EFFICIENT SYNTHESIS OF FUSED ISOTHIAZOLO C-NUCLEOSIDES. III. SYNTHESIS OF 7-SUBSTITUTED ISOTHIAZOLO[4,5-d]PYRIMIDINE C-NUCLEOSIDES*

The Divakar-Reese procedure has been successfully applied for transforming 7-oxo-isothiazolo[4,5-d]pyrimidine C-nucleosides (4a,b, 5a,b, 6a) via 1,2,4-triazol-1-yl intermediates (7a,b, 8a,b) into various 7-substituted C-nucle- osides 15a,b, 16a,b, 17a, 18a, 19a,b, 20a,b; their subsequent deprotection provides novel types of unusual C-glycosides 22b, 23a, 24a,b, 25b, 26b.

C-Nucleosides, possessing on its heterocyclic base other than naturally occuring oxo- or amino substituents, are important model compounds for biological or medicinal studies [2a] Hanessian, S. and Pernet, A. G. 1976. Adv. Carbohydr. Chem. Biochem., 32: 111–188. cf. [Google Scholar], [2b] Mizuno, Y. 1986. The Organic Chemistry of Nucleic Acids Amsterdam: Elsevier.  [Google Scholar], [2c] Huryn, D. M. and Okabe, M. 1992. Chem. Rev., 92: 1745–1768. [Crossref], [Web of Science ®] , [Google Scholar], [2d] Häbich, D. 1991. Chem. in uns. Zeit, 25: 295–307.  [Google Scholar], [2e] Uhlmann, E. and Peyman, A. 1990. Chem. Rev., 90: 543–584. [Crossref], [Web of Science ®] , [Google Scholar], [2f] Thuong, N. T. and Helene, C. 1993. Angew. Chem., 105: 697–723. [Crossref] , [Google Scholar], [2g] 1993. Angew. Chem. Int. Ed. Engl., 33: 666–690.  [Google Scholar], [2h] Yarchoan, R., Mitsuya, H., Zhomas, R. V., Pluda, J. M., Hartman, N. R., Perno, C. F., Marczyk, K. S., Allain, J. P., Johns, D. G. and Broder, S. 1989. Science, 245: 412–414. [Crossref], [PubMed], [Web of Science ®] , [Google Scholar], [2i] Tanaka, H., Baba, M., Hayakawa, H., Sakamaki, T., Miyasaka, T., Ubasawa, M., Takashima, H., Sekiya, E., Nitta, I., Shigeta, S., Walker, R. T., Balzarini, J. and De Clerq, E. 1991. J. Med. Chem., 34: 349–357. [Crossref], [PubMed], [Web of Science ®] , [Google Scholar] [3a] Koyama, G., Maeda, K., Umezawa, H. and Iitaka, Y. 1966. Tetrahedron Lett., : 597–602. Some C-glycosides with antibiotic, antiviral (HIV), and anticancer activities [Google Scholar], [3b] Hori, M., Wakashiro, T., Ito, E., Sawa, T., Takeuchi, T. and Umezawa, H. J. 1968. J. Antibiot., 21A: 264–270. [Chem. Abstr. 1968, 69, 11356j] [Google Scholar], [3c] Farkas, J. and ?orm. 1972. F. Collect. Czech. Chem. Commun., 37: 2798–2803.  [Google Scholar], [3d] Acton, E. M., Ryan, K. J., Henry, D. W. and Goodman, L. 1971. J. Chem. Soc., Chem. Commun., : 986–988.  [Google Scholar], [3e] Nakagawa, Y., Kano, H., Tsukuda, Y. and Koyama, H. 1967. Tetrahedron Lett., : 4105–4109.  [Google Scholar], [3f] Inoue, I. and Kuwaijama, I. 1980. J. Chem. Soc., Chem. Commun., : 251–253.  [Google Scholar], [3g] Buchanan, J. G., Stobie, A. and Wightman, R. H. 1980. ibid., : 916–917. [Crossref] , [Google Scholar], [3h] Hildebrand, S. and Leumann, C. 1996. Angew. Chem., 108: 2100–2102. Angew. Chem. Int. Ed. Engl. 1996, 35, 1968–1970 [Google Scholar]. We want to report on the synthesis of novel 7-substituted isothiazolo = [4,5-d]pyrimidine C-nucleosides. As we could show in previous papers [1] Wamhoff, H., Berressem, R. and Nieger, M. 1994. J. Org. Chem., 59: 1912–1917. Part 2 [Google Scholar], [4] Wamhoff, H., Berressem, R. and Nieger. 1993. M. J. Org. Chem., 58: 5181–5185.  [Google Scholar], there exists a simple approach to the protected C-glycosides 4–6.

     

Translational control of gurken mRNA in Drosophila development

Localized mRNA translation is a widespread mechanism for targeting protein synthesis, important for cell fate, motility and pathogenesis. In Drosophila, the spatiotemporal control of gurken/TGF-α mRNA translation is required for establishing the embryonic body axes. A number of recent studies have highlighted key aspects of the mechanism of gurken mRNA translational control at the dorsoanterior corner of the mid-stage oocyte. Orb/CPEB and Wispy/GLD-2 are required for polyadenylation of gurken mRNA, but unlocalized gurken mRNA in the oocyte is not fully polyadenylated.¹ Norvell A, Wong J, Randolph K, Thompson L. Wispy and Orb cooperate in the cytoplasmic polyadenylation of localized gurken mRNA. Dev Dyn Off Publ Am Assoc Anat 2015; 244:1276-1285. [Google Scholar] At the dorsoanterior corner, Orb and gurken mRNA have been shown to be enriched at the edge of Processing bodies, where translation occurs.² Weil TT, Parton RM, Herpers B, Soetaert J, Veenendaal T, Xanthakis D, Dobbie IM, Halstead JM, Hayashi R, Rabouille C, et al. Drosophila patterning is established by differential association of mRNAs with P bodies. Nat Cell Biol 2012; 14:1305-1313; PMID:23178881; http://dx.doi.org/10.1038/ncb2627[Crossref], [PubMed], [Web of Science ®] [Google Scholar] Over-expression of Orb in the adjacent nurse cells, where gurken mRNA is transcribed, is sufficient to cause mis-expression of Gurken protein.³ Davidson A, Parton RM, Rabouille C, Weil TT, Davis I. Localized translation of gurken/TGF-α mRNA during axis specification is controlled by access to Orb/CPEB on processing bodies. Cell Rep 2016; 14:2451-2462; PMID:26947065; http://dx.doi.org/10.1016/j.celrep.2016.02.038[Crossref], [PubMed], [Web of Science ®] [Google Scholar] In orb mutant egg chambers, reducing the activity of CK2, a Serine/Threonine protein kinase, enhances the ventralized phenotype, consistent with perturbation of gurken translation.⁴ Wong LC, Costa A, McLeod I, Sarkeshik A, Yates J 3rd, Kyin S, Perlman D, Schedl P, et al. The functioning of the drosophila CPEB protein Orb is regulated by phosphorylation and requires casein kinase 2 activity. PLoS One 2011; 6:e24355; PMID:21949709; http://dx.doi.org/10.1371/journal.pone.0024355[Crossref], [PubMed], [Web of Science ®] [Google Scholar] Here we show that sites phosphorylated by CK2 overlap with active Orb and with Gurken protein expression. Together with our new findings we consolidate the literature into a working model for gurken mRNA translational control and review the role of kinases, cell cycle factors and polyadenylation machinery highlighting a multitude of conserved factors and mechanisms in the Drosophila egg chamber.     

Reducing GWAS Complexity

Genome-wide association studies (GWAS) have revealed numerous genomic 'hits' associated with complex phenotypes. In most cases these hits, along with surrogate genetic variation as measure by numerous single nucleotide polymorphisms (SNPs) that are in linkage disequilibrium, are not in coding genes making assignment of functionality or causality intractable. Here we propose that fine-mapping along with the matching of risk SNPs at chromatin biofeatures lessen this complexity by reducing the number of candidate functional/causal SNPs. For example, we show here that only on average 2 SNPs per prostate cancer risk locus are likely candidates for functionality/causality; we further propose that this manageable number should be taken forward in mechanistic studies. The candidate SNPs can be looked up for each prostate cancer risk region in 2 recent publications in 2015^1,2 Han Y, Hazelett DJ, Wiklund F, Schumacher FR, Stram DO, Berndt SI, Wang Z, Rand KA, Hoover RN, Machiela MJ, et al. Integration of Multiethnic Fine-mapping and Genomic Annotation to Prioritize Candidate Functional SNPs at Prostate Cancer Susceptibility Regions. Hum Mol Genet 2015; 24(19):5603–18. Amin Al Olama A, Dadaev T, Hazelett DJ, Li Q, Leongamornlert D, Saunders EJ, Stephens S, Cieza-Borrella C, Whitmore I, Benlloch Garcia S, et al. Multiple novel prostate cancer susceptibility signals identified by fine-mapping of known risk loci among Europeans. Hum Mol Genet 2015; 24(19):5589–602.  from our groups.     

Tyr(less) kinase signaling during mitosis

Tyrosine phosphorylation is rare, representing only about 0.5% of phosphorylations in the cell under basal conditions. While mitogenic tyrosine kinase signaling has been extensively explored, the role of phosphotyrosine signaling across the cell cycle and in particular during mitosis is poorly understood.

Two recent, independent studies tackled this question from different angles to reveal exciting new insights into the role of this modification during cell division. Caron et al.¹ Caron D, Byrne DP, Thebault P, Soulet D, Landry CR, Eyers PA, Elowe S. Mitotic phosphotyrosine network analysis reveals that tyrosine phosphorylation regulates Polo-like kinase 1 (PLK1). Sci Signal 2016; 9:rs14; PMID:27965426; http://dx.doi.org/10.1126/scisignal.aah3525[Crossref], [PubMed], [Web of Science ®] [Google Scholar] exploited mitotic phosphoproteomics data sets to determine the extent of mitotic tyrosine phosphorylation, and St-Denis et al.² St-Denis N, Gupta GD, Lin ZY, Gonzalez-Badillo B, Veri AO, Knight JD, Rajendran D, Couzens AL, Currie KW, Tkach JM, et al. Phenotypic and interaction profiling of the human phosphatases identifies diverse mitotic regulators. Cell Rep 2016; 17:2488-501; PMID:27880917; http://dx.doi.org/10.1016/j.celrep.2016.10.078[Crossref], [PubMed], [Web of Science ®] [Google Scholar] identified protein tyrosine phosphatases from all subfamilies as regulators of mitotic progression or spindle formation. These studied collectively revealed that tyrosine phosphorylation may play a more prominent and active role in mitotic progression than previously appreciated.     

Buried Tracks: Ichnological Applications of High-Frequency Georadar

Recognition and sampling of traces in unconsolidated sands present a major challenge for ichnologists. This can be partially remedied through the application of high-resolution geophysical techniques, such as ground-penetrating radar (GPR or georadar), which uses electromagnetic impulse for continuous imaging of shallow subsurface. It addition to geological applications, GPR imaging has been used in several studies focused on animal traces as related to conservation of endangered fossorial species (Kinlaw et al., 2007 Kinlaw, A. E., Conyers, L. B. and Zajac, W. 2007. Use of ground penetrating radar to image burrows of the gopher tortoise (Gopherus polyphemus). Herpetological Review, 38: 50–56.  [Google Scholar]; Martin et al., 2011 Martin, A. J., Skaggs, S. A., Vance, R. K. and Greco, V. 2011. Ground-penetrating radar investigation of gopher-tortoise burrows: Refining the characterization of modern vertebrate burrows and associated commensal traces. Geological Society of America Abstracts with Programs, 43: 381 [Google Scholar]), slope and levee stability (Nichol et al., 2003 Nichol, D., Lenham, J. W. and Reynolds, J. M. 2003. Application of ground-penetrating radar to investigate the effects of badger setts on slope stability at St. Asaph Bypass, North Wales. Quarterly Journal of Engineering Geology and Hydrogeology, 36: 143–153.  [Google Scholar]; Di Prinzio et al., 2010 Di Prinzio, M, Bittelli, M., Castellarin, A. and Pisa, P. R. 2010. Application of GPR to the monitoring of river embankments. Journal of Applied Geophysics, 7: 53–61.  [Google Scholar]), and mapping of fossil tracks (Matthews et al., 2006 Matthews, N. A., Noble, T. A. and Breithaupt, B. H. 2006. “The application of photogrammetry, remote sensing and geographic information systems (GIS) to fossil resource management”. In Fossils from Federal Lands, Edited by: Lucas, S. G., Spielmann, J. A., Hester, P. M., Kenworthy, J. P. and Santucci, V. L. Vol. 34, 119–131. New Mexico Museum of Natural History and Science Bulletin.  [Google Scholar]; Aucoin and Hasbargen, 2010 Aucoin, C. D. and Hasbargen, L. 2010. Using GPR, GPS and close-range photography to map and characterize dinosaur tracks in the Connecticut River valley. Geological Society of America Abstracts with Programs, 42: 276 [Google Scholar]) and tracking surfaces (Webb, 2007 Webb, S. 2007. Further research of the Willandra Lakes fossil footprint site, southeastern Australia. Journal of Human Evolution, 52: 711–715. [Crossref], [PubMed], [Web of Science ®] , [Google Scholar]). Few efforts have been dedicated specifically to characterizing burrow and track characteristics (Stott, 1996 Stott, P. 1996. Ground-penetrating radar: a technique for investigating the burrow structure of fossorial vertebrates. Wildlife Research, 22: 519–530.  [Google Scholar]; Sensors & Software Inc., 2010 [compilation on geophysical projects related to animal burrows]; Buynevich and Hasiotis, 2011; Buynevich et al., 2011; Martin et al., 2011) and most of the above studies are published in journals not routinely accessed by ichnologists.     

Dissociation of recombinant prion autocatalysis from infectivity

ABSTRACT

Within the mammalian prion field, the existence of recombinant prion protein (PrP) conformers with self-replicating (ie. autocatalytic) activity in vitro but little to no infectious activity in vivo challenges a key prediction of the protein-only hypothesis of prion replication – that autocatalytic PrP conformers should be infectious. To understand this dissociation of autocatalysis from infectivity, we recently performed a structural and functional comparison between a highly infectious and non-infectious pair of autocatalytic recombinant PrP conformers derived from the same initial prion strain.¹ Noble GP, Wang DW, Walsh DJ, Barone JR, Miller MB, Nishina KA, Li S, Supattapone S. A Structural and Functional Comparison Between Infectious and Non-Infectious Autocatalytic Recombinant PrP Conformers. PLoS Pathog 2015; 11:e1005017; PMID:26125623; http://dx.doi.org/10.1371/journal.ppat.1005017[Crossref], [PubMed], [Web of Science ®] , [Google Scholar] We identified restricted, C-terminal structural differences between these 2 conformers and provided evidence that these relatively subtle differences prevent the non-infectious conformer from templating the conversion of native PrP^C substrates containing a glycosylphosphatidylinositol (GPI) anchor.¹ Noble GP, Wang DW, Walsh DJ, Barone JR, Miller MB, Nishina KA, Li S, Supattapone S. A Structural and Functional Comparison Between Infectious and Non-Infectious Autocatalytic Recombinant PrP Conformers. PLoS Pathog 2015; 11:e1005017; PMID:26125623; http://dx.doi.org/10.1371/journal.ppat.1005017[Crossref], [PubMed], [Web of Science ®] , [Google Scholar] In this article we discuss a model, consistent with these findings, in which recombinant PrP, lacking post-translational modifications and associated folding constraints, is capable of adopting a wide variety of autocatalytic conformations. Only a subset of these recombinant conformers can be adopted by post-translationally modified native PrP^C, and this subset represents the recombinant conformers with high specific infectivity. We examine this model's implications for the generation of highly infectious recombinant prions and the protein-only hypothesis of prion replication.     

Computer simulations of the two-species model for the melting curve maximum

The existence of extrema, maximum and/or minimum and negative gradients of melting curves observed for several elements at high pressure is investigated by molecular dynamics simulation using the two-species model (TSM) proposed by Ogura et al. [1 OguraH, MatsudaH, OgawaT, OgitaN, UedaA. Computer simulation for the melting curve maximum phenomenon. Prog Theor Phys. 1977;58:419–433.[Crossref], [Web of Science ®] , [Google Scholar]]. TSM is a model which imitates the change in the electronic structure of an atom in terms of a species change in particles. The TSM phase diagram has two solid phases and one liquid phase with a solid–solid–liquid triple point which corresponds to the melting curve minimum. The melting curve has both a maximum and a minimum, and the gradient of the melting curve is negative between these extrema. These peculiar melting curve properties and phase diagram are common to alkali metals and some other elements.     

Charge heterogeneity: Basic antibody charge variants with increased binding to Fc receptors

We identified active isoforms of the chimeric anti-GD2 antibody, ch14.18, a recombinant antibody produced in Chinese hamster ovary cells, which is already used in clinical trials.^1,2,3 Ladenstein R, Weixler S, Baykan B, Bleeke M, Kunert R, Katinger D, Pribill I, Glander P, Bauer S, Pistoia V, et al. Ch14.18 antibody produced in CHO cells in relapsed or refractory Stage 4 neuroblastoma patients: a SIOPEN Phase 1 study. MAbs 2013; 5:801-9; PMID:23924804; http://dx.doi.org/10.4161/mabs.25215 Desai AV, Fox E, Smith LM, Lim AP, Maris JM, Balis FM. Pharmacokinetics of the chimeric anti-GD2 antibody, ch14.18, in children with high-risk neuroblastoma. Cancer Chemother Pharmacol 2014; 74:1047-55; PMID:25212536; http://dx.doi.org/10.1007/s00280-014-2575-9 Siebert N, Eger C, Seidel D, Jüttner M, Zumpe M, Wegner D, Kietz S, Ehlert K, Veal GJ, Siegmund W, et al. Pharmacokinetics and pharmacodynamics of ch14.18/CHO in relapsed/refractory high-risk neuroblastoma patients treated by long-term infusion in combination with IL-2. MAbs 2016; 8:604-16; PMID:26785755; http://dx.doi.org/10.1080/19420862.2015.1130196  We separated the antibody by high resolution ion-exchange chromatography with linear pH gradient elution into acidic, main and basic charge variants on a preparative scale yielding enough material for an in-depth study of the sources and the effects of microheterogeneity. The binding affinity of the charge variants toward the antigen and various cell surface receptors was studied by Biacore. Effector functions were evaluated using cellular assays for antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity. Basic charge variants showed increased binding to cell surface receptor FcγRIIIa, which plays a major role in regulating effector functions. Furthermore, increased binding of the basic fractions to the neonatal receptor was observed. As this receptor mediates the prolonged half-life of IgG in human serum, this data may well hint at an increased serum half-life of these basic variants compared to their more acidic counterparts. Different glycoform patterns, C-terminal lysine clipping and N-terminal pyroglutamate formation were identified as the main structural sources for the observed isoform pattern. Potential differences in structural stability between individual charge variant fractions by nano differential scanning calorimetry could not been detected. Our in-vitro data suggests that the connection between microheterogeneity and the biological activity of recombinant antibody therapeutics deserves more attention than commonly accepted.     

Hitchhiking vesicular transport routes to the vacuole: Amyloid recruitment to the Insoluble Protein Deposit (IPOD)

Sequestration of aggregates into specialized deposition sites occurs in many species across all kingdoms of life ranging from bacteria to mammals and is commonly believed to have a cytoprotective function. Yeast cells possess at least 3 different spatially separated deposition sites, one of which is termed “Insoluble Protein Deposit (IPOD)” and harbors amyloid aggregates. We have recently discovered that recruitment of amyloid aggregates to the IPOD uses an actin cable based recruitment machinery that also involves vesicular transport.¹ Kumar R, Nawroth PP, Tyedmers J. Prion aggregates are recruited to the insoluble protein deposit (IPOD) via myosin 2-based vesicular transport. PLoS Genet 2016; 12:e1006324; PMID:27689885; http://dx.doi.org/10.1371/journal.pgen.1006324[Crossref], [PubMed], [Web of Science ®] [Google Scholar] Here we discuss how different proteins known to be involved in vesicular transport processes to the vacuole might act to guide amyloid aggregates to the IPOD. These factors include the Myosin V motor protein Myo2 involved in transporting vacuolar vesicles along actin cables, the transmembrane protein Atg9 involved in the recruitment of large precursor hydrolase complexes to the vacuole, the phosphatidylinositol/ phosphatidylcholine (PI/PC) transfer protein Sec 14 and the SNARE chaperone Sec 18. Furthermore, we present new data suggesting that the yeast dynamin homolog Vps1 is also crucial for faithful delivery of the amyloid model protein PrD-GFP to the IPOD. This is in agreement with a previously identified role for Vps1 in recruitment of heat-denatured aggregates to a perivacuolar deposition site.² Hill SM, Hao X, Gronvall J, Spikings-Nordby S, Widlund PO, Amen T, Jörhov A, Josefson R, Kaganovich D, Liu B, et al. Asymmetric inheritance of aggregated proteins and age reset in yeast are regulated by Vac17-dependent vacuolar functions. Cell Rep 2016; 16:826-38; PMID:27373154[Crossref], [PubMed], [Web of Science ®] [Google Scholar]     

A review of drug therapy for sporadic fatal insomnia

Background: Sporadic fatal insomnia (sFI) is a rapid progressive neurodegenerative disease characterised by gradual to perpetual insomnia, followed by dysautonomia, coma and death.¹ Lugaresi E, Medori R, Montagna P, Baruzzi A, Cortelli P, Lugaresi A, Tinuper P, Zucconi M, Gambetti P. Fatal familial insomnia and dysautonomia with selective degeneration of thalamic nuclei. New England J of Med. 1986;315:997–1003. doi:10.1056/NEJM198610163151605.[Crossref], [PubMed], [Web of Science ®] [Google Scholar] The cause of sFI was recently mapped to a mutation in a protein, the prion, found in the human brain. It is the unfolding of the prion that leads to the generation of toxic oligomers that destroy brain tissue and function. Recent studies have confirmed that a methionine mutation at codon 129 of the human Prion is characteristic of sFI. Current treatment slows down the progression of the disease, but no cure has been found, yet. Methods: We used Molecular Docking and Molecular Dynamics simulation methods, to study the toxic Fatal-Insomnia-prion conformations at local unfolding. The idea was to determine these sites and to stabilise these regions against unfolding and miss-folding, using a small ligand, based on a phenothiazine "moiety". Conclusion: As a result we here discuss current fatal insomnia therapy and present seven novel possible compounds for in vitro and in vivo screening.     

ARTD1 regulates cyclin E expression and consequently cell-cycle re-entry and G1/S progression in T24 bladder carcinoma cells

ADP-ribosylation is involved in a variety of biological processes, many of which are chromatin-dependent and linked to important functions during the cell cycle. However, any study on ADP-ribosylation and the cell cycle faces the problem that synchronization with chemical agents or by serum starvation and subsequent growth factor addition already activates ADP-ribosylation by itself. Here, we investigated the functional contribution of ARTD1 in cell cycle re-entry and G₁/S cell cycle progression using T24 urinary bladder carcinoma cells, which synchronously re-enter the cell cycle after splitting without any additional stimuli. In synchronized cells, ARTD1 knockdown, but not inhibition of its enzymatic activity, caused specific down-regulation of cyclin E during cell cycle re-entry and G₁/S progression through alterations of the chromatin composition and histone acetylation, but not of other E2F-1 target genes. Although Cdk2 formed a functional complex with the residual cyclin E, p27^{Kip1 Murray AH, Hunt T. The cell cycle: an introduction. New York: Oxford University Press, 1993. [Google Scholar]} protein levels increased in G₁ upon ARTD1 knockdown most likely due to inappropriate cyclin E-Cdk2-induced phosphorylation-dependent degradation, leading to decelerated G₁/S progression. These results provide evidence that ARTD1 regulates cell cycle re-entry and G₁/S progression via cyclin E expression and p27^{Kip1 Murray AH, Hunt T. The cell cycle: an introduction. New York: Oxford University Press, 1993. [Google Scholar]} stability independently of its enzymatic activity, uncovering a novel cell cycle regulatory mechanism.     

Creating a human head finite element model using a multi-block approach for predicting skull response and brain pressure

To better understand head injuries, human head finite element (FE) models have been reported in the literature. In scenarios where the head is directly impacted and measurements of head accelerations are not available, a high-quality skull model, as well as a high-quality brain model, is needed to predict the effect of impact on the brain through the skull. Furthermore, predicting cranial bone fractures requires comprehensively validated skull models. Lastly, high-quality meshes for both the skull and brain are needed for accurate strain/stress predictions across the entire head. Hence, we adopted a multi-block approach to develop hexahedral meshes for the brain, skull, and scalp simultaneously, a first approach in its kind. We then validated our model against experimental data of brain pressures (Nahum et al., 1977 Nahum AM, Smith R, Ward CC. 1977. Intracranial pressure dynamics during head impact. Proceedings of the 21st Stapp Car Crash Conference, SAE Paper No. 770922; Warrendale, PA: Society of Automotive Engineers.[Crossref] , [Google Scholar]) and comprehensive skull responses (Yoganandan et al., 1995 Yoganandan N, Pintar FA, Sances A, Jr., Walsh PR, Ewing CL, Thomas DJ, Snyder RG. 1995. Biomechanics of skull fracture. J Neurotrauma. 12(4):659–668.[Crossref], [PubMed], [Web of Science ®] , [Google Scholar], Yoganandan et al., 2004 Yoganandan N, Zhang J, Pintar FA. 2004. Force and acceleration corridors from lateral head impact. Traffic Injury Prevention. 5(4):368–373.[Taylor &; Francis Online] , [Google Scholar], and Raymond et al., 2009 Raymond D, Van Ee C, Crawford G, Bir C. 2009. Tolerance of the skull to blunt ballistic temporo-parietal impact. J Biomech. 42(15):2479–2485.[Crossref], [PubMed], [Web of Science ®] , [Google Scholar]). We concluded that a human head FE model was developed with capabilities to predict blunt- and ballistic-impact-induced skull fractures and pressure-related brain injuries.     

Small Amphibian and Reptile Footprints from the Permian Carapacha Basin,Argentina

This paper contains a taxonomic study of the Permian tetrapod ichnofauna from the Carapacha Basin. Tetrapod traces are analyzed in their environmental context and compared with similar faunas from Europe and North America. This ichnofauna is particularly relevant because of the scarcity of Permian tetrapod tracks from South America and also of Permian tetrapod fossils from Argentina. Ephemeral fluvial and shallow lacustrine deposits compose the sedimentary succession of the basin, which is represented by the Carapacha Formation. Most of the tracks have been collected from the upper member of the formation (Urre-Lauquen Member), mainly from freshwater ephemeral lake deposits as well as from playa-lake mudflats. The deposits of this member have been attributed to the early Late Permian on the basis of a Glossopteris fossil flora. Ichnotaxonomic designations of tetrapod traces are made on the basis of morphologic features that reflect the anatomy of the producer and special attention has been paid to extramorphologic deformations observed in the track assemblage. A total of four footprint ichnotaxa have been recognized, namely Batrachichnus salamandroides (Geinitz, 1861 Geinitz, H. B. 1861. Dyas oder die Zechsteinformation und das Rhotliegende—Die animalischen Überreste der Dyas, 130 ppLeipzig: Wilhelm Engelmann.  [Google Scholar]), Hyloidichnus bifurcatus Gilmore, 1927 Gilmore, C. W. 1927. Fossil footprints from the Grand Canyon: second contribution. Smithsonian Miscellaneous Collection, 80(3): 1–78.  [Google Scholar], cf. Amphisauropus isp. and cf. Varanopus isp. These track taxa are associated with two forms of vertebrate swimming traces (Characichnos isp. and type A swimming trace) and a possible fish trail. Invertebrate trace fossils include abundant arthropod locomotion traces and Scoyenia isp. The ichnofauna is composed of six tetrapod ichnocoenoses that are dominated by tiny amphibian tracks attributed to Temnospondyli (Batrachichnus and type A swimming trace) and Seymouriamorpha (Amphisauropus), and also contain the footprints of small reptiles, mostly Captorhinomorpha and possibly Pelycosauria (Hyloidichnus and Varanopus). Even if the ichnofauna of the Carapacha Basin is slightly younger than typical examples from the literature of the Early Permian “red bed ichnofacies” (Hunt et al., 1995b Hunt, A. P., Lucas, S. G., Lockley, M. G., Haubold, H. and Braddy, S. 1995b. Tetrapod ichnofacies in Early Permian red beds of the American Southwest. Bulletin New Mexico Museum of Natural History and Science, 6: 295–301. Early Permian footprint facies [Google Scholar]), a comparison is made. However, further detailed case studies are needed to formally define this “red bed ichnofacies” and its prospective subdivisions.     

Of Cecropias,Snarks and Boojums

We reply to Webber et al. (2011 Webber, BL, Born, C, Conn, BJ, Hadiah, JT and Zalamea, P-C. 2011. What is in a name? That which we call Cecropia peltata by any other name would be as invasive?. Plant Ecology & Diversity, 3: 241–245.  [Google Scholar]). Our study, Sheil and Padmanaba (2011 Sheil, D and Padmanaba, M. 2011. Innocent invaders? A preliminary assessment of Cecropia, an American tree, in Java. Plant Ecology & Diversity, 3: 231–240.  [Google Scholar]), was a preliminary assessment of the alien Neotropical tree Cecropia around Bogor, West Java, Indonesia. We highlighted the need for low-cost assessment approaches for addressing alien species and considered what might be achieved by local actors with limited resources. We successfully characterised the local distribution of Cecropia and identified, addressed and illustrated various concerns. Based on these results we asked how such studies might be useful and how they could be improved. Webber et al. criticise our study but fail to engage with its goals. Certainly, low-input local approaches lack the sophistication and rigour of larger international efforts, but such comparisons are unhelpful. Aside from a number of misunderstandings, we find Webber et al.’s principal arguments overstated, and unsupported by published information. The evidence that Cecropia causes harm is unconvincing; species-level identification is overemphasised; and the insistence on completing detailed research before considering management options is problematic. We offer some positive conclusions and repeat our plea for the development of effective low-input methods for evaluating and addressing naturalised organisms.     

Direct mass spectrometric characterization of disulfide linkages

Disulfide linkage is critical to protein folding and structural stability. The location of disulfide linkages for antibodies is routinely discovered by comparing the chromatograms of the reduced and non-reduced peptide mapping with location identification confirmed by collision-induced dissociation (CID) mass spectrometry (MS)/MS. However, CID product spectra of disulfide-linked peptides can be difficult to interpret, and provide limited information on the backbone region within the disulfide loop. Here, we applied an electron-transfer dissociation (ETD)/CID combined fragmentation method that identifies the disulfide linkage without intensive LC comparison, and yet maps the disulfide location accurately. The native protein samples were digested using trypsin for proteolysis. The method uses RapiGest SF Surfactant and obviates the need for reduction/alkylation and extensive sample manipulation. An aliquot of the digest was loaded onto a C₄ analytical column. Peptides were gradient-eluted and analyzed using a Thermo Scientific LTQ Orbitrap Elite mass spectrometer for the ETD-triggered CID MS³ Wypych J, Li M, Guo A, Zhang Z, Martinez T, Allen MJ, Fodor S, Kelner DN, Flynn GC, Liu YD, et al. Human IgG2 antibodies display disulfide-mediated structural isoforms. J Biol Chem. 2008;283:16194–205. doi:10.1074/jbc.M709987200. PMID:18339624[Crossref], [PubMed], [Web of Science ®] , [Google Scholar] experiment. Survey MS scans were followed by data-dependent scans consisting of ETD MS² scans on the most intense ion in the survey scan, followed by 5 MS³ CID scans on the 5 most intense ions in the ETD MS² scan. We were able to identify the disulfide-mediated structural variants A and A/B forms and their corresponding disulfide linkages in an immunoglobulin G2 monoclonal antibody with λ light chain (IgG2λ), where the location of cysteine linkages were unambiguously determined.     

Sampling Plan Optimization: A Data Review and Sampling Frequency Evaluation Process

Lawrence Livermore National Laboratory (LLNL) uses a cost-effective sampling (CES) methodology to evaluate and review ground water contaminant data and optimize the site's ground water monitoring plan. The CES methodology is part of LLNL's regulatory approved compliance monitoring plan (Lamarre et al., 1996 Lamarre, A. L., Nichols, E. M., Berg, L. L., Dresen, M. D., Gelinas, R. J., Bainer, R. W. and Folsom, E. N. 1996. Compliance monitoring plan for the Lawrence Livermore National Laboratory Livermore Site UCRL-AR-120936 [Google Scholar]). It allows LLNL to adjust the ground water sampling plan every quarter in response to changing conditions at the site. Since the use of the CES methodology has been approved by the appropriate regulatory agencies, such adjustments do not need additional regulatory approval. This permits LLNL to respond more quickly to changing conditions. The CES methodology bases the sampling frequency for each location on trend, variability, and magnitude statistics describing the contaminants at that location, and on the input of the technical staff (hydrologists, chemists, statisticians, and project leaders). After initial setup is complete, each application of CES takes only a few days for as many as 400 wells. Effective use of the CES methodology requires sufficient data, an understanding of contaminant transport at the site, and an adequate number of monitoring wells downgradient of the contamination. The initial implementation of CES at LLNL in 1992 produced a 40% reduction in the required number of annual routine ground water samples at LLNL. This has saved LLNL $390,000 annually in sampling, analysis, and data management costs.