bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of an N-terminal inhibitory extension as the primary mechanosensory regulator of twitchin kinase
Authors: von Castelmur Eleonore , Strümpfer Johan , Franke Barbara , Bogomolovas Julijus , Barbieri Sonia , Qadota Hiroshi , Konarev Petr V. , Svergun Dmitri I. , Labeit Siegfried , Benian Guy M. , Schulten Klaus , Mayans Olga
Published: August 2012

Titin-like kinases are an important class of cytoskeletal kinases that intervene in the response of muscle to mechanical stimulation, being central to myofibril homeostasis and development. These kinases exist in autoinhibited states and, allegedly, become activated during muscle activity by the elastic unfolding of a C-terminal regulatory segment (CRD). However, this mechano-activation model remains controversial. Here we explore the structural, catalytic, and tensile properties of the multidomain kinase region of
Caenorhabditis elegans
twitchin (Fn
31
-Nlinker-kinase-CRD-Ig
26
) using X-ray crystallography, small angle X-ray scattering, molecular dynamics simulations, and catalytic assays. This work uncovers the existence of an inhibitory segment that flanks the kinase N-terminally (N-linker) and that acts synergistically with the canonical CRD tail to silence catalysis. The N-linker region has high mechanical lability and acts as the primary stretch-sensor in twitchin kinase, while the CRD is poorly responsive to pulling forces. This poor response suggests that the CRD is not a generic mechanosensor in this kinase family. Instead, the CRD is shown here to be permissive to catalysis and might protect the kinase active site against mechanical damage. Thus, we put forward a regulatory model where kinase inhibition results from the combined action of both N- and C-terminal tails, but only the N-terminal extension undergoes mechanical removal, thereby affording partial activation. Further, we compare invertebrate and vertebrate titin-like kinases and identify variations in the regulatory segments that suggest a mechanical speciation of these kinase classes.
Structures of dolomite at ultrahigh pressure and their influence on the deep carbon cycle
Authors: Merlini Marco , Crichton Wilson A. , Hanfland Michael , Gemmi Mauro , Müller Harald , Kupenko Ilya , Dubrovinsky Leonid
Published: August 2012

Carbon-bearing solids, fluids, and melts in the Earth's deep interior may play an important role in the long-term carbon cycle. Here we apply synchrotron X-ray single crystal micro-diffraction techniques to identify and characterize the high-pressure polymorphs of dolomite. Dolomite-II, observed above 17 GPa, is triclinic, and its structure is topologically related to CaCO
3
-II. It transforms above 35 GPa to dolomite-III, also triclinic, which features carbon in [3 + 1] coordination at the highest pressures investigated (60 GPa). The structure is therefore representative of an intermediate between the low-pressure carbonates and the predicted ultra-high pressure carbonates, with carbon in tetrahedral coordination. Dolomite-III does not decompose up to the melting point (2,600 K at 43 GPa) and its thermodynamic stability demonstrates that this complex phase can transport carbon to depths of at least up to 1,700 km. Dolomite-III, therefore, is a likely occurring phase in areas containing recycled crustal slabs, which are more oxidized and Ca-enriched than the primitive lower mantle. Indeed, these phases may play an important role as carbon carriers in the whole mantle carbon cycling. As such, they are expected to participate in the fundamental petrological processes which, through carbon-bearing fluids and carbonate melts, will return carbon back to the Earth’s surface.
How an interacting many-body system tunnels through a potential barrier to open space
Authors: Lode Axel U.J. , Streltsov Alexej I. , Sakmann Kaspar , Alon Ofir E. , Cederbaum Lorenz S.
Published: August 2012

The tunneling process in a many-body system is a phenomenon which lies at the very heart of quantum mechanics. It appears in nature in the form of α-decay, fusion and fission in nuclear physics, and photoassociation and photodissociation in biology and chemistry. A detailed theoretical description of the decay process in these systems is a very cumbersome problem, either because of very complicated or even unknown interparticle interactions or due to a large number of constituent particles. In this work, we theoretically study the phenomenon of quantum many-body tunneling in a transparent and controllable physical system, an ultracold atomic gas. We analyze a full, numerically exact many-body solution of the Schrödinger equation of a one-dimensional system with repulsive interactions tunneling to open space. We show how the emitted particles dissociate or fragment from the trapped and coherent source of bosons: The overall many-particle decay process is a quantum interference of single-particle tunneling processes emerging from sources with different particle numbers taking place simultaneously. The close relation to atom lasers and ionization processes allows us to unveil the great relevance of many-body correlations between the emitted and trapped fractions of the wave function in the respective processes.
A quantitative version of the commutator theorem for zero trace matrices
Authors: Johnson William B. , Ozawa Narutaka , Schechtman Gideon
Published: August 2012

Let
A
be an
m
 × 
m
complex matrix with zero trace and let
ε
 > 0. Then there are
m
 × 
m
matrices
B
and
C
such that
A
 = [
B
,
C
] and ‖
B
‖‖
C
‖ ≤ 
K

ε

m

ε

‖
A
‖ where
K

ε

depends only on
ε
. Moreover, the matrix
B
can be taken to be normal.
Oncogenic
Myc
translocations are independent of chromosomal location and orientation of the immunoglobulin heavy chain locus
Authors: Spehalski Elizabeth , Kovalchuk Alexander L. , Collins John T. , Liang Genqing , Dubois Wendy , Morse Herbert. C. , Ferguson David O. , Casellas Rafael , Dunnick Wesley A.
Published: August 2012

Many tumors are characterized by recurrent translocations between a tissue-specific gene and a proto-oncogene. The juxtaposition of the Ig heavy chain gene and
Myc
in Burkitt’s lymphoma and in murine plasmacytoma is a classic example. Regulatory elements within the heavy chain constant region locus are required for
Myc
translocation and/or deregulation. However, many genes are regulated by cis-acting elements at distances up to 1,000 kb outside the locus. Such putative distal elements have not been examined for the heavy chain locus, particularly in the context of
Myc
translocations. We demonstrate that a transgene containing the Ig heavy chain constant region locus, inserted into five different chromosomal locations, can undergo translocations involving
Myc
. Furthermore, these translocations are able to generate plasmacytomas in each transgenic line. We conclude that the heavy chain constant region locus itself includes all of the elements necessary for both the translocation and the deregulation of the proto-oncogene.
Interpreting the widespread nonlinear force spectra of intermolecular bonds
Authors: Friddle Raymond W. , Noy Aleksandr , De Yoreo James J.
Published: August 2012

Single molecule force spectroscopy probes the strength, lifetime, and energetic details of intermolecular interactions in a simple experiment. A growing number of these studies have reported distinctly nonlinear trends in rupture force with loading rate that are typically explained in conventional models by invoking complex escape pathways. Recent analyses suggested that these trends should be expected even for simple barriers based on the basic assumptions of bond rupture dynamics and thus may represent the norm rather than the exception. Here we explore how these nonlinear trends reflect the two fundamental regimes of bond rupture: (
i
) a near-equilibrium regime, produced either by bond reforming in the case of a single bond or by asynchronized rupture of multiple individual bonds, and (
ii
) a kinetic regime produced by fast, non-equilibrium bond rupture. We analyze both single- and multi-bonded cases, describe the full evolution of the system as it transitions between near- and far-from-equilibrium loading regimes, and show that both interpretations produce essentially identical force spectra. Data from 10 different molecular systems show that this model provides a comprehensive description of force spectra for a diverse suite of bonds over experimentally relevant loading rates, removes the inconsistencies of previous interpretations of transition state distances, and gives ready access to both kinetic and thermodynamic information about the interaction. These results imply that single-molecule binding free energies for a vast number of bonds have already been measured.
Nuclear receptor PPARγ-regulated monoacylglycerol
O
-acyltransferase 1 (MGAT1) expression is responsible for the lipid accumulation in diet-induced hepatic steatosis
Authors: Lee Yoo Jeong , Ko Eun Hee , Kim Ji Eun , Kim Eunha , Lee Hyemin , Choi Hyeonjin , Yu Jung Hwan , Kim Hyo Jung , Seong Je-Kyung , Kim Kyung-Sup , Kim Jae-woo
Published: August 2012

Recently, hepatic peroxisome proliferator-activated receptor (PPAR)γ has been implicated in hepatic lipid accumulation. We found that the C3H mouse strain does not express PPARγ in the liver and, when subject to a high-fat diet, is resistant to hepatic steatosis, compared with C57BL/6 (B6) mice. Adenoviral PPARγ2 injection into B6 and C3H mice caused hepatic steatosis, and microarray analysis demonstrated that hepatic PPARγ2 expression is associated with genes involved in fatty acid transport and the triglyceride synthesis pathway. In particular, hepatic PPARγ2 expression significantly increased the expression of monoacylglycerol
O
-acyltransferase 1 (MGAT1). Promoter analysis by luciferase assay and electrophoretic mobility shift assay as well as chromatin immunoprecipitation assay revealed that PPARγ2 directly regulates the MGAT1 promoter activity. The MGAT1 overexpression in cultured hepatocytes enhanced triglyceride synthesis without an increase of PPARγ expression. Importantly, knockdown of MGAT1 in the liver significantly reduced hepatic steatosis in 12-wk-old high-fat–fed mice as well as ob/ob mice, accompanied by weight loss and improved glucose tolerance. These results suggest that the MGAT1 pathway induced by hepatic PPARγ is critically important in the development of hepatic steatosis during diet-induced obesity.
BRCA1 and HSP90 cooperate in homologous and non-homologous DNA double-strand-break repair and G2/M checkpoint activation
Authors: Stecklein Shane R. , Kumaraswamy Easwari , Behbod Fariba , Wang Wenjia , Chaguturu Vamsee , Harlan-Williams Lisa M. , Jensen Roy A.
Published: August 2012

Expression of functional breast cancer susceptibility gene 1 (BRCA1) in human breast and ovarian cancers is associated with resistance to platinum-based chemotherapeutics and poly(ADP ribose) polymerase (PARP) inhibitors. BRCA1 is a nuclear tumor suppressor that is critical for resolving double-strand DNA breaks (DSBs) and interstrand crosslinks (ICLs) by homologous recombination (HR). In vitro, animal and human clinical data have demonstrated that BRCA1-deficient cancers are highly sensitive to ICL-inducing chemotherapeutic agents, are amenable to synthetic lethal approaches that exploit defects in DSB/ICL repair, and may be associated with improved survival. Conversely, high or restored expression of BRCA1 in breast and ovarian cancer is associated with therapeutic resistance and poor prognosis. There has been much interest in identifying agents that interfere with BRCA1-dependent DSB/ICL repair to restore or enhance sensitivity to cancer therapeutics. We demonstrate that the heat-shock protein 90 (HSP90) inhibitor 17-allylamino-17-demethoxygeldanamycin [17-AAG (Tanespimycin)], currently in Phase II/III clinical evaluation for several cancers, induces BRCA1 ubiquitination and proteasomal degradation, resulting in compromised repair of ionizing radiation- and platinum-induced DNA damage. We show that loss of HSP90 function abolishes BRCA1-dependent DSB repair and that BRCA1-deficient cells are hypersensitive to 17-AAG due to impaired Gap 2/Mitosis (G2/M) checkpoint activation and resultant mitotic catastrophe. In summary, we document an upstream HSP90-dependent regulatory point in the Fanconi anemia/BRCA DSB/ICL repair pathway, illuminate the role of BRCA1 in regulating damage-associated checkpoint and repair responses to HSP90 inhibitors, and identify BRCA1 as a clinically relevant target for enhancing sensitivity in refractory and/or resistant malignancies.
Monotonicity and its analytic and geometric implications
Authors: Colding Tobias Holck , Minicozzi William P.
Published: August 2012

In this expository article, we discuss various monotonicity formulas for parabolic and elliptic operators and explain how the analysis of function spaces and the geometry of the underlining spaces are intertwined. After briefly discussing some of the well-known analytical applications of monotonicity for parabolic operators, we turn to their elliptic counterparts, their geometric meaning, and some geometric consequences.
Quantitation of ten 30S ribosomal assembly intermediates using fluorescence triple correlation spectroscopy
Authors: Ridgeway William K. , Millar David P. , Williamson James R.
Published: August 2012

The self-assembly of bacterial 30S ribosomes involves a large number of RNA folding and RNA-protein binding steps. The sequence of steps determines the overall assembly mechanism and the structure of the mechanism has ramifications for the robustness of biogenesis and resilience against kinetic traps. Thermodynamic interdependencies of protein binding inferred from omission-reconstitution experiments are thought to preclude certain assembly pathways and thus enforce ordered assembly, but this concept is at odds with kinetic data suggesting a more parallel assembly landscape. A major challenge is deconvolution of the statistical distribution of intermediates that are populated during assembly at high concentrations approaching in vivo assembly conditions. To specifically resolve the intermediates formed by binding of three ribosomal proteins to the full length 16S rRNA, we introduce Fluorescence Triple-Correlation Spectroscopy (F3CS). F3CS identifies specific ternary complexes by detecting coincident fluctuations in three-color fluorescence data. Triple correlation integrals quantify concentrations and diffusion kinetics of triply labeled species, and F3CS data can be fit alongside auto-correlation and cross-correlation data to quantify the populations of 10 specific ribosome assembly intermediates. The distribution of intermediates generated by binding three ribosomal proteins to the entire native 16S rRNA included significant populations of species that were not previously thought to be thermodynamically accessible, questioning the current interpretation of the classic omission-reconstitution experiments. F3CS is a general approach for analyzing assembly and function of macromolecular complexes, especially those too large for traditional biophysical methods.
RAB-10-GTPase–mediated regulation of endosomal phosphatidylinositol-4,5-bisphosphate
Authors: Shi Anbing , Liu Ou , Koenig Sabine , Banerjee Riju , Chen Carlos Chih-Hsiung , Eimer Stefan , Grant Barth D.
Published: August 2012

Caenorhabditis elegans
RAB-10 and mammalian Rab10 are key regulators of endocytic recycling, especially in the basolateral recycling pathways of polarized epithelial cells. To understand better how RAB-10 contributes to recycling endosome function, we sought to identify RAB-10 effectors. One RAB-10-binding partner that we identified, CNT-1, is the only
C. elegans
homolog of the mammalian Arf6 GTPase-activating proteins ACAP1 and ACAP2. Arf6 is known to regulate endosome-to-plasma membrane transport, in part through activation of type I phophatidylinositol-4-phosphate 5 kinase. Here we show that CNT-1 binds to RAB-10 through its C-terminal ankyrin repeats and colocalizes with RAB-10 and ARF-6 on recycling endosomes in vivo. Furthermore, we find that RAB-10 is required for the recruitment of CNT-1 to endosomal membranes in the intestinal epithelium. Consistent with negative regulation of ARF-6 by RAB-10 and CNT-1, we found overaccumulation of endosomal phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] in
cnt-1
and
rab-10
mutants and reduced endosomal PI(4,5)P2 levels in
arf-6
mutants. These mutants produced similar effects on endosomal recruitment of the PI(4,5)P2-dependent membrane-bending proteins RME-1/Ehd and SDPN-1/Syndapin/Pacsin and resulted in endosomal trapping of specific recycling cargo. Our studies identify a RAB-10–to–ARF-6 regulatory loop required to regulate endosomal PI(4,5)P2, a key phosphoinositide in membrane traffic.
General and inducible hypermutation facilitate parallel adaptation in
Pseudomonas aeruginosa
despite divergent mutation spectra
Authors: Weigand Michael R. , Sundin George W.
Published: August 2012

The successful growth of hypermutator strains of bacteria contradicts a clear preference for lower mutation rates observed in the microbial world. Whether by general DNA repair deficiency or the inducible action of low-fidelity DNA polymerases, the evolutionary strategies of bacteria include methods of hypermutation. Although both raise mutation rate, general and inducible hypermutation operate through distinct molecular mechanisms and therefore likely impart unique adaptive consequences. Here we compare the influence of general and inducible hypermutation on adaptation in the model organism
Pseudomonas aeruginosa
PAO1 through experimental evolution. We observed divergent spectra of single base substitutions derived from general and inducible hypermutation by sequencing
rpoB
in spontaneous rifampicin-resistant (Rif
R
) mutants. Likewise, the pattern of mutation in a draft genome sequence of a derived inducible hypermutator isolate differed from those of general hypermutators reported in the literature. However, following experimental evolution, populations of both mutator types exhibited comparable improvements in fitness across varied conditions that differed from the highly specific adaptation of nonmutators. Our results suggest that despite their unique mutation spectra, general and inducible hypermutation can analogously influence the ecology and adaptation of bacteria, significantly shaping pathogenic populations where hypermutation has been most widely observed.
Long-range exciton dissociation in organic solar cells
Authors: Caruso Domenico , Troisi Alessandro
Published: August 2012

It is normally assumed that electrons and holes in organic solar cells are generated by the dissociation of excitons at the interface between donor and acceptor materials in strongly bound hole-electron pairs. We show in this contribution that excitons can dissociate tens of angstroms away from the interface and generate partially separated electrons and holes, which can more easily overcome their coulombic attraction and form free charges. We first establish under what conditions long-range exciton dissociation is likely (using a kinetic model and a microscopic model for the calculation of the long-range electron transfer rate). Then, defining a rather general model Hamiltonian for the donor material, we show that the phenomenon is extremely common in the majority of polymer:fullerene bulk heterojunction solar cells.
Rational, combinatorial, and genomic approaches for engineering L-tyrosine production in
Escherichia coli
Authors: Santos Christine Nicole S. , Xiao Wenhai , Stephanopoulos Gregory
Published: August 2012

Although microbial metabolic engineering has traditionally relied on rational and knowledge-driven techniques, significant improvements in strain performance can be further obtained through the use of combinatorial approaches exploiting phenotypic diversification and screening. Here, we demonstrate the combined use of global transcriptional machinery engineering and a high-throughput L-tyrosine screen towards improving L-tyrosine production in
Escherichia coli
. This methodology succeeded in generating three strains from two separate mutagenesis libraries (
rpoA
and
rpoD
) exhibiting up to a 114% increase in L-tyrosine titer over a rationally engineered parental strain with an already high capacity for production. Subsequent strain characterization through transcriptional analysis and whole genome sequencing allowed complete phenotype reconstruction from well-defined mutations and point to important roles for both the acid stress resistance pathway and the stringent response of
E. coli
in imparting this phenotype. As such, this study presents one of the first examples in which cell-wide measurements have helped to elucidate the genetic and biochemical underpinnings of an engineered cellular property, leading to the total restoration of metabolite overproduction from specific chromosomal mutations.
Role of satellite cells versus myofibers in muscle hypertrophy induced by inhibition of the myostatin/activin signaling pathway
Authors: Lee Se-Jin , Huynh Thanh V. , Lee Yun-Sil , Sebald Suzanne M. , Wilcox-Adelman Sarah A. , Iwamori Naoki , Lepper Christoph , Matzuk Martin M. , Fan Chen-Ming
Published: August 2012

Myostatin and activin A are structurally related secreted proteins that act to limit skeletal muscle growth. The cellular targets for myostatin and activin A in muscle and the role of satellite cells in mediating muscle hypertrophy induced by inhibition of this signaling pathway have not been fully elucidated. Here we show that myostatin/activin A inhibition can cause muscle hypertrophy in mice lacking either syndecan4 or Pax7, both of which are important for satellite cell function and development. Moreover, we show that muscle hypertrophy after pharmacological blockade of this pathway occurs without significant satellite cell proliferation and fusion to myofibers and without an increase in the number of myonuclei per myofiber. Finally, we show that genetic ablation of
Acvr2b
, which encodes a high-affinity receptor for myostatin and activin A specifically in myofibers is sufficient to induce muscle hypertrophy. All of these findings are consistent with satellite cells playing little or no role in myostatin/activin A signaling in vivo and render support that inhibition of this signaling pathway can be an effective therapeutic approach for increasing muscle growth even in disease settings characterized by satellite cell dysfunction.
ERK1/2 activation is a therapeutic target in age-related macular degeneration
Authors: Dridi Sami , Hirano Yoshio , Tarallo Valeria , Kim Younghee , Fowler Benjamin J. , Ambati Balamurali K. , Bogdanovich Sasha , Chiodo Vince A. , Hauswirth William W. , Kugel Jennifer F. , Goodrich James A. , Ponicsan Steven L. , Hinton David R. , Kleinman Mark E. , Baffi Judit Z. , Gelfand Bradley D. , Ambati Jayakrishna
Published: August 2012

Deficient expression of the RNase III DICER1, which leads to the accumulation of cytotoxic Alu RNA, has been implicated in degeneration of the retinal pigmented epithelium (RPE) in geographic atrophy (GA), a late stage of age-related macular degeneration that causes blindness in millions of people worldwide. Here we show increased extracellular-signal-regulated kinase (ERK) 1/2 phosphorylation in the RPE of human eyes with GA and that RPE degeneration in mouse eyes and in human cell culture induced by DICER1 depletion or Alu RNA exposure is mediated via ERK1/2 signaling. Alu RNA overexpression or DICER1 knockdown increases ERK1/2 phosphorylation in the RPE in mice and in human cell culture. Alu RNA-induced RPE degeneration in mice is rescued by intravitreous administration of PD98059, an inhibitor of the ERK1/2-activating kinase MEK1, but not by inhibitors of other MAP kinases such as p38 or JNK. These findings reveal a previously unrecognized function of ERK1/2 in the pathogenesis of GA and provide a mechanistic basis for evaluation of ERK1/2 inhibition in treatment of this disease.
Resolution of ray-finned fish phylogeny and timing of diversification
Authors: Near Thomas J. , Eytan Ron I. , Dornburg Alex , Kuhn Kristen L. , Moore Jon A. , Davis Matthew P. , Wainwright Peter C. , Friedman Matt , Smith W. Leo
Published: August 2012

Ray-finned fishes make up half of all living vertebrate species. Nearly all ray-finned fishes are teleosts, which include most commercially important fish species, several model organisms for genomics and developmental biology, and the dominant component of marine and freshwater vertebrate faunas. Despite the economic and scientific importance of ray-finned fishes, the lack of a single comprehensive phylogeny with corresponding divergence-time estimates has limited our understanding of the evolution and diversification of this radiation. Our analyses, which use multiple nuclear gene sequences in conjunction with 36 fossil age constraints, result in a well-supported phylogeny of all major ray-finned fish lineages and molecular age estimates that are generally consistent with the fossil record. This phylogeny informs three long-standing problems: specifically identifying elopomorphs (eels and tarpons) as the sister lineage of all other teleosts, providing a unique hypothesis on the radiation of early euteleosts, and offering a promising strategy for resolution of the “bush at the top of the tree” that includes percomorphs and other spiny-finned teleosts. Contrasting our divergence time estimates with studies using a single nuclear gene or whole mitochondrial genomes, we find that the former underestimates ages of the oldest ray-finned fish divergences, but the latter dramatically overestimates ages for derived teleost lineages. Our time-calibrated phylogeny reveals that much of the diversification leading to extant groups of teleosts occurred between the late Mesozoic and early Cenozoic, identifying this period as the “Second Age of Fishes.”
Hydrogen-limited growth of hyperthermophilic methanogens at deep-sea hydrothermal vents
Authors: Ver Eecke Helene C. , Butterfield David A. , Huber Julie A. , Lilley Marvin D. , Olson Eric J. , Roe Kevin K. , Evans Leigh J. , Merkel Alexandr Y. , Cantin Holly V. , Holden James F.
Published: August 2012

Microbial productivity at hydrothermal vents is among the highest found anywhere in the deep ocean, but constraints on microbial growth and metabolism at vents are lacking. We used a combination of cultivation, molecular, and geochemical tools to verify pure culture H2threshold measurements for hyperthermophilic methanogenesis in low-temperature hydrothermal fluids from Axial Volcano and Endeavour Segment in the northeastern Pacific Ocean. TwoMethanocaldococcusstrains from Axial andMethanocaldococcus jannaschiishowed similar Monod growth kinetics when grown in a bioreactor at varying H2concentrations. Their H2half-saturation value was 66 μM, and growth ceased below 17–23 μM H2, 10-fold lower than previously predicted. By comparison, measured H2and CH4concentrations in fluids suggest that there was generally sufficient H2forMethanocaldococcusgrowth at Axial but not at Endeavour. Fluids from one vent at Axial (Marker 113) had anomalously high CH4concentrations and contained various thermal classes of methanogens based on cultivation andmcrA/mrtAanalyses. At Endeavour, methanogens were largely undetectable in fluid samples based on cultivation and molecular screens, although abundances of hyperthermophilic heterotrophs were relatively high. Where present,Methanocaldococcusgenes were the predominantmcrA/mrtAsequences recovered and comprised ∼0.2–6% of the total archaeal community. Field and coculture data suggest that H2limitation may be partly ameliorated by H2syntrophy with hyperthermophilic heterotrophs. These data support our estimated H2threshold for hyperthermophilic methanogenesis at vents and highlight the need for coupled laboratory and field measurements to constrain microbial distribution and biogeochemical impacts in the deep sea.
Mass spectrometry study of a transferrin-based protein drug reveals the key role of protein aggregation for successful oral delivery
Authors: Bobst Cedric E. , Wang Shunhai , Shen Wei-Chiang , Kaltashov Igor A.
Published: August 2012

A recently designed human growth hormone/transferrin fusion protein (GHT) remains one of the very few examples of a protein capable of eliciting measurable therapeutic response after oral administration. To better understand the underlying factors that resulted in this rare success of nonparenteral protein drug delivery, we analyzed proteolytic stability and receptor binding properties of this protein, the key factors in overcoming the primary barriers to successful oral delivery. Analysis of GHT by a combination of size exclusion chromatography and mass spectrometry revealed that a significant protein population exists in an oligomeric (GHTx) state in addition to the anticipated monomer (GHT1). These states of GHT were evaluated for their survivability in stomach-like conditions, as well as their ability to bind transferrin receptor (TfR). Our results reveal an exceptional stability of GHTx, as well as the preserved ability to bind TfR, a critical first step in crossing the epithelial–intestinal barrier through receptor-mediated transcytosis.
The Ser/Thr protein kinase AfsK regulates polar growth and hyphal branching in the filamentous bacteria
Streptomyces
Authors: Hempel Antje M. , Cantlay Stuart , Molle Virginie , Wang Sheng-Bing , Naldrett Mike J. , Parker Jennifer L. , Richards David M. , Jung Yong-Gyun , Buttner Mark J. , Flärdh Klas
Published: August 2012

In cells that exhibit apical growth, mechanisms that regulate cell polarity are crucial for determination of cellular shape and for the adaptation of growth to intrinsic and extrinsic cues. Broadly conserved pathways control cell polarity in eukaryotes, but less is known about polarly growing prokaryotes. An evolutionarily ancient form of apical growth is found in the filamentous bacteria
Streptomyces
, and is directed by a polarisome-like complex involving the essential protein DivIVA. We report here that this bacterial polarization machinery is regulated by a eukaryotic-type Ser/Thr protein kinase, AfsK, which localizes to hyphal tips and phosphorylates DivIVA. During normal growth, AfsK regulates hyphal branching by modulating branch-site selection and some aspect of the underlying polarisome-splitting mechanism that controls branching of
Streptomyces
hyphae. Further, AfsK is activated by signals generated by the arrest of cell wall synthesis and directly communicates this to the polarisome by hyperphosphorylating DivIVA. Induction of high levels of DivIVA phosphorylation by using a constitutively active mutant AfsK causes disassembly of apical polarisomes, followed by establishment of multiple hyphal branches elsewhere in the cell, revealing a profound impact of this kinase on growth polarity. The function of AfsK is reminiscent of the phoshorylation of polarity proteins and polarisome components by Ser/Thr protein kinases in eukaryotes.
Structural insights into neuronal K
+
channel–calmodulin complexes
Authors: Mruk Karen , Shandilya Shiven M.D. , Blaustein Robert O. , Schiffer Celia A. , Kobertz William R.
Published: August 2012

Calmodulin (CaM) is a ubiquitous intracellular calcium sensor that directly binds to and modulates a wide variety of ion channels. Despite the large repository of high-resolution structures of CaM bound to peptide fragments derived from ion channels, there is no structural information about CaM bound to a fully folded ion channel at the plasma membrane. To determine the location of CaM docked to a functioning KCNQ K
+
channel, we developed an intracellular tethered blocker approach to measure distances between CaM residues and the ion-conducting pathway. Combining these distance restraints with structural bioinformatics, we generated an archetypal quaternary structural model of an ion channel–CaM complex in the open state. These models place CaM close to the cytoplasmic gate, where it is well positioned to modulate channel function.
Sequencing papaya X and Y
h
chromosomes reveals molecular basis of incipient sex chromosome evolution
Authors: Wang Jianping , Na Jong-Kuk , Yu Qingyi , Gschwend Andrea R. , Han Jennifer , Zeng Fanchang , Aryal Rishi , VanBuren Robert , Murray Jan E. , Zhang Wenli , Navajas-Pérez Rafael , Feltus F. Alex , Lemke Cornelia , Tong Eric J. , Chen Cuixia , Man Wai Ching , Singh Ratnesh , Wang Ming-Li , Min Xiang Jia , Alam Maqsudul , Charlesworth Deborah , Moore Paul H. , Jiang Jiming , Paterson Andrew H. , Ming Ray
Published: August 2012

Sex determination in papaya is controlled by a recently evolved XY chromosome pair, with two slightly different Y chromosomes controlling the development of males (Y) and hermaphrodites (Y
h
). To study the events of early sex chromosome evolution, we sequenced the hermaphrodite-specific region of the Y
h
chromosome (HSY) and its X counterpart, yielding an 8.1-megabase (Mb) HSY pseudomolecule, and a 3.5-Mb sequence for the corresponding X region. The HSY is larger than the X region, mostly due to retrotransposon insertions. The papaya HSY differs from the X region by two large-scale inversions, the first of which likely caused the recombination suppression between the X and Y
h
chromosomes, followed by numerous additional chromosomal rearrangements. Altogether, including the X and/or HSY regions, 124 transcription units were annotated, including 50 functional pairs present in both the X and HSY. Ten HSY genes had functional homologs elsewhere in the papaya autosomal regions, suggesting movement of genes onto the HSY, whereas the X region had none. Sequence divergence between 70 transcripts shared by the X and HSY revealed two evolutionary strata in the X chromosome, corresponding to the two inversions on the HSY, the older of which evolved about 7.0 million years ago. Gene content differences between the HSY and X are greatest in the older stratum, whereas the gene content and order of the collinear regions are identical. Our findings support theoretical models of early sex chromosome evolution.
Oligomycin frames a common drug-binding site in the ATP synthase
Authors: Symersky Jindrich , Osowski Daniel , Walters D. Eric , Mueller David M.
Published: August 2012

We report the high-resolution (1.9 Å) crystal structure of oligomycin bound to the subunit c
10
ring of the yeast mitochondrial ATP synthase. Oligomycin binds to the surface of the c
10
ring making contact with two neighboring molecules at a position that explains the inhibitory effect on ATP synthesis. The carboxyl side chain of Glu59, which is essential for proton translocation, forms an H-bond with oligomycin via a bridging water molecule but is otherwise shielded from the aqueous environment. The remaining contacts between oligomycin and subunit c are primarily hydrophobic. The amino acid residues that form the oligomycin-binding site are 100% conserved between human and yeast but are widely different from those in bacterial homologs, thus explaining the differential sensitivity to oligomycin. Prior genetics studies suggest that the oligomycin-binding site overlaps with the binding site of other antibiotics, including those effective against
Mycobacterium tuberculosis
, and thereby frames a common “drug-binding site.” We anticipate that this drug-binding site will serve as an effective target for new antibiotics developed by rational design.
A search for thermal excursions from ancient extraterrestrial impacts using Hadean zircon Ti-U-Th-Pb depth profiles
Authors: Abbott Sunshine S. , Harrison T. Mark , Schmitt Axel K. , Mojzsis Stephen J.
Published: August 2012

Few terrestrial localities preserve more than a trace lithic record prior toca.3.8 Ga greatly limiting our understanding of the first 700 Ma of Earth history, a period inferred to have included a spike in the bolide flux to the inner solar system atca.3.85–3.95 Ga (the Late Heavy Bombardment, LHB). An accessible record of this era may be found in Hadean detrital zircons from the Jack Hills, Western Australia, in the form of μm-scale epitaxial overgrowths. By comparing crystallization temperatures of pre-3.8 Ga zircon overgrowths to the archive of zircon temperature spectra, it should, in principle, be possible to identify a distinctive impact signature. We have developed Ti-U-Th-Pb ion microprobe depth profiling to obtain age and temperature information within these zircon overgrowths and undertaken a feasibility study of its possible use in identifying impact events. Of eight grains profiled in this fashion, four have overgrowths of LHB-era age. Age vs. temperature profiles reveal a period betweenca.3.85–3.95 Ga (i.e., LHB era) characterized by significantly higher temperatures (approximately 840–875 °C) than do older or younger zircons or zircon domains (approximately 630–750 °C). However, temperatures approaching 900 °C can result in Pb isotopic exchange rendering interpretation of these profiles nonunique. Coupled age-temperature depth profiling shows promise in this role, and the preliminary data we report could represent the first terrestrial evidence for impact-related heating during the LHB.
Protection from the acquisition of
Staphylococcus aureus
nasal carriage by cross-reactive antibody to a pneumococcal dehydrogenase
Authors: Lijek Rebeccah S. , Luque Santiago L. , Liu Qian , Parker Dane , Bae Taeok , Weiser Jeffrey N.
Published: August 2012

Nasal colonization by
Staphylococcus aureus
is the major risk factor for disease and transmission. Epidemiological studies have reported a reduced risk of
S. aureus
carriage in immunocompetent but not in immunocompromised children colonized by
Streptococcus pneumoniae
. We investigate the hypothesis that the immune response to pneumococcal colonization affects
S. aureus
colonization. We demonstrate that pneumococcal colonization in mice inhibits subsequent
S. aureus
acquisition in an antibody-dependent manner and elicits antibody that cross-reacts with
S. aureus
. We identify the staphylococcal target of cross-reactive antibody as 1-pyrroline-5-carboxylate dehydrogenase (P5CDH), and the homologous immunogen in
S. pneumoniae
as SP_1119, both of which are conserved dehydrogenases. These antigens are necessary and sufficient to inhibit the acquisition of
S. aureus
colonization in a mouse model. Our findings demonstrate that immune-mediated cross-reactivity between
S. pneumoniae
and
S. aureus
protects against
S. aureus
nasal acquisition and thus reveal a paradigm for identifying protective antigens against
S. aureus
.
Ritual Black Drink consumption at Cahokia
Authors: Crown Patricia L. , Emerson Thomas E. , Gu Jiyan , Hurst W. Jeffrey , Pauketat Timothy R. , Ward Timothy
Published: August 2012

Chemical analyses of organic residues in fragments of pottery from the large site of Cahokia and surrounding smaller sites in Illinois reveal theobromine, caffeine, and ursolic acid, biomarkers for species of
Ilex
(holly) used to prepare the ritually important Black Drink. As recorded during the historic period, men consumed Black Drink in portions of the American Southeast for ritual purification. This first demonstrated discovery of biomarkers for
Ilex
occurs in beaker vessels dating between A.D. 1050 and 1250 from Cahokia, located far north of the known range of the holly species used to prepare Black Drink during historic times. The association of
Ilex
and beaker vessels indicates a sustained ritual consumption of a caffeine-laced drink made from the leaves of plants grown in the southern United States.
IgG1
+
ovalbumin-specific B-cell transnuclear mice show class switch recombination in rare allelically included B cells
Authors: Dougan Stephanie K. , Ogata Souichi , Hu Chih-Chi Andrew , Grotenbreg Gijsbert M. , Guillen Eduardo , Jaenisch Rudolf , Ploegh Hidde L.
Published: August 2012

We used somatic cell nuclear transfer (SCNT) to generate a mouse from the nucleus of an IgG1
+
ovalbumin-specific B cell. The resulting OBI mice show generally normal B-cell development, with elevated percentages of marginal zone B cells and a reduction in B-1 B cells. Whereas OBI RAG1
−/−
mice have exclusively IgG1 anti-ovalbumin in their serum, OBI mice show elevated levels of anti-ovalbumin of nearly all isotypes 3′ of the γ1 constant region in the IgH locus, indicating that class switch recombination (CSR) occurs in the absence of immunization with ovalbumin. This CSR is associated with the presence of IgM
+
IgG1
+
double producer B cells that represent <1% of total B cells, accumulate in the peritoneal cavity, and account for near-normal levels of serum IgM and IgG3.
Reprogramming IgH isotype-switched B cells to functional-grade induced pluripotent stem cells
Authors: Wesemann Duane R. , Portuguese Andrew J. , Magee Jennifer M. , Gallagher Michael P. , Zhou Xiaolong , Panchakshari Rohit A. , Alt Frederick W.
Published: August 2012

Induced pluripotent stem cells (iPSCs) can be formed from somatic cells by a defined set of genetic factors; however, aberrant epigenetic silencing of the imprintedDlk1-Dio3gene cluster often hinders their developmental potency and ability to contribute to high-grade chimerism in mice. Here, we describe an approach that allows splenic B cells activated to undergo Ig heavy-chain (IgH) class-switch recombination (CSR) to be reprogrammed into iPSCs that contribute to high-grade chimerism in mice. Treatment of naïve splenic B cells in culture with anti-CD40 plus IL-4 inducesIgHCSR from IgM to IgG1 and IgE. CSR leads to irreversibleIgHlocus deletions wherein the IgM-producing Cμ exons are permanently excised from the B-cell genome. We find that anti-CD40 plus IL-4–activated B cells produce iPSCs that are uniformly hypermethylated in the imprintedDlk1-Dio3gene cluster and fail to produce chimerism in mice. However, treatment of activated B cells with the methyltransferase inhibitor 5-aza-2′-deoxycytidine before and at early stages of reprogramming attenuates hypermethylation of theDlk1-Dio3locus in resultant iPSCs and enables them to form high-grade chimerism in mice. These conditions allowed us to produce chimeric mice in which all mature B cells were derived entirely from IgG1-expressing B-cell–derived iPSCs. We conclude that culture conditions of activated B cells before and at early stages of reprogramming influence the developmental potency of resultant iPSCs.
An accumulator model for spontaneous neural activity prior to self-initiated movement
Authors: Schurger Aaron , Sitt Jacobo D. , Dehaene Stanislas
Published: August 2012

A gradual buildup of neuronal activity known as the “readiness potential” reliably precedes voluntary self-initiated movements, in the average time locked to movement onset. This buildup is presumed to reflect the final stages of planning and preparation for movement. Here we present a different interpretation of the premovement buildup. We used a leaky stochastic accumulator to model the neural decision of “when” to move in a task where there is no specific temporal cue, but only a general imperative to produce a movement after an unspecified delay on the order of several seconds. According to our model, when the imperative to produce a movement is weak, the precise moment at which the decision threshold is crossed leading to movement is largely determined by spontaneous subthreshold fluctuations in neuronal activity. Time locking to movement onset ensures that these fluctuations appear in the average as a gradual exponential-looking increase in neuronal activity. Our model accounts for the behavioral and electroencephalography data recorded from human subjects performing the task and also makes a specific prediction that we confirmed in a second electroencephalography experiment: Fast responses to temporally unpredictable interruptions should be preceded by a slow negative-going voltage deflection beginning well before the interruption itself, even when the subject was not preparing to move at that particular moment.
Spatial ordering of chromosomes enhances the fidelity of chromosome partitioning in cyanobacteria
Authors: Jain Isha H. , Vijayan Vikram , O’Shea Erin K.
Published: August 2012

Many cyanobacteria have been shown to harbor multiple chromosome copies per cell, yet little is known about the organization, replication, and segregation of these chromosomes. Here, we visualize individual chromosomes in the cyanobacterium
Synechococcus elongatus
via time-lapse fluorescence microscopy. We find that chromosomes are equally spaced along the long axis of the cell and are interspersed with another regularly spaced subcellular compartment, the carboxysome. This remarkable organization of the cytoplasm along with accurate midcell septum placement allows for near-optimal segregation of chromosomes to daughter cells. Disruption of either chromosome ordering or midcell septum placement significantly increases the chromosome partitioning error. We find that chromosome replication is both asynchronous and independent of the position of the chromosome in the cell and that spatial organization is preserved after replication. Our findings on chromosome organization, replication, and segregation in
S. elongatus
provide a basis for understanding chromosome dynamics in bacteria with multiple chromosomes.
Electron spin resonance of nitrogen-vacancy centers in optically trapped nanodiamonds
Authors: Horowitz Viva R. , Alemán Benjamín J. , Christle David J. , Cleland Andrew N. , Awschalom David D.
Published: August 2012

Using an optical tweezers apparatus, we demonstrate three-dimensional control of nanodiamonds in solution with simultaneous readout of ground-state electron-spin resonance (ESR) transitions in an ensemble of diamond nitrogen-vacancy color centers. Despite the motion and random orientation of nitrogen-vacancy centers suspended in the optical trap, we observe distinct peaks in the measured ESR spectra qualitatively similar to the same measurement in bulk. Accounting for the random dynamics, we model the ESR spectra observed in an externally applied magnetic field to enable dc magnetometry in solution. We estimate the dc magnetic field sensitivity based on variations in ESR line shapes to be approximately. This technique may provide a pathway for spin-based magnetic, electric, and thermal sensing in fluidic environments and biophysical systems inaccessible to existing scanning probe techniques.
SAMBA, a plant-specific anaphase-promoting complex/cyclosome regulator is involved in early development and A-type cyclin stabilization
Authors: Eloy Nubia B. , Gonzalez Nathalie , Van Leene Jelle , Maleux Katrien , Vanhaeren Hannes , De Milde Liesbeth , Dhondt Stijn , Vercruysse Leen , Witters Erwin , Mercier Raphaël , Cromer Laurence , Beemster Gerrit T. S. , Remaut Han , Van Montagu Marc C. E. , De Jaeger Geert , Ferreira Paulo C. G. , Inzé Dirk
Published: August 2012

The anaphase-promoting complex/cyclosome (APC/C) is a large multiprotein E3 ubiquitin ligase involved in ubiquitin-dependent proteolysis of key cell cycle regulatory proteins, including the destruction of mitotic cyclins at the metaphase-to-anaphase transition. Despite its importance, the role of the APC/C in plant cells and the regulation of its activity during cell division remain poorly understood. Here, we describe the identification of a plant-specific negative regulator of the APC/C complex, designated SAMBA. In
Arabidopsis thaliana
,
SAMBA
is expressed during embryogenesis and early plant development and plays a key role in organ size control.
Samba
mutants produced larger seeds, leaves, and roots, which resulted from enlarged root and shoot apical meristems, and, additionally, they had a reduced fertility attributable to a hampered male gametogenesis. Inactivation of
SAMBA
stabilized A2-type cyclins during early development. Our data suggest that SAMBA regulates cell proliferation during early development by targeting CYCLIN A2 for APC/C-mediated proteolysis.