion suggest that they are produced by cleavage from a higher order repeat structure, the 33S particle representing the monomer. At an intermediate stage of digestion, about 75 % of the nuclear chromatin can be recovered as monomers to trimers of this higher order structure. Sedimentation profiles indicate that monomer particles containing 7–8 nucleosomes occur at the highest frequency. The DNA fragments in monomers have a size corresponding to hepta- and octanucleosomes, and those in dimers have a size corresponding to chains of sixteen nucleosomes. The higher order repeat structure is only stable between 30 and 200 mM NaCl; the particles unfold below 30 and above 200 mM NaCl. When examined by electron microscopy, monomers and dimers appear as compact globular structures. Relaxation by lowering the salt concentration results in the appearance of polynucleosomes with a chain length of eight beads in the monomer and sixteen in the dimer particle. These results indicate that the unit particle of the higher order repeat structure of rat liver chromatin contains eight nucleosomes.     

Heterogeneity of spacer lengths in circles of amplified ribosomal DNA of two insect species, Dytiscus marginalis and Acheta domesticus.

Crystals of subtilopeptidase A have been studied by means of X-ray diffraction. They are orthorhombic, space group P2₁2₁2₁ with $\text{a = 76 73}\text{A}\text{?}\text{, b = 55.35}\text{A}\text{?}\text{and}\text{c = 53.00}\text{A}\text{?}$. There is one molecule per asymmetric unit and the solvent content is 42%. The crystals diffract very well and appear suitable for detailed study of the three-dimensional structure of the enzyme.     

Incorporation of Wild-Type and C-Terminally Truncated Human Epidermal Growth Factor Receptor into Human Immunodeficiency Virus-Like Particles: Insight into the Processes Governing Glycoprotein Incorporation into Retroviral Particles

Previous results have indicated that incorporation of surface glycoprotein into retroviral particles is not a specific process and that many heterologous viral and cellular glycoproteins can be incorporated as long as they do not have long cytoplasmic C-terminal regions which were presumed to be sterically inhibitory. In this study, this concept has been directly examined by analyzing the incorporation of the wild-type human epidermal growth factor receptor (Wt-EGFR) and of a C-terminally truncated mutant of Wt-EGFR (Tr-EGFR) into human immunodeficiency virus (HIV)-like particles. Incorporation was directly analyzed at the protein level and by immunogold labelling of enriched HIV-like particles. In agreement with the above concept, Tr-EGFR, with only 7 C-terminal amino acids (aa), was efficiently incorporated into HIV-like particles. Incorporation of the Wt-EGFR species, with 542 C-terminal cytoplasmic aa, was reduced by a factor of about 5 in comparison to that of the Tr-EGFR species. However, the Wt-EGFR species was still very significantly present in the HIV-like particles. A series of control experiments verified that this represents genuine incorporation of Wt-EGFR into the membrane of HIV-like particles. These observations allow further speculation as to the processes governing glycoprotein incorporation into retroviral particles and indicate that the internal virus structure of HIV (in particular the matrix layer [MA]) can accommodate much larger heterologous cytoplasmic domains in incorporated glycoproteins than previously assumed.     

Receptor recognition by a hepatitis B virus reveals a novel mode of high affinity virus-receptor interaction

The duck hepatitis B virus model system was used to elucidate the characteristics of receptor (carboxypeptidase D, gp180) interaction with polypeptides representing the receptor binding site in the preS part of the large viral surface protein. We demonstrate the pivotal role of carboxypeptidase D for virus entry and show its C-domain represents the virus attachment site, which binds preS with extraordinary affinity. Combining results from surface plasmon resonance spectroscopy and two-dimensional NMR analysis we resolved the contribution of preS sequence elements to complex stability and show that receptor binding potentially occurs in two steps. Initially, a short alpha-helix in the C-terminus of the receptor binding domain facilitates formation of a primary complex. This complex is stabilized sequentially, involving approximately 60 most randomly structured amino acids preceding the helix. Thus, hepadnaviruses exhibit a novel mechanism of high affinity receptor interaction by conserving the potential to adapt structure during binding rather than to preserve it per se. We propose that this process represents an alternative strategy to escape immune surveillance and the evolutionary pressure inherent in the compact hepadnaviral genome organization.     

Homo- and heterodimerization of APP family members promotes intercellular adhesion

The amyloid precursor protein (APP) plays a central role in Alzheimer's disease, but its physiological function and that of its mammalian paralogs, the amyloid precursor-like proteins 1 and 2 (APLPs), is still poorly understood. APP has been proposed to form dimers, a process that could promote cell adhesion via trans-dimerization. We investigated the dimerization and cell adhesion properties of APP/APLPs and provide evidence that all three paralogs are capable of forming homo- and heterocomplexes. Moreover, we show that trans-interaction of APP family proteins promotes cell-cell adhesion in a homo- and heterotypic fashion and that endogenous APLP2 is required for cell-cell adhesion in mouse embryonic fibroblasts. We further demonstrate interaction of all the three APP family members in mouse brain, genetic interdependence, and molecular interaction of APP and APLPs in synaptically enriched membrane compartments. Together, our results provide evidence that homo- and heterocomplexes of APP/APLPs promote trans-cellular adhesion in vivo.     

The correlation between oxidative stress and leaf senescence during plant development

In plants, besides being the final step leading to the death of the whole organism, senescence has a developmental function involving the coordinated degradation of macromolecules and the mobilization of nutrients out of senescing tissues into developing parts of the plant. Free radicals are thought to play an essential role in senescence, especially those derived from oxygen. Since these molecules are extremely toxic, the levels of the different reactive oxygen species have to be tightly regulated. However, at low concentrations, hydrogen peroxide may also serve as a signalling molecule. Therefore, a coordinated regulation of the free radical scavenging system, which comprises enzymatic components such as catalase, superoxide dismutase and ascorbate peroxidase, and non-enzymatic molecules such as ascorbate and glutathione is essential. The increased radical levels displayed during senescence are not only caused by the elevated production of radicals but also by a loss in antioxidant capacity.     

Characterization of prototype foamy virus gag late assembly domain motifs and their role in particle egress and infectivity

Foamy viruses (FV) are unusual among retroviruses since they require both Gag and Env structural proteins for particle egress. Recently significant progress has been made towards the mechanistic understanding of the viral release process, in particular that of retroviruses, and the viral domains and cellular pathways involved. However little is currently known about domains of FV structural proteins and cellular proteins engaged in this process. By mutational analysis of sequence motifs in prototype FV (PFV) Gag, bearing homology to known late assembly (L) domains, a PSAP motif with L domain function that was functionally interchangeable by heterologous L domains was identified. In contrast the inactivation of a PPPI motif had no significant influence on PFV particle release, although mutant viral particles displayed reduced infectivity. Similarly mutation of an evolutionary conserved YXXL motif revealed no classical L-domain function but resulted in release of noninfectious viruslike particles. Biochemical and electron microscopy analysis demonstrated that these mutant particles incorporated all viral structural proteins but contained aberrantly capsid structures, suggesting a role in capsid assembly for this PFV Gag sequence motif. In line with the mutational analysis, overexpression of dominant negative (DN) mutants and wild-type TSG101 but not the DN mutant of AIP-1/ALIX reduced PFV particle release and infectivity. Furthermore, DN mutants of Vps4A, Vps4B, and CHMP3 inhibited PFV egress and infectivity. Taken together these results demonstrate that PFV, like other viruses, requires components of the vacuolar protein sorting (VPS) machinery for egress and enters the VPS pathway through interaction with TSG101.     

Senescence-related gene expression profiles of rosette leaves of Arabidopsis thaliana: leaf age versus plant age

Abstract: Senescence is a form of programmed cell death (PCD) which leads to the death of whole organs, e.g., leaves or flowers, and eventually to the death of entire plants. Like all forms of PCD, senescence is a highly regulated and energy consuming process. Senescence parameters, like protein content, chlorophyll content, expression of photosynthesis-associated genes or senescence-associated genes (SAGs), reveal that senescence occurs in old leaves derived from young plants (6 week old) as well as in young leaves derived from older plants (8 week old), indicating that it is governed by the actual age of the leaves. In order to analyse the differential gene expression profiles during leaf senescence, hybridizations of high-density genome arrays were performed with: i) individual leaves within the rosette of a 6-week-old plant and ii) leaves of the same position within the rosette but harvested from plants of different ages, ranging from 5 to 8 weeks. Cluster and genetree analyses, according to the expression pattern revealed that genes which are up-regulated with respect to the age of the entire plant, showed completely different expression profiles with respect to the age of the individual leaves within one rosette. This was observed even though the actual difference in leaf age was approximately the same. This indicates that gene expression appears to be governed by different parameters: i) the age of the individual leaf and ii) the age and developmental stage of the entire plant.