Studies on the chemoprophylaxis of subperiodic Brugia malayi infection in the leaf monkey (Presbytis melalophos) with diethylcarbamazine citrate

The chemoprophylactic use of diethylcarbamazine citrate at total oral doses of 15--180 mg/kg body weight was tested against subperiodic Brugia malayi infection in the leaf monkey (Presbytis melalophos). A total dose of 45 mg/kg body weight given over 9 days killed all developing infective larvae. Similarly, a total dose of 35 mg/kg body weight given over 7 days killed all fourth stage larvae. The minimum effective dose that prevents infection would be 5 mg/kg body weight daily for 7 days every month.     

Structural and trace element changes in scalp hair of radiographers

Scalp hair samples were collected from medical radiographers and nonradiographers of matching age groups. Structural morphology of hair was studied by scanning electron microscopy, and the trace element profiles in hair were measured using neutron activation analysis. The structural damage to the hair follicles of the radiographers was quite obvious, and this may be a good qualitative indicator of radiation damage at low doses. The concentrations of aluminum (Al), potassium (K), and vanadium (V) in hair of the radiographers were significantly higher, whereas those of antimony (Sb) and magnesium (Mg) were significantly lower than those of nonradiographers. Some of our findings were quite consistent with those of others in determining the changes in trace element concentrations in irradiated tissue.     

Increased gene coverage and Alu frequency in large linkage disequilibrium blocks of the human genome

The human genome has linkage disequilibrium (LD) blocks, within which single-nucleotide polymorphisms show strong association with each other. We examined data from the International HapMap Project to define LD blocks and to detect DNA sequence features inside of them. We used permutation tests to determine the empirical significance of the association of LD blocks with genes and Alu repeats. Very large LD blocks (>200 kb) have significantly higher gene coverage and Alu frequency than the outcome obtained from permutation-based simulation, whereas there was no significant positive correlation between gene density and block size. We also observed a reduced frequency of Alu repeats at the gaps between large LD blocks, indicating that their enrichment in large LD blocks does not introduce recombination hotspots that would cause these gaps.     

A data-driven model of a modal gated ion channel: The inositol 1,4,5-trisphosphate receptor in insect Sf9 cells

The inositol 1,4,5-trisphosphate (IP(3)) receptor (IP(3)R) channel is crucial for the generation and modulation of intracellular Ca(2+) signals in animal cells. To gain insight into the complicated ligand regulation of this ubiquitous channel, we constructed a simple quantitative continuous-time Markov-chain model from the data. Our model accounts for most experimentally observed gating behaviors of single native IP(3)R channels from insect Sf9 cells. Ligand (Ca(2+) and IP(3)) dependencies of channel activity established six main ligand-bound channel complexes, where a complex consists of one or more states with the same ligand stoichiometry and open or closed conformation. Channel gating in three distinct modes added one complex and indicated that three complexes gate in multiple modes. This also restricted the connectivity between channel complexes. Finally, latencies of channel responses to abrupt ligand concentration changes defined a model with specific network topology between 9 closed and 3 open states. The model with 28 parameters can closely reproduce the equilibrium gating statistics for all three gating modes over a broad range of ligand concentrations. It also captures the major features of channel response latency distributions. The model can generate falsifiable predictions of IP(3)R channel gating behaviors and provide insights to both guide future experiment development and improve IP(3)R channel gating analysis. Maximum likelihood estimates of the model parameters and of the parameters in the De Young-Keizer model yield strong statistical evidence in favor of our model. Our method is simple and easily applicable to the dynamics of other ion channels and molecules.     

Cortactin regulates podosome formation: roles of the protein interaction domains

Cortactin, a multi-domain scaffolding protein involved in actin polymerization, is enriched in podosomes induced by phorbol ester in vascular smooth muscle cells. We generated several functional and truncation mutants of cortactin to probe the roles of various protein interaction domains in the regulation of the dynamics of podosome formation. At the onset of podosome genesis, cortactin clustered near the ends of stress fibers that appeared to act as nucleation platforms onto which the actin polymerization machinery assembled. Translocation of cortactin to these pre-podosome clusters required the intact N-WASp-binding SH3 domain. Overexpression of the C-terminal third of cortactin containing the intact SH3 domain inhibited podosome formation presumably by sequestering of N-WASp and prevented cortactin clustering. Subsequent assembly of the actin-rich core of podosomes required translocation of additional cortactin to the actin core, a process that required the actin-binding repeats, but not the Arp2/3-binding N-terminal acidic region nor the SH3 domain. These results suggest that the SH3 domain and the actin-binding repeat region are involved, respectively, in the early and late stages of podosome formation process.     

Phosphorylation of cortactin by p21-activated kinase

Cortactin is an F-actin binding protein that is enriched in dynamic cytoskeletal organelles such as podosomes, membrane ruffles, and lamellipodia. We have shown previously that Src-phosphorylation of cortactin is not required for its translocation to phorbol-ester induced podosomes in A7r5 aortic smooth muscle cells, but may be important for stability and turnover of podosomes. However, little is known of the role of Ser/Thr kinases in the regulation of cortactin. Here, we report that p21-associated kinase (PAK), which plays a crucial role in the formation of podosome and membrane ruffles, is able to phosphorylate cortactin in vitro. The predominant phosphorylation site is located at Ser113 in the first actin-binding repeat. Phosphorylation by PAK is not required for the translocation of cortactin to podosomes, lamellipodia, or membrane ruffles in A7r5 smooth muscle cells. However, binding of cortactin to F-actin is significantly reduced by PAK-phosphorylation. Taken together, these results suggest a role for PAK-phosphorylation of cortactin in the regulation of the dynamics of branched actin filaments in dynamic cytoskeletal organelles.     

New remains of <Emphasis Type="Italic">Hungarosaurus tormai</Emphasis> (Ankylosauria,Dinosauria) from the Upper Cretaceous of Hungary: skeletal reconstruction and body mass estimation

Hungarosaurus tormai is a medium-sized nodosaurid ankylosaur that was described on the basis of four partial skeletons from the Santonian (Upper Cretaceous) of Hungary. In this paper, a new fifth skeleton and several isolated remains are described which greatly improve our knowledge of this primitive nodosaurid ankylosaur. Isolated cranial remains referred to juvenile individuals provide new information on the development of cranial ornamentation in nodosaurid ankylosaurs. Apart from both preserved mandibles with in situ dentition, the fifth partial skeleton contains several previously unknown limb elements (humerus, ulnae, radius) that indicate unusual limb proportions for Hungarosaurus compared with other ankylosaurs. On the basis of the five partial skeletons and the isolated remains, a skeletal and dermal armor reconstruction is attempted. Body mass calculations using three different methods yield an estimate of 650 kg for H. tormai.      

Phorbol ester-induced podosomes in normal human bronchial epithelial cells

Spreading and migration of the basal cells neighboring a wound is essential for airway epithelial repair. To gain insight into the molecular mechanisms that govern these cellular processes, we asked whether normal human airway epithelial cells can form podosomes, a cellular structure discovered from cancer and mesenchymal cells that controls migration and invasion. Herein, we report that phorbol-12, 13-dibutyrate (PDBu), a protein kinase C activator, induced reorganization of cytoskeletal structure in primary normal human bronchial epithelial cells, and in normal human airway epithelial BEAS2B cells. Z-stack scanning confocal microscopy showed that PDBu-induced podosome-like structures contain actin-rich columns that arise from the ventral surface of the cell, and also revealed the presence of circular ruffles/waves at the dorsal cell surface. The molecular components of these cytoskeletal structures were determined with immunofluorescent staining. Using in situ zymography, we demonstrated that PDBu-induced podosomes were capable of degrading fibronectin-gelatin-sucrose matrix. PDBu also increased epithelial cell invasion across Transwell chamber. Podosomes and circular dorsal ruffles may be important for epithelial cell migration and invasion, thus contributing to respiratory epithelial repair and regeneration.     

Accurately Measuring Recombination between Closely Related HIV-1 Genomes

Retroviral recombination is thought to play an important role in the generation of immune escape and multiple drug resistance by shuffling pre-existing mutations in the viral population. Current estimates of HIV-1 recombination rates are derived from measurements within reporter gene sequences or genetically divergent HIV sequences. These measurements do not mimic the recombination occurring in vivo, between closely related genomes. Additionally, the methods used to measure recombination make a variety of assumptions about the underlying process, and often fail to account adequately for issues such as co-infection of cells or the possibility of multiple template switches between recombination sites. We have developed a HIV-1 marker system by making a small number of codon modifications in gag which allow recombination to be measured over various lengths between closely related viral genomes. We have developed statistical tools to measure recombination rates that can compensate for the possibility of multiple template switches. Our results show that when multiple template switches are ignored the error is substantial, particularly when recombination rates are high, or the genomic distance is large. We demonstrate that this system is applicable to other studies to accurately measure the recombination rate and show that recombination does not occur randomly within the HIV genome.