Dementia Revealed: Novel Chromosome 6 Locus for Late-Onset Alzheimer Disease Provides Genetic Evidence for Folate-Pathway Abnormalities

Genome-wide association studies (GWAS) of late-onset Alzheimer disease (LOAD) have consistently observed strong evidence of association with polymorphisms in APOE. However, until recently, variants at few other loci with statistically significant associations have replicated across studies. The present study combines data on 483,399 single nucleotide polymorphisms (SNPs) from a previously reported GWAS of 492 LOAD cases and 496 controls and from an independent set of 439 LOAD cases and 608 controls to strengthen power to identify novel genetic association signals. Associations exceeding the experiment-wide significance threshold () were replicated in an additional 1,338 cases and 2,003 controls. As expected, these analyses unequivocally confirmed APOE''s risk effect (rs2075650, ). Additionally, the SNP rs11754661 at 151.2 Mb of chromosome 6q25.1 in the gene MTHFD1L (which encodes the methylenetetrahydrofolate dehydrogenase (NADP+ dependent) 1-like protein) was significantly associated with LOAD (; Bonferroni-corrected P = 0.022). Subsequent genotyping of SNPs in high linkage disequilibrium () with rs11754661 identified statistically significant associations in multiple SNPs (rs803424, P = 0.016; rs2073067, P = 0.03; rs2072064, P = 0.035), reducing the likelihood of association due to genotyping error. In the replication case-control set, we observed an association of rs11754661 in the same direction as the previous association at P = 0.002 ( in combined analysis of discovery and replication sets), with associations of similar statistical significance at several adjacent SNPs (rs17349743, P = 0.005; rs803422, P = 0.004). In summary, we observed and replicated a novel statistically significant association in MTHFD1L, a gene involved in the tetrahydrofolate synthesis pathway. This finding is noteworthy, as MTHFD1L may play a role in the generation of methionine from homocysteine and influence homocysteine-related pathways and as levels of homocysteine are a significant risk factor for LOAD development.     

Sequence of plasmid pBS228 and reconstruction of the IncP-1alpha phylogeny

The antibiotic resistance plasmid pBS228 has been completely sequenced, and revealed to be descended from a plasmid virtually identical to the Birmingham IncP-1alpha plasmid RK2/RP4/RP1. However, it has three additional transposon insertions, one of which is responsible for the extra antibiotic resistances conferred. Loss of kanamycin resistance, which is characteristic of most IncP-1alpha plasmids, is the result of this insertion. A second transposon causes inactivation of the mating pair formation apparatus, rendering the plasmid non-self-transmissible. Comparison with the published data for other IncP-1alpha plasmids gives insight into the recent evolutionary history of this group as well as the acquisition and transmission of one of the first ampicillin resistance transposons discovered.     

Screening Marine Fungi for Inhibitors of the C4 Plant Enzyme Pyruvate Phosphate Dikinase: Unguinol as a Potential Novel Herbicide Candidate

A total of 2,245 extracts, derived from 449 marine fungi cultivated in five types of media, were screened against the C₄ plant enzyme pyruvate phosphate dikinase (PPDK), a potential herbicide target. Extracts from several fungal isolates selectively inhibited PPDK. Bioassay-guided fractionation of one isolate led to the isolation of the known compound unguinol, which inhibited PPDK with a 50% inhibitory concentration of 42.3 ± 0.8 μM. Further kinetic analysis revealed that unguinol was a mixed noncompetitive inhibitor of PPDK with respect to the substrates pyruvate and ATP and an uncompetitive inhibitor of PPDK with respect to phosphate. Unguinol had deleterious effects on a model C₄ plant but no effect on a model C₃ plant. These results indicate that unguinol inhibits PPDK via a novel mechanism of action which also translates to an herbicidal effect on whole plants.     

Characterisation of host defence proteins in milk using a proteomic approach

Besides providing nutrition to the newborn, milk also protects the neonate and the mammary gland against infection. As well as the six major proteins, bovine milk contains minor proteins, not all of which have been characterized. In this study, we have subjected bovine skim milk, whey, and milk fat globule membrane (MFGM) fractions to both direct liquid chromatography-tandem mass spectrometry (LC-MS/MS), and two-dimensional electrophoresis (2-DE) followed by matrix assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry (MS) of individual protein spots to better characterize the repertoire of minor milk proteins, particularly those involved with host defense. Milk from peak lactation as well as during the period of colostrum formation and during mastitis were analyzed to gain a more complete sampling of the milk proteome. In total, 2903 peptides were detected by LC-MS and 2770 protein spots by 2-DE. From these, 95 distinct gene products were identified, comprising 53 identified through direct LC-MS/MS and 57 through 2-DE-MS. The latter were derived from a total of 363 spots analyzed with 181 being successfully identified. At least 15 proteins were identified that are involved in host defense. These results demonstrate that the proteome of milk is more complex than has previously been reported and a significant fraction of minor milk proteins are involved in protection against infection.     

Insights into Ubiquitination from the Unique Clamp-like Binding of the RING E3 AO7 to the E2 UbcH5B

RING proteins constitute the largest class of E3 ubiquitin ligases. Unlike most RINGs, AO7 (RNF25) binds the E2 ubiquitin-conjugating enzyme, UbcH5B (UBE2D2), with strikingly high affinity. We have defined, by co-crystallization, the distinctive means by which AO7 binds UbcH5B. AO7 contains a structurally unique UbcH5B binding region (U5BR) that is connected by an 11-amino acid linker to its RING domain, forming a clamp surrounding the E2. The U5BR interacts extensively with a region of UbcH5B that is distinct from both the active site and the RING-interacting region, referred to as the backside of the E2. An apparent paradox is that the high-affinity binding of the AO7 clamp to UbcH5B, which is dependent on the U5BR, decreases the rate of ubiquitination. We establish that this is a consequence of blocking the stimulatory, non-covalent, binding of ubiquitin to the backside of UbcH5B. Interestingly, when non-covalent backside ubiquitin binding cannot occur, the AO7 clamp now enhances the rate of ubiquitination. The high-affinity binding of the AO7 clamp to UbcH5B has also allowed for the co-crystallization of previously described and functionally important RING mutants at the RING-E2 interface. We show that mutations having marked effects on function only minimally affect the intermolecular interactions between the AO7 RING and UbcH5B, establishing a high degree of complexity in activation through the RING-E2 interface.     

Historical demography of freshwater mussels (Bivalvia: Unionidae): genetic evidence for population expansion and contraction during the late Pleistocene and Holocene

Genetic variation was examined in two endangered mussel species, Epioblasma brevidens and Epioblasma capsaeformis, and in a non‐listed species, Lampsilis fasciola, in the Clinch River, Tennessee, USA, by screening mitochondrial DNA (mtDNA) sequences and nuclear DNA microsatellites. Patterns of mtDNA polymorphism exhibited different trends in long‐term population sizes for each species during the late Pleistocene and Holocene (～20 000 ya to present); namely, E. brevidens has declined over time, E. capsaeformis has remained demographically stable, and L. fasciola has expanded. However, analyses using microsatellites did not exhibit similar trends, perhaps because homoplasy had eliminated long‐term population signatures for the loci examined. For both marker types, long‐term effective population size (N_e) was low in E. brevidens, intermediate in E. capsaeformis, and high in L. fasciola. Moderately diverged mtDNA lineages, perhaps indicative of secondary contact, were observed in E. brevidens and E. capsaeformis. Perhaps the most surprising result of this study was the high level of genetic variation observed at both mtDNA and microsatellite DNA markers for L. fasciola, variation seemingly contrary to the relatively small demes that currently reside in the Clinch River. However, the data are consistent with known demographic and life‐history traits of these three mussel species and their fish hosts, namely that they each use hosts with different dispersal capabilities, ranging from low, moderate, and high, respectively. The low divergence of mtDNA sequence variation reported in this and other recent mussel studies indicates that considerable extant population genetic variation probably originated during the late Pleistocene and Holocene. © 2015 The Linnean Society of London, Biological Journal of the Linnean Society, 2015, 114 , 376–397.     

Structural characterization of a ribose-5-phosphate isomerase B from the pathogenic fungus Coccidioides immitis

Background

F-spondin is a multi-domain extracellular matrix (ECM) protein and a contact-repellent molecule that directs axon outgrowth and cell migration during development. The reelin_N domain and the F-spondin domain (FS domain) comprise a proteolytic fragment that interacts with the cell membrane and guides the projection of commissural axons to floor plate. The FS domain is found in F-spondins, mindins, M-spondin and amphiF-spondin.

Results

We present the crystal structure of human F-spondin FS domain at 1.95Å resolution. The structure reveals a Ca²⁺-binding C2 domain variant with an 8-stranded antiparallel β-sandwich fold. Though the primary sequences of the FS domains of F-spondin and mindin are less than 36% identical, their overall structures are very similar. The unique feature of F-spondin FS domain is the presence of three disulfide bonds associated with the N- and C-termini of the domain and a highly conserved N-linked glycosylation site. The integrin-binding motif found in mindin is not conserved in the F-spondin FS domain.

Conclusion

The structure of the F-spondin FS domain completes the structural studies of the multiple-domain ECM molecule. The homology of its core structure to a common Ca²⁺- and lipid-binding C2 domain suggests that the F-spondin FS domain may be responsible for part of the membrane targeting of F-spondin in its regulation of axon development. The structural properties of the FS domain revealed in this study pave the way for further exploration into the functions of F-spondin.     

Bryophytes as food: comparative consumption and utilization of mosses by a generalist insect herbivore

Compared with angiosperms, bryophytes are seldom fed upon by insects, despite being commonly used for shelter. Bryophytes are assumed to be unpalatable, and three classes of mechanisms have been suggested as possible barriers to bryophagy: chemical defenses, low digestibility, and low nutrient content. However, very few studies have tested these hypotheses. The present study examines pre‐ and post‐ingestive defenses of mosses. The acceptability and quality of four species of moss –Bryum argenteum Hedw. (Bryales: Bryaceae), Climacium americanum Brid. (Leucodontales: Climaciaceae), Leucobryum glaucum (Hedw.) (Dicranales: Leucobryaceae), and Sphagnum warnstorfii Russ. (Sphagnales: Sphagnaceae) – were compared with two control diets using the generalist caterpillar, Trichoplusia ni Hübner (Lepidoptera: Noctuidae: Plusiinae). In no‐choice trials, caterpillars consumed much less of any of the mosses than lettuce or wheat germ. The only moss consumed in sufficient quantities to evaluate post‐ingestive responses was C. americanum. Digestibility, assimilation, and overall utilization efficiency of C. americanum did not differ from that of lettuce, although C. americanum and lettuce were both less digestible than artificial diet. Choice assays using leaf discs showed that ethanol extract of L. glaucum, the least consumed moss, was deterrent, implying that chemical defenses play a major role in deterring feeding on L. glaucum. This study suggested that pre‐ingestive mechanisms are more important than post‐ingestive mechanisms in discouraging herbivory on mosses, and offered evidence that mosses are not simply nutrient poor.