Distal Sequences Research Articles

FOXF1 transcription factor promotes lung morphogenesis by inducing cellular proliferation in fetal lung mesenchyme

Organogenesis is regulated by mesenchymal-epithelial signaling events that induce expression of cell-type specific transcription factors critical for cellular proliferation, differentiation and appropriate tissue patterning. While mesenchymal transcription factors play a key role in mesenchymal-epithelial interactions, transcriptional networks in septum transversum and splanchnic mesenchyme remain poorly characterized. Forkhead Box F1 (FOXF1) transcription factor is expressed in mesenchymal cell lineages; however, its role in organogenesis remains uncharacterized due to early embryonic lethality of Foxf1-/- mice. In the present study, we generated mesenchyme-specific Foxf1 knockout mice (Dermo1-Cre Foxf1-/-) and demonstrated that FOXF1 is required for development of respiratory, cardiovascular and gastrointestinal organ systems. Deletion of Foxf1 from mesenchyme caused embryonic lethality in the middle of gestation due to multiple developmental defects in the heart, lung, liver and esophagus. Deletion of Foxf1 inhibited mesenchyme proliferation and delayed branching lung morphogenesis. Gene expression profiling of micro-dissected distal lung mesenchyme and ChIP sequencing of fetal lung tissue identified multiple target genes activated by FOXF1, including Wnt2, Wnt11, Wnt5A and Hoxb7. FOXF1 decreased expression of the Wnt inhibitor Wif1 through direct transcriptional repression. Furthermore, using a global Foxf1 knockout mouse line (Foxf1-/-) we demonstrated that FOXF1-deficiency disrupts the formation of the lung bud in foregut tissue explants. Finally, deletion of Foxf1 from smooth muscle cell lineage (smMHC-Cre Foxf1-/-) caused hyper-extension of esophagus and trachea, loss of tracheal and esophageal muscle, mispatterning of esophageal epithelium and decreased proliferation of smooth muscle cells. Altogether, FOXF1 promotes lung morphogenesis by regulating mesenchymal-epithelial signaling and stimulating cellular proliferation in fetal lung mesenchyme.

A promoter interaction map for cardiovascular disease genetics.

Over 500 genetic loci have been associated with risk of cardiovascular diseases (CVDs); however, most loci are located in gene-distal non-coding regions and their target genes are not known. Here, we generated high-resolution promoter capture Hi-C (PCHi-C) maps in human induced pluripotent stem cells (iPSCs) and iPSC-derived cardiomyocytes (CMs) to provide a resource for identifying and prioritizing the functional targets of CVD associations. We validate these maps by demonstrating that promoters preferentially contact distal sequences enriched for tissue-specific transcription factor motifs and are enriched for chromatin marks that correlate with dynamic changes in gene expression. Using the CM PCHi-C map, we linked 1999 CVD-associated SNPs to 347 target genes. Remarkably, more than 90% of SNP-target gene interactions did not involve the nearest gene, while 40% of SNPs interacted with at least two genes, demonstrating the importance of considering long-range chromatin interactions when interpreting functional targets of disease loci.

Strontium Isotope Stratigraphy: Principles and State of the Art

The methodical basis, development, and current state of a new method of chronostratigraphic studies, i.e., strontium isotope stratigraphy (SIS), are considered. This method makes it possible to date and correlate geographically distant sedimentary sequences without involving the biostratigraphic and isotope geochronological data. SIS is based on secular variations in 87Sr/86Sr in the paleocean, resulting from the redistribution of the roles of two global strontium flows formed in the mantle and continental reservoirs of the Earth. Isotopic homogeneity of Sr in the paleoceans and in the linked seas leads to the fact that the 87Sr/86Sr ratio in the sea basins is individual for each geological time point and is inherited in marine chemogenic sediments under deposition of dissolved Sr as an isomorphic impurity. Low-Mg calcite and also fragments of fossilized paleontological remains buried in situ are the best minerals that are capable of retaining the Sr isotopic signature of the sedimentation environment. SIS is carried out with geochemical diagnostics of secondary alteration of the studied material and selective dissolution of the samples to produce a carbonate material that adequately reflects isotopic signature of the sedimentary basin. Interregional correlations of the Proterozoic and Cenozoic sea sediments and their relation to the SIS-based stratigraphic scale are given as an example.

How is structural divergence related to evolutionary information?

The analysis of evolutionary information in a protein family, such as conservation and covariation, is often linked to its structural information. Multiple sequence alignments of distant homologous sequences are used to measure evolutionary variables. Although high structural differences between proteins can be expected in such divergent alignments, most works linking evolutionary and structural information use a single structure ignoring the structural variability within protein families.The goal of this work is to elucidate the relevance of structural divergence when sequence-based measures are integrated with structural information.We found that inter-residue contacts and solvent accessibility undergo large variations in protein families. Our results show that high covariation scores tend to reveal residue contacts that are conserved in the family, instead of protein or conformer specific contacts.We also found that residue accessible surface area shows a high variability between structures of the same family. As a consequence, the mean relative solvent accessibility of multiple structures correlates better with the conservation pattern than the relative solvent accessibility of a single structure.We conclude that the use of comprehensive structural information allows a more accurate interpretation of the information computed from sequence alignments. Therefore, considering structural divergence would lead to a better understanding of protein function, dynamics, and evolution.

Quantitative Analysis of Proximal and Distal Kinetic Chain Musculature During Dynamic Exercises.

Oliver, GD, Washington, JK, Barfield, JW, Gascon, SS, and Gilmer, G. Quantitative analysis of proximal and distal kinetic chain musculature during dynamic exercises. J Strength Cond Res 32(6): 1545-1553, 2018-Proximal to distal sequencing for the dynamic movement of throwing is dependent on the movement and stability of the lumbopelvic-hip complex (LPHC) and scapula. Although the need for proximal stability for distal mobility has been vastly documented, pre-throwing programs tend to focus on the traditional rotator cuff activation exercises before long toss. Thus, it was the purpose of this study to describe muscle activations of LPHC stabilizing musculature (bilateral gluteus medius and maximus) and scapular stabilizing musculature (dominant side latissimus dorsi, lower trapezius, upper trapezius, and serratus anterior) during 5 kinetic chain exercises that could be implemented in a throwing program. It was hypothesized that both the LPHC and the scapular stabilizing musculature would exhibit moderate to high activation during all the selected kinetic chain exercises. Nineteen healthy college students (23.2 ± 7.2 years; 176.7 ± 17.9 cm; 78.0 ± 28.6 kg) participated. Surface electromyography was used to measure muscle activity in the LPHC and scapular stabilizing musculature during 5 kinetic chain exercises. A nonparametric Friedman test revealed significantly different muscle activations as a factor of exercise for each muscle, χ(18) = 417.220, p < 0.001. The 5 kinetic chain exercises successfully elicited moderate to high muscle activation in all musculature, except the upper trapezius. Because greater muscle activation of the LPHC and scapular stabilizers are crucial during a throwing task, these exercises are recommended for pre-throwing program implementation because they efficiently prepare the stabilizing musculature for lengthy or strenuous throwing tasks, resulting in a potential decrease in injury susceptibility.

Selective Activation of Alternative MYC Core Promoters by Wnt-Responsive Enhancers.

In Metazoans, transcription of most genes is driven by the use of multiple alternative promoters. Although the precise regulation of alternative promoters is important for proper gene expression, the mechanisms that mediates their differential utilization remains unclear. Here, we investigate how the two alternative promoters (P1, P2) that drive MYC expression are regulated. We find that P1 and P2 can be differentially regulated across cell-types and that their selective usage is largely mediated by distal regulatory sequences. Moreover, we show that in colon carcinoma cells, Wnt-responsive enhancers preferentially upregulate transcription from the P1 promoter using reporter assays and in the context of the endogenous Wnt induction. In addition, multiple enhancer deletions using CRISPR/Cas9 corroborate the regulatory specificity of P1. Finally, we show that preferential activation between Wnt-responsive enhancers and the P1 promoter is influenced by the distinct core promoter elements that are present in the MYC promoters. Taken together, our results provide new insight into how enhancers can specifically target alternative promoters and suggest that formation of these selective interactions could allow more precise combinatorial regulation of transcription initiation.

Divergent rectrix moult: the implications and conditions of moult sequence

Wing and tail morphology strongly affect flight performance which may consequently decline during feather moult due to the creation of feather gaps in the flight‐surface. Hence, the size and shape of moult‐related gaps may directly affect flight capacity. Here, I examined the divergent rectrix moult sequence compared to the more common distal moult sequence. In the divergent moult, the focus of rectrix moult is shifted from the tail centre (R1; rectrices numbered distally from mid‐tail outward) to another rectrix (R2 or R3), and then rectrices are moulted bidirectionally, towards the tail centre and outwards. The result of this moult sequence is the splitting of the tail gap into multiple smaller gaps. Using a large moult database including 5669 individuals of 47 Western Palaearctic passerine species, I found evidence of divergent moult sequence for only seven species. Using comparative and experimental approaches, I found that the divergent rectrix sequence is correlated with higher moult speed and lower aerodynamic cost. Furthermore, the divergent rectrix sequence is more common among adults than juveniles. This work focused on the feather moult sequence – a seldom studied aspect of the avian life‐history. I propose that moult‐related aerodynamic costs may be an important evolutionary factor not only in moult speed, but also in moult sequence.

Molecular typing of Staphylococcus aureus based on coagulase gene.

AimThis study was conducted to study the coagulase gene-based genetic diversity of Staphylococcus aureus, isolated from different samples of cattle using restriction fragment length polymorphism (RFLP) and their sequence-based phylogenetic analysis.Materials and MethodsA total of 192 different samples from mastitic milk, nasal cavity, and pus from skin wounds of cattle from Military Dairy Farm, Jammu, India, were screened for the presence of S. aureus. The presumptive isolates were confirmed by nuc gene-based polymerase chain reaction (PCR). The confirmed S. aureus isolates were subjected to coagulase (coa) gene PCR. Different coa genotypes observed were subjected to RFLP using restriction enzymes Hae111 and Alu1, to obtain the different restriction patterns. One isolate from each restriction pattern was sequenced. These sequences were aligned for maximum homology using the Bioedit softwareandsimilarity in the sequences was inferred with the help of sequence identity matrix.ResultsOf 192 different samples,39 (20.31%) isolates of S. aureus were confirmed by targeting nuc gene using PCR. Of 39 S. aureus isolates, 25 (64.10%) isolates carried coa gene. Four different genotypes of coa gene, i.e., 514 bp, 595 bp, 757 bp, and 802 bp were obtained. Two coa genotypes, 595 bp (15 isolates) and 802 bp (4 isolates), were observed in mastitic milk. 514 bp (2 isolates) and 757 bp (4 isolates) coa genotypes were observed from nasal cavity and pus from skin wounds, respectively. On RFLP using both restriction enzymes, four different restriction patterns P1, P2, P3, and P4 were observed. On sequencing, four different sequences having unique restriction patterns were obtained. The most identical sequences with the value of 0.810 were found between isolate S. aureus 514 (nasal cavity) and S. aureus 595 (mastitic milk), and thus, they are most closely related. While as the most distant sequences with the value of 0.483 were found between S. aureus 514 and S. aureus 802 isolates.ConclusionThe study, being localized to only one farm, yielded different RFLP patterns as observed from different sampling sites, which indicates that different S. aureus coagulase typeshave a site-specific predilection. Two coa patterns were observed in mastitic milk indicating multiple origins of infection, with 595 bp coa genotype being predominant in mastitic milk. The coa genotypes and their restriction patterns observed in the present study are novel, not published earlier. 514 and 595 coa variants of S. aureus are genetically most related.

Developing in 3D: the role of CTCF in cell differentiation.

CTCF is a highly conserved zinc-finger DNA-binding protein that mediates interactions between distant sequences in the genome. As a consequence, CTCF regulates enhancer-promoter interactions and contributes to the three-dimensional organization of the genome. Recent studies indicate that CTCF is developmentally regulated, suggesting that it plays a role in cell type-specific genome organization. Here, we review these studies and discuss how CTCF functions during the development of various cell and tissue types, ranging from embryonic stem cells and gametes, to neural, muscle and cardiac cells. We propose that the lineage-specific control of CTCF levels, and its partnership with lineage-specific transcription factors, allows for the control of cell type-specific gene expression via chromatin looping.

Reccurrent F8 Intronic Deletion Found in Mild Hemophilia A Causes Alu Exonization

Incorporation of distant intronic sequences in mature mRNA is an underappreciated cause of genetic disease. Several disease-causing pseudoexons have been found to contain repetitive elements such as Alu elements. This study describes an original pathological mechanism by which a small intronic deletion leads to Alu exonization. We identified an intronic deletion, c.2113+461_2113+473del, in the F8 intron 13, in two individuals with mild hemophilia A. Invivo and invitro transcript analysis found an aberrant transcript, with an insertion of a 122-bp intronic fragment (c.2113_2114ins2113+477_2113+598) at the exon 13-14 junction. This out-of-frame insertion is predicted to lead to truncated protein (p.Gly705Aspfs∗37). DNA sequencing analysis found that the pseudoexon corresponds to antisense AluY element and the deletion removed a part of the poly(T)-tail from the right arm of these AluY. The heterogenous nuclear riboprotein C1/C2 (hnRNP C) is an important antisense Alu-derived cryptic exon silencer and binds to poly(T)-tracts. Disruption of the hnRNP C binding site in AluY T-tract by mutagenesis or hnRNP C knockdown using siRNA in HeLa cells reproduced the effect of c.2113+461_2113+473del. The screening of 114 unrelated families with mild hemophilia A in whom no genetic event was previously identified found a deletion in the poly(T)-tail of AluY in intron 13 in 54% of case subjects (n = 61/114). In conclusion, this study describes a deletion leading to Alu exonization found in 6.1% of families with mild hemophila A in France.

Dengue virus infection induces chromatin remodeling at locus AAEL006536 in the midgut of Aedes aegypti.

To identify and characterize Aedes aegypti's AAEL006536 gene proximal upstream cis-regulatory sequences activated by dengue virus infection. A. aegypti Rockefeller strain mosquitoes were blood fed or infected with dengue virus 2. Open chromatinprofiling was then carried out in pools of midguts from each group of mosquitoes. The proximal upstream region does not contain open chromatin sites in the midguts of blood-fed mosquitoes as detected by FAIRE-qPCR. In contrast, two cis-regulatory sites were identified in the same upstream region of dengue virus-infected mosquito midguts. The distal sequence contains STAT-, REL- and C/EBP-type transcription factor binding sites. The activation of two proximal cis-regulatory sequences, induced by dengue virus infection, is mediated by chromatin remodeling mechanisms. Binding sites suggest a dengue virus infectioninduced participation of immunity transcription factors in the up-regulation of this gene. This suggests the participation of the AAEL006536 gene in the mosquito's antiviral innate immune response.

Mutations in the Influenza A Virus M1 Protein Enhance Virus Budding To Complement Lethal Mutations in the M2 Cytoplasmic Tail.

The influenza A virus M1 and M2 proteins play important roles in virus assembly and in the morphology of virus particles. Mutations in the distal cytoplasmic tail region of M2, and in particular a tyrosine-to-alanine mutation at residue 76 (Y76A), were essential for infectious virus production and filament formation while having limited effects on total virus particle budding. Using a novel selection method, mutations at seven different M1 amino acids (residue 73, 94, 135, 136, or 138 or a double mutation, 93/244) that are not found in circulating influenza virus strains or have not been previously identified to play a role in influenza A virus assembly were found to complement the lethal M2Y76A mutation. These M1 suppressor mutations restored infectious virus production in the presence of M2Y76A and mediated increased budding and filament formation even in the absence of M2. However, the efficiency of infectious virus replication was still dependent on the presence of the distal region of the M2 cytoplasmic tail. The data suggest that influenza A virus budding and genome incorporation can occur independently and provide further support for complementary roles of the M1 and M2 proteins in virus assembly.IMPORTANCE Influenza virus particle assembly involves the careful coordination of various viral and host factors to optimally produce infectious virus particles. We have previously identified a mutation at position 76 of the influenza A virus M2 protein that drastically reduces infectious virus production and filament formation with minimal effects on virus budding. In this work, we identified suppressor mutations in the M1 protein which complement this lethal M2 mutation by increasing the efficiency with which virus particles bud from infected cells and restoring filament formation at the infected-cell surface. M2 distal cytoplasmic domain sequences were still required for optimal infectivity. This indicates that M1 and M2 can functionally replace each other in some, but not all, aspects of virus particle assembly.

Architectural Effects on Floral Traits in Sedum praealtum Dc. (Crassulaceae) in Mexico

Morphological and functional characteristics of flowers have been the primary criterion for traditional plant taxonomy and systematic classification because of their assumed constancy among individuals within a population. The effects of resource competition and pollen limitation from those of inflorescence architecture were separately studied during flowering season of the species to investigate patterns of investment allocation and fruit set among flowers within inflorescences of Sedum praealtum. I found that architecture was primarily accountable for the variation observed in reproductive traits of flowers within inflorescences. Specifically, the S. praealtum inflorescences exhibit a proximal to distal investment allocation sequence from female-biased to male-biased flowers, whereas the observed differences in fruit set were at least partially attributable to pollen limitation in distal-positioned flowers. Given the opportunity for different contributions of female and male functions to reproductive success, potential gender specialization of flowers at different positions within the inflorescence are discussed.

Structural features of the TatC membrane protein that determine docking and insertion of a twin-arginine signal peptide

Twin-arginine translocation (Tat) systems transport folded proteins across cellular membranes with the concerted action of mostly three membrane proteins: TatA, TatB, and TatC. Hetero-oligomers of TatB and TatC form circular substrate-receptor complexes with a central binding cavity for twin-arginine-containing signal peptides. After binding of the substrate, energy from an electro-chemical proton gradient is transduced into the recruitment of TatA oligomers and into the actual translocation event. We previously reported that Tat-dependent protein translocation into membrane vesicles of Escherichia coli is blocked by the compound N,N'-dicyclohexylcarbodiimide (DCCD, DCC). We have now identified a highly conserved glutamate residue in the transmembrane region of E. coli TatC, which when modified by DCCD interferes with the deep insertion of a Tat signal peptide into the TatBC receptor complex. Our findings are consistent with a hydrophobic binding cavity formed by TatB and TatC inside the lipid bilayer. Moreover, we found that DCCD mediates discrete intramolecular cross-links of E. coli TatC involving both its N- and C-tails. These results confirm the close proximity of two distant sequence sections of TatC proposed to concertedly function as the primary docking site for twin-arginine signal peptides.

Foreshocks and delayed triggering of the 2016 MW7.1 Te Araroa earthquake and dynamic reinvigoration of its aftershock sequence by the MW7.8 Kaikōura earthquake, New Zealand

We analyze the preparatory period of the September 2016 MW7.1 Te Araroa foreshock–mainshock sequence in the Northern Hikurangi margin, New Zealand, and subsequent reinvigoration of Te Araroa aftershocks driven by a large distant earthquake (the November 2016 MW7.8 Kaikōura earthquake). By adopting a matched-filter detection workflow using 582 well-defined template events, we generate an improved foreshock and aftershock catalog for the Te Araroa sequence (>8,000 earthquakes over 66 d). Templates characteristic of the MW7.1 sequence (including the mainshock template) detect several highly correlating events (ML2.5–3.5) starting 12 min after a MW5.7 foreshock. These pre-cursory events occurred within ∼1 km of the mainshock and migrate bilaterally, suggesting precursory slip was triggered by the foreshock on the MW7.1 fault patch prior to mainshock failure. We extend our matched-filter routine to examine the interactions between high dynamic stresses resulting from passing surface waves of the November 2016 MW7.8 Kaikōura earthquake, and the evolution of the Te Araroa aftershock sequence. We observe a sudden spike in moment release of the aftershock sequence immediately following peak dynamic Coulomb stresses of 50–150 kPa on the MW7.1 fault plane. The triggered increase in moment release culminated in a MW5.1 event, immediately followed by a ∼3 h temporal stress shadow. Our observations document the preparatory period of a major subduction margin earthquake following a significant foreshock, and quantify dynamic reinvigoration of a distant on-going major aftershock sequence amid a period of temporal clustering of seismic activity in New Zealand.

Conserved amino acid networks modulate discrete functional properties in an enzyme superfamily

In this work, we applied the sequence-based statistical coupling analysis approach to characterize conserved amino acid networks important for biochemical function in the pancreatic-type ribonuclease (ptRNase) superfamily. This superfamily-wide analysis indicates a decomposition of the RNase tertiary structure into spatially distributed yet physically connected networks of co-evolving amino acids, termed sectors. Comparison of this statistics-based description with new NMR experiments data shows that discrete amino acid networks, termed sectors, control the tuning of distinct functional properties in different enzyme homologs. Further, experimental characterization of evolutionarily distant sequences reveals that sequence variation at sector positions can distinguish homologs with a conserved dynamic pattern and optimal catalytic activity from those with altered dynamics and diminished catalytic activities. Taken together, these results provide important insights into the mechanistic design of the ptRNase superfamily, and presents a structural basis for evolutionary tuning of function in functionally diverse enzyme homologs.

Geochemical characterization of the middle and late Pleistocene alluvial fan-dominated infill of the northern part of the Weihe Basin, Central China

Major reorganizations in climate and tectonic regime occurred in East Asia during the Pleistocene, resulting in large-scale environmental changes. In this paper a detailed geochemical and mineralogical record of these changes is presented from a distal alluvial fan sedimentary sequence in the northern Weihe Basin. We established that, in addition to glacial-interglacial variation, there are three distinctly different units deposited over the past 1m.y. These units are the result of variations in the overall tectonic regime in the northern Weihe Basin.Fine-grained detrital minerals were predominantly delivered during colder climatic periods, whereas evaporative minerals were dominantly deposited during the warmer, interglacial periods, probably as a result of strong seasonal contrast. This compositional variation demonstrates the importance of climate control on hinterland erosion, surface runoff, chemical weathering and evaporation.Al-normalized ratios of indicative major elements relative to average loess composition, indicate important variations in sedimentary processes, mostly related to sediment flux. Si-enrichment is an index for past flooding events, while Fe enrichment, just like K and Ti, reflects influx of clays. In contrast, Ca and Mn are strongly enriched throughout the core, associated with the authigenic precipitation of carbonates, especially during interglacial periods.The lower (~1000–690ka) and upper (~330–0ka) units of the core are characterized by relative intense and frequent flooding, which coincided with extensive ponding in the study area. In the middle unit (~690–330ka) increased salinity levels caused by evaporation, as reflected in the high Sr/Ca ratio and dolomite abundance, led to increased carbonate precipitation. Simultaneously, the increased influx of fine sediments indicates increased clay production in the source area as a result of a more intense summer monsoon strength after 600ka.

GTP‐loading Activity of TC10/TCL Chimeras Underscores Important Allosteric Regulatory Regions of TCL

TCL/RhoJ is a Rho family GTPase that functions as a regulator of angiogenesis and may also contribute to tumor growth and metastasis in some melanomas and gastric cancers. Despite these interesting functions, the biochemistry of TCL is poorly understood. Recently our lab has investigated primary sequence differences between TCL and its closest homologs TC10 and Cdc42. TCL has an additional 20 amino acids in its N‐terminus compared to Cdc42, and deletion or mutation of amino acids 17–20 inhibit the ability of TCL to exchange guanine nucleotides. However, a similar N‐terminal deletion (ΔN) within its closest homologue, TC10, does not have a similar affect. Chimeras of TCL and TC10 identified amino acids 121–129 (designated as region R2c) of TCL as a distal sequence within TCL that influences the ability of the N‐terminus to regulate nucleotide loading. To further investigate this mode of allosteric regulation, chimeras were produced that replaced the R2c sequence of a TC10 ΔN construct, and mutation of this region was found to be sufficient to change the nucleotide binding characteristics of TC10 ΔN so it mimicked TCL ΔN, further supporting the importance of the R2c region for TCL GTP‐loading. Additional chimeras will be produced to test whether adding the N‐terminus of TCL to the TCL/TC10 R2c chimera restores nucleotide binding. Overall, the results demonstrate TCL is a unique GTPase in regard to its nucleotide exchange mechanism and may be a selectively targetable GTPase for chemotherapeutic intervention.

Synergistic effect of peptide inhibitors derived from the extracellular and intracellular domain of αIIb subunit of integrin αIIbβ3 on platelet activation and aggregation

αIIbβ3, the major platelet integrin, plays a central role in hemostasis and thrombosis. Upon platelet activation, conformation of αIIbβ3 changes and allows fibrinogen binding and, subsequently, platelet aggregation. It was previously shown that a lipid-modified platelet permeable peptide, which corresponds to the intracellular acidic membrane distal sequence 1000LEEDDEEGE1008 of αIIb (pal-K-LEEDDEEGE or pal-K-1000-1008), inhibits thrombin-induced human platelet aggregation, by inhibiting talin association with the integrin. YMESRADR, a peptide corresponding to the extracellular sequence 313–320 of αIIb, is also a potent platelet aggregation inhibitor by mimicking the effect of a clasp between the head domains of αIIb and β3. The aim of the present study was to investigate the synergistic effect of the intra- and extracellular- peptide inhibitors on platelet aggregation, as well as on the phosphorylation of two signaling proteins, focal adhesion kinase (FAK) and extracellular signal-regulated kinase (ERK). Platelet preincubation with Pal-K-LEEDDEGE followed by YMESRADR showed a synergistic inhibitory activity on platelet aggregation. Platelet incubation with threshold inhibitory concentrations of both peptides provoked almost the total inhibition of aggregation, PAC-1 binding, and fibrinogen binding, but not P-selectin exposure on activated platelets’ surface. Like RGDS peptide, this mixture inhibits FAK phosphorylation whose phosphorylation is well known to be consecutive to the aggregation (postoccupancy events). However, in contrast to RGDS peptide that enhances ERK phosphorylation and activation, the mixture of threshold inhibitory concentrations of Pal-K-LEEDDEEGE and YMESRADR inhibits ERK phosphorylation. We suggest that the use of the intracellular in combination with the extracellular peptide inhibitor, acting with a non-RGD-like mechanism, may provide an alternative way to antagonize integrin αIIbβ3 activation.

Unifying tephrostratigraphic approaches to redefine major Holocene marker tephras, Mt. Taranaki, New Zealand

In this study, geochemical fingerprinting of glass shards and titanomagnetite phenocrysts was used to match twenty complex pyroclastic deposits from the flanks of Mt. Taranaki to major tephra fall “marker beds” in medial and distal deposition sites. These correlations hinged upon identifying time-bound compositional changes (a chemostratigraphy) in distal Taranaki tephra-fall sequences preserved in lake and peat sediment records around the volcano. The current work shows that previous soil-stratigraphy based studies led to miscorrelations, because they relied upon radiocarbon dates, a “counting back” approach, and an underestimate of the number of eruptions that actually occurred in any time frame. The new tephrostratigraphy proposed at Mt. Taranaki resulted from stratigraphic rearranging of several earlier-defined units. Some tephra units are older than previously determined (e.g., Waipuku, Tariki, and Mangatoki; ~6 to 9calkaBP), while one of the most prominent Taranaki marker tephra deposit, the Korito, is shown to lie stratigraphically above a widespread rhyolitic marker bed from Taupo volcano, the Stent Tephra (also known as unit Q; ~4.3calkaBP). Pyroclastic tephra deposits previously dated between ~6 to 4calkaBP at a key tephra section, c. 40km NE of Mt. Taranaki's summit, were misidentified and are now shown to comprise new marker tephra deposits, including the Kokowai (~4.7calkaBP), which is a prominent marker horizon on the eastern flanks of the volcano. A new local proximal stratigraphy for <5calkaBP tephra units can be well correlated to tephra layers within distal lake and peat sequences, but the differences between the two records indicates an overall larger number of eruptions have occurred at this volcano than previously thought. This study additionally demonstrates the utility of titanomagnetite chemistry for discrimination and correlation of groups or sequences of tephra deposits – even if unique compositions cannot be identified.