The impact of cyclin-dependent kinase 5 depletion on poly(ADP-ribose) polymerase activity and responses to radiation

Cyclin-dependent kinase 5 (Cdk5) has been identified as a determinant of sensitivity to poly(ADP-ribose) polymerase (PARP) inhibitors. Here, the consequences of its depletion on cell survival, PARP activity, the recruitment of base excision repair (BER) proteins to DNA damage sites, and overall DNA single-strand break (SSB) repair were investigated using isogenic HeLa stably depleted (KD) and Control cell lines. Synthetic lethality achieved by disrupting PARP activity in Cdk5-deficient cells was confirmed, and the Cdk5^KD cells were also found to be sensitive to the killing effects of ionizing radiation (IR) but not methyl methanesulfonate or neocarzinostatin. The recruitment profiles of GFP-PARP-1 and XRCC1-YFP to sites of micro-irradiated Cdk5^KD cells were slower and reached lower maximum values, while the profile of GFP-PCNA recruitment was faster and attained higher maximum values compared to Control cells. Higher basal, IR, and hydrogen peroxide-induced polymer levels were observed in Cdk5^KD compared to Control cells. Recruitment of GFP-PARP-1 in which serines 782, 785, and 786, potential Cdk5 phosphorylation targets, were mutated to alanines in micro-irradiated Control cells was also reduced. We hypothesize that Cdk5-dependent PARP-1 phosphorylation on one or more of these serines results in an attenuation of its ribosylating activity facilitating persistence at DNA damage sites. Despite these deficiencies, Cdk5^KD cells are able to effectively repair SSBs probably via the long patch BER pathway, suggesting that the enhanced radiation sensitivity of Cdk5^KD cells is due to a role of Cdk5 in other pathways or the altered polymer levels.     

Genomic insights into the other malaria

Plasmodium vivax has received less attention and study than Plasmodium falciparum, due in part to difficulties in culturing this pathogen. Whole-genome sequencing of both P. vivax and Plasmodium cynomolgi and characterization of genetic variation in these species provide a genetic toolbox for tertian malaria and new insights into the monkey malaria clade.     

Nesting phenology and productivity of birds in the White and Inyo Mountains, California, as assessed with nest-boxes

Nest-boxes were monitored from 1988 to 1992 on 5 grids in the pinyon-juniper ( Pinus-Juniperus ) woodlands of the White and Inyo Mountains, California, to determine breeding phenology and productivity of cavity-nesting birds, and characteristics of used and non-used nest-boxes. We found a total of 112 nests of 6 species. Bewicks Wrens ( Thryomanes bewicki i; 64 nests), Mountain Chickadees ( Poecile gambeli ; 18), Ash-throated Flycatchers ( Myiarchus cinerascens ; 12), and Juniper Titmice ( Baeolophus ridgwayi ; 11) were the most common species utilizing the boxes. Nest phenology and numbers of young were similar to values reported elsewhere in the literature for the species, although nesting success for Mountain Chickadees appeared lower in our study. Chickadees were associated with relatively dense, mature vegetation on southern slopes. Juniper Titmice used areas with tall juniper shrubs and generally sparse vegetation. Bewicks Wrens used areas with short trees, sparse vegetation, and many stumps. Nest-boxes were underutilized (< 15% use) by most species relative to their availability and relative to detected abundances of the species. We suggest that nest-boxes can provide valuable breeding biology information and can potentially increase the productivity of rare and cavity-limited species.     

Regional and seasonal diet of the Western Burrowing Owl in south central Nevada

We examined diets of Western Burrowing Owls ( Athene cunicularia hypugaea ) based on contents of pellets and large prey remains collected year-round at burrows in each of the 3 regions in south central Nevada (Mojave Desert, Great Basin Desert, and Transition region). The most common prey items, based on percent frequency of occurrence, were crickets and grasshoppers, beetles, rodents, sun spiders, and scorpions. The most common vertebrate prey was kangaroo rats ( Dipodomys spp.). True bugs (Hemiptera), scorpions, and western harvest mice ( Reithrodontomys megalotis ) occurred most frequently in pellets from the Great Basin Desert region. Kangaroo rats ( Dipodomys spp.) and pocket mice (Perognathinae) were the most important vertebrate prey items in the Transition and Mojave Desert regions, respectively. Frequency of occurrence of any invertebrate prey was high (>80%) in samples year-round but dropped in winter samples, with scorpions and sun spiders exhibiting the steepest declines. Frequency of occurrence of any vertebrate prey peaked in spring samples, was intermediate for winter and summer samples, and was lowest in fall samples. With the possible exception of selecting for western harvest mice in the Great Basin Desert region, Western Burrowing Owls in our study appeared to be opportunistic foragers with a generalist feeding strategy.     

Complex reorganization and predominant non-homologous repair following chromosomal breakage in karyotypically balanced germline rearrangements and transgenic integration

We defined the genetic landscape of balanced chromosomal rearrangements at nucleotide resolution by sequencing 141 breakpoints from cytogenetically interpreted translocations and inversions. We confirm that the recently described phenomenon of 'chromothripsis' (massive chromosomal shattering and reorganization) is not unique to cancer cells but also occurs in the germline, where it can resolve to a relatively balanced state with frequent inversions. We detected a high incidence of complex rearrangements (19.2%) and substantially less reliance on microhomology (31%) than previously observed in benign copy-number variants (CNVs). We compared these results to experimentally generated DNA breakage-repair by sequencing seven transgenic animals, revealing extensive rearrangement of the transgene and host genome with similar complexity to human germline alterations. Inversion was the most common rearrangement, suggesting that a combined mechanism involving template switching and non-homologous repair mediates the formation of balanced complex rearrangements that are viable, stably replicated and transmitted unaltered to subsequent generations.     

Synthesis of brome mosaic virus subgenomic RNA in vitro by internal initiation on (-)-sense genomic RNA

The genomes of many (+)-stranded RNA viruses, including plant viruses and alphaviruses, consist of polycistronic RNAs whose internal genes are expressed via subgenomic messenger RNAs. The mechanism(s) by which these subgenomic mRNAs arise are poorly understood. Based on indirect evidence, three models have been proposed: (1) internal initiation by the replicase on the (-)-strand of genomic RNA, (2) premature termination during (-)-strand synthesis, followed by independent replication of the subgenomic RNA and (3) processing by nuclease cleavage of genome-length RNA. Using an RNA-dependent RNA polymerase (replicase) preparation from barley leaves infected with brome mosaic virus (BMV) to synthesize the viral subgenomic RNA in vitro, we now provide evidence that subgenomic RNA arises by internal initiation on the (-)-strand of genomic RNA. We believe that this also represents the first in vitro demonstration of a replicase from a eukaryotic (+)-stranded RNA virus capable of initiating synthesis of (+)-sense RNA.     

Growth mechanisms of Ag and Cu nanodendrites via Galvanic replacement reactions

Dendritic silver and copper crystals were produced via Galvanic replacement reactions on zinc and aluminum plates, respectively. The growth orientations of these metals were determined using electron microscopy. The results showed that a fast crystal growth associated with a high concentration of metal cations led to kinetically controlled growth along the <112> axes of the cubic close-packed structures. However, a slow growth rate resulted in thermodynamically controlled growth along the [111] axis. The crystal growth was not found to rely upon the direct deposition of metal cations at crystallographic sites on crystal facets, but instead, hydrated metal cations deposited on the crystal surface to form an amorphous coating layer, followed by the reduction of metal cations and crystallization at the crystal/coating interface. Twin defects and stacking faults were often observed across the whole particle and commonly observed ?{422} diffraction spots were explained by stacking faults rather than by the possible presence of any superstructures. The present work offers evidences to claim that both the crystal growth rate and Coulomb interaction between negatively charged crystal surface and metal cations play an important role in the formation of metal dendrites in replacement reactions.