Cryo‐EM structure of native human uromodulin,a zona pellucida module polymer

Assembly of extracellular filaments and matrices mediating fundamental biological processes such as morphogenesis, hearing, fertilization, and antibacterial defense is driven by a ubiquitous polymerization module known as zona pellucida (ZP) “domain”. Despite the conservation of this element from hydra to humans, no detailed information is available on the filamentous conformation of any ZP module protein. Here, we report a cryo‐electron microscopy study of uromodulin (UMOD)/Tamm–Horsfall protein, the most abundant protein in human urine and an archetypal ZP module‐containing molecule, in its mature homopolymeric state. UMOD forms a one‐start helix with an unprecedented 180‐degree twist between subunits enfolded by interdomain linkers that have completely reorganized as a result of propeptide dissociation. Lateral interaction between filaments in the urine generates sheets exposing a checkerboard of binding sites to capture uropathogenic bacteria, and UMOD‐based models of heteromeric vertebrate egg coat filaments identify a common sperm‐binding region at the interface between subunits.     

Integrative Structural Investigation on the Architecture of Human Importin4_Histone H3/H4_Asf1a Complex and Its Histone H3 Tail Binding

Importin4 transports histone H3/H4 in complex with Asf1a to the nucleus for chromatin assembly. Importin4 recognizes the nuclear localization sequence located at the N-terminal tail of histones. Here, we analyzed the structures and interactions of human Importin4, histones and Asf1a by cross-linking mass spectrometry, X-ray crystallography, negative-stain electron microscopy, small-angle X-ray scattering and integrative modeling. The cross-linking mass spectrometry data showed that the C-terminal region of Importin4 was extensively cross-linked with the histone H3 tail. We determined the crystal structure of the C-terminal region of Importin4 bound to the histone H3 peptide, thus revealing that the acidic patch in Importin4 accommodates the histone H3 tail, and that histone H3 Lys14 contributes to the interaction with Importin4. In addition, we show that Asf1a modulates the binding of histone H3/H4 to Importin4. Furthermore, the molecular architecture of the Importin4_histone H3/H4_Asf1a complex was produced through an integrative modeling approach. Overall, this work provides structural insights into how Importin4 recognizes histones and their chaperone complex.     

Holarctic phylogeography of an asexual species complex – II. Allozymic variation and clonal structure in Arctic Daphnia

Previous mitochondrial (mt)DNA sequencing and restriction fragment length polymorphism (RFLP) studies have shown that the Holarctic Daphnia pulex complex is divisible into two major groups (pulicaria and tenebrosa) that exhibit distinct phylogeographic patterns. Here we examine allozymic variation at six polymorphic enzyme loci to ascertain clonal structure and clonal distribution patterns within each group. Specimens were collected from a total of 850 populations encompassing the Arctic. A significant negative relationship (Mantel test) between similarity of regional clonal arrays and geographical distance was observed. A small fraction of clones in each group was widespread (in the order of 1000s of kilometres). However, most clones were restricted to single regions, and were often found only in a single population. These data indicate that the population genetic structure is highly fragmented in this complex, but the potential for long-distance passive dispersal exists. Further, ‘hot spots’ of high clonal richness and diversity were found in each group, which is concordant with earlier work. In addition, ≈ 20% of pulicaria group clones possess nuclear genes from tenebrosa, while approximately 10% of tenebrosa group clones harbour pulicaria nuclear genes. These data indicate nuclear introgression between the two groups, which was found to be prominent in a broad zone of secondary contact encompassing parts of northwestern Russia, northern Fennoscandia, Svalbard, and extending into the high eastern Canadian Arctic.     

Virulence of Septoria nodorum as Affected by Passage Through Detached Leaves of Wheat and Barley Cultivars1

Abstract Three genetically marked, single–spore isolates of Septoria nodorum from wheat were passed through detached leaves of wheat cvs Blueboy and Coker 747 and the barley cv. Boone to produce three sub–isolates per original isolate. Each sub–isolate was cultured for three pycnidiospore generations on its respective host. Virulence of each sub–isolate on detached leaves of Blueboy, Caldwell, Coker 747, and NK81W701 wheat, and Boone and Surry barley was compared with that of the original single–spore isolate from which it was derived. In most cases, sub–isolates passed through wheat were significantly more virulent than the originals on wheat cultivars. They also were more virulent to barley than the original isolates but they were less virulent to barley than to wheat cultivars. Isolate × cultivar interactions were statistically significant (P < .0001) for isolates passed through wheat or barley and were greater than isolate × cultivar interactions among the original isolates. In seven of eight isolates passed through wheat or barley, only the original genetic marker was recovered after three generations, indicating that cross–contamination could not account for the observed change of virulence. In the single case of apparent contamination, of a sub–isolate, virulence declined.     

Directed termination of the polymerase chain reaction: kinetics and applications in mutation detection.

We describe a PCR-based method (DT-PCR) that integrates both DNA amplification and directed chain termination into a single-step process. This method exploits unbalanced nucleotide concentrations to induce the polymerase chain reaction to terminate at specific nucleotide sites, leading to the generation of two sets of nested termination fragments from genomic DNA. The kinetic mechanism underlying the termination process is outlined and the application of this method to the detection and characterization of mutations in fragments as long as 1 kb is described. The method is effective for the analysis of both haploid and diploid genomes, and not only allows the recognition of both indels (insertions and deletions) and nucleotide substitutions, but also enables the determination of their position in a single-step fashion.     

A general method of isolating high molecular weight DNA from methanogenic archaea (archaebacteria).

High molecular weight DNA was readily isolated from all methanogens treated, as well as from thermophilic anaerobic eubacteria, by grinding cells frozen in liquid N2, prior to lysis with SDS. DNA can subsequently be purified by the usual phenol-chloroform extractions. The procedure yields DNA readily cut by restriction enzymes and suitable for oligonucleotide probing, as well as for mole percent G + C content determination by thermal denaturation. The method routinely yields DNA of high molecular weight and is an improvement over DNA isolation methods for many methanogens, which often involve an initial breakage of the cells in a French pressure cell.     

A Psychiatric Home Care Program: A Report on Three Years' Experience (1962-1964)

The activities covering a three-year period of a psychiatric home care treatment program attached to a psychiatric unit of a general hospital are described. A detailed account of its operation and the roles played by each member of the team is given. This service frequently provides a substitute for hospitalization in the management of both acute and chronic psychiatric states and thereby constitutes an important preventive measure in the field of public health. Even if the initial attitude of the patient is negative it is possible to gain the co-operation of the family who become a useful ally in the treatment. The co-operation of the patient is not as essential as has been thought. The traditional role of the psychiatrist is reversed by virtue of his attending the patient at home. The active participation of social agencies is an integral part of the treatment.     

Clonal diversity in high arctic ostracodes

It remains unclear why the majority of parthenogenetic lineages persist for only brief periods of evolutionary time. However, by characterizing their patterns of genetic variation, it is possible to gain insights regarding their evolutionary origin and potential. We examined clonal diversity patterns in high arctic populations of freshwater ostracodes with the goal of clarifying the factors promoting genotypic diversity. Allozyme electrophoresis showed that the three dominant ostracode species in high arctic ponds reproduce via apomictic parthenogenesis that two of these species (Prionocypris glacialis, Candona rectangulata) were both highly clonally diverse, and had allozyme phenotypes suggestive of polyploidy. Scanning microdensitometry confirmed that many clones of P. glacialis at Igloolik were polyploid. In contrast to most other polyploids, clones of P. glacialis seem to be autopolyploids. Although clonal variation in P. glacialis may reflect multiple transitions to parthenogenesis in an undetected sexual population, it seems likely that genomic recombination associated with polyploidy has also played a role in generating local diversity following the transition to parthenogenesis.