bluuebunny/crossref_metadata_2025 huggingface dataset:
Foetal programming of immune competence
Authors: Cronjé P. B.
Published: December 2003

A considerable body of evidence has accumulated to indicate that nutrition and stress during gestation can programme gene expression of the immune system and the hypothalamic-pituitary-adrenal axis in the developing foetus and that these effects may persist for the lifetime of an animal. Several studies have implicated foetal programming of these axes as a cause of reduced immune competence. Low protein diets and stress during gestation have been shown to result in blunted acute phase responses, reduced thymic and splenic lymphocyte proliferation and decreased splenic natural killer cell activity in the progeny. These effects have been linked to an increased incidence of asthma in adolescent humans and increased pre-weaning morbidity and mortality in piglets. Low protein diets and stress during gestation also alter the hypothalamic-pituitary-adrenal axis of the progeny, and many studies have shown that this increases basal corticosteroid blood concentrations and results in an exaggerated response to stress. These effects have been linked to increased preweaning mortality and slower wound healing in piglets. Most of the advances in this field have been made using rodent models and few studies have been conducted with ruminants. There are no studies in any species that have examined the impact of foetal programming on susceptibility to parasites. It is suggested that foetal programming may represent a fruitful avenue for improvement of the resistance of sheep to parasites.
Impact of non-protein nitrogen supplements on nematode infected sheep
Authors: Knox M. R.
Published: December 2003

Inadequate nutrition and gastrointestinal nematode parasitism are major constraints to livestock production. In livestock rearing enterprises that rely on low quality roughage for the provision of digestible carbohydrate, the most critical nutritional deficiency is often nitrogen. The provision of non-protein nitrogen in the diet can compensate for this deficiency and enable increased productivity from the available feed resource. Numerous studies have shown that increasing the supply of protein for intestinal absorption can alleviate the detrimental effects of nematode parasites on production and can also result in improved protective immunological responses to infection. In many situations, the practical implementation of such a strategy for improved nematode parasite control is hampered by the high cost and/or unavailability of high quality protein sources for use as ruminant livestock feed. Evidence is presented from studies with young sheep to demonstrate that supplementation with urea can achieve similar qualitative benefits in reducing the effects and level of infection with parasitic nematodes to that achieved with protein supplements. Urea–molasses blocks are a popular low-cost means of delivering non-protein nitrogen and experimental evidence shows that the use of urea–molasses blocks can have a beneficial impact on enhancing the resilience and resistance of sheep to infection with nematode parasites. Consistent with other nutritional studies, urea appeared to confer increased benefits proportional to increasing supplement intake as shown by reduced pathological effects, parasite numbers and faecal egg output. It is suggested that application of these findings in Australian pasture systems may require revision of current systems for the delivery of non-protein nitrogen supplements to increase the level of urea consumed.
Potential impact of nematode parasitism on nutrient partitioning forwool production, growth and reproduction in sheep
Authors: Liu S. M. , Masters D. G. , Adams N. R.
Published: December 2003

Most ovine studies indicate that protein supply is a key determinant of the ability to mount a strong immunity-associated response to gastrointestinal parasites. This paper quantifies the demands placed on protein supply by various physiological states, to determine why this is so. Protein requirements in fast growing lambs and ewes in late pregnancy and early lactation are substantially increased, amounting to a 1.43-, 0.69- and 2.50-fold increase in maintenance requirement for dietary protein. Infestation with internal parasites may result in a total net loss of about 12 g protein/day, which consists of a net loss of about 6 g from the gastrointestinal tract and a further loss of about 6 g through increased amino acid oxidation in the body. This loss from the infection accounts for about 0.57-, 0.71-, 0.14- and 0.77-fold the protein requirements respectively for body growth, pregnancy, lactation and wool production, and creates strong competition for protein. Such considerable demands for protein by activation of immunity-associated response diminishes the animal's nutrient status in late pregnancy and particularly in early lactation. The total demand for protein cannot be met by pastures during the reproductive season in Mediterranean environments, and it is this nutrient deficiency, relative to the demand of sheep, which results in weak immune responses over these periods. The immunity-associated response requires more protein rather than energy, particularly sulfur amino acids such as cysteine and probably methionine. Therefore, parasite infection has a more severe effect on wool- than meat-producing sheep. Body condition of sheep could be a predominant factor in determining immune status and current intake and quality of pastures needs to be considered to further boost immunity-associated response. Wool- and meat-producing genotypes, and parasite-resistant sheep in particular, may have different interactions with nutrition in response to parasite infection. These factors need to be integrated with anthelmintic treatment to establish an effective strategy to control parasite infection in Mediterranean environments.
Validation of age and growth estimates using thin otolith sections from the purple wrasse, Notolabrus fucicola
Authors: Ewing Graeme P. , Welsford Dirk C. , Jordan Alan R. , Buxton Colin
Published: December 2003

The age of Notolabrus fucicola from the east and southeast coasts of Tasmania was estimated from counts of opaque (transmitted light) growth zones in thin transverse sections of sagittal otoliths. The position of the first annulus was validated through the otolith radii of known-age juveniles. Marginal increment analysis was performed by examining the growing edge of otoliths over 2 years. This showed that opaque zones were formed annually and were consistently deposited from late September to early December (spring season), which is coincident with annual increases in somatic growth and water temperature during this season. A criterion was developed to ensure accurate age estimation for individuals sampled during the time of year when deposition of opaque material was incomplete or not apparent. The maximum age estimated was 20 years and the relationship between fork length and age was described by the von Bertalanffy growth function (L∞ = 368 mm, K = 0.116 and t0 = –1.87 for males and L∞ = 385.7 mm, K = 0.109 and t0 = –1.96 for females). The clarity of increment structure and rigorous validation protocol that was adopted minimised the risks associated with using marginal increment analysis to validate the periodicity of increment formation and it provided robust age and growth estimates for N. fucicola in Tasmanian waters.
Validation of the ageing of 0+ carp (Cyprinus carpio L.)
Authors: Smith Benjamin B. , Walker Keith F.
Published: December 2003

Forty carp larvae were reared from eggs spawned in the Torrens River, Adelaide, South Australia, and their otoliths were examined at the time of hatching or at 6, 10, 15 or 20 days after hatching (post-hatch). Using light- and scanning electron microscopy (SEM), otolith increments were counted and compared with known post-hatch ages. Typically, the counts were one day (range 0–2 days) more than the known post-hatch age, but increment formation was daily to at least age 20 days. Thus, age estimates derived from otolith increment counts of wild-caught 0+ carp should be reduced by one day. Comparison with SEM data showed that light microscopy alone offers sufficient resolution for ageing 0+ carp.
Validation of otolith age determination in Australian longfinned river eels, Anguilla reinhardtii
Authors: Pease Bruce C. , Reynolds Darren P. , Walsh Christopher T.
Published: December 2003

The primary objective of the present study was to validate the annual nature of growth rings in otoliths of 'yellow eel stage' (fully pigmented, premigratory) Australian longfinned eels (Anguilla reinhardtii) in New South Wales, Australia, using a combination of laboratory and field experiments. Eels were injected with oxytetracycline (OTC) and tagged with external 'T-bar' tags. Microscopic examination of thin transverse sections of the sagittal otoliths from recaptured eels showed that one opaque annulus was typically formed in the otolith during each year subsequent to OTC marking. The seasonal timing of opaque annulus formation was highly variable, but generally occurred between May and November. Supernumerary (false or incomplete) rings were observed in many of the otoliths. Examination of otolith sections from a sample of very small, untagged yellow eels verifed the age at first annual increment formation. Marked and tagged longfinned eels did not have a significantly higher mortality rate than controls in the laboratory experiment, but tag loss rates may be high. The field study also indicates that Australian longfinned eels generally have a very restricted home range of 300 m or less. This study is believed to provide the first otolith age validation of a tropical anguillid.
An account of the Hapalidiaceae (Corallinales, Rhodophyta) in south-eastern Australia
Authors: Harvey Adele S. , Woelkerling Wm J. , Millar Alan J. K.
Published: December 2003

This paper provides the first monographic account of south-eastern Australian representatives of the Hapalidiaceae (Corallinales, Rhodophyta). The Hapalidiaceae includes those Corallinales whose tetrasporangia produce zonately arranged spores, and whose tetrasporangia/bisporangia are borne in conceptacles, produce apical plugs, develop beneath multiporate plates or roofs, and are not enclosed individually within calcified sporangial compartments. The Hapalidiaceae contains the subfamilies Choreonematoideae, Melobesioideae and Austrolithoideae, all formerly placed in the Corallinaceae sensu lato. The Choreonematoideae is represented in south-eastern Australia by a single species, Choreonema thuretii. The Melobesioideae is represented by five genera (Melobesia, Lithothamnion, Phymatolithon, Mesophyllum, Synarthrophyton) and eight species. Synarthrophyton pseudosorus sp. nov. is newly described. The presence of tetrasporangial conceptacles occurring in irregularly shaped dense clusters that contain both discrete conceptacles and fused groups of conceptacles delimits this species from all other species of Synarthrophyton. Morphological and anatomical accounts are provided, including keys, information on distribution, habitat and nomenclature. Brief biogeographic comparisons between south-eastern Australia and neighbouring regions are also made.
Catalogue of the marine benthic macroalgae of the Fiji Islands, South Pacific
Authors: South G. Robin , Skelton Posa A.
Published: December 2003

A catalogue of the marine benthic macroalgae of Fiji is provided. All records are fully referenced, and type localities and local distributions (accompanied by listings of voucher specimens) are provided. Taxonomic notes are also included where appropriate. Records from SUVA-A, UC and BISH examined by the authors, together with updates from recent publications, form the substance of the report. A total of 448 taxa are included, comprising 136 Chlorophyta, 46 Phaeophyta and 266 Rhodophyta. This represents an increase of 116 new records since 1996, of which 39 are newly reported by the authors, and the remainder are recently published records from the Suva Lagoon (29) and the Great Astrolabe Reef and Lagoon (48). Also included are a total of 36 nomenclatural changes made since 1996, a compilation of 33 uncertain records, and a list of 34 species with type localities in Fiji, Samoa and Tonga. Apart from the far more extensive region of Micronesia, the flora of Fiji is the richest in the south and central Pacific Islands.
Sex- and age-related variations in cranial measurements and suture closure in the Australian sea lion, Neophoca cinerea (Peron, 1816)
Authors: Tedman R.
Published: December 2003

A total of 65 skulls of the Australian sealion, Neophoca cinerea, was examined to investigate the extent to which sexual dimorphism is reflected in cranial dimensions (n = 32) and skull growth, and to determine whether cranial sutures (n = 18) can be useful in age determination. All adult skull dimensions studied display significant sexual dimorphism. Skull growth ceases close to 4–7 years of age for females (Suture Fusion Rating, SFR 25–34) but skull growth in males continues until at least 16 years of age. In animals with a SFR ≥� 25, male skulls have a minimum condylobasal length of 272 mm, whereas female skulls have a maximum condylobasal length of 259 mm. The relatively early closure of the cranial vault sutures (cessation of brain growth) is balanced by the continued growth of the bony projections that provide muscle attachment (e.g. mastoid width). The later fusion of the snout and palate sutures corresponds with the continued growth of the snout and palate to match the prolonged growth of the mandibles. The upper sixth postcanine tooth was present in 43% of the adult female skulls, but only 15% of the adult male skulls. The data suggest that it may be possible to determine age(s) from examination of the sequence of fusion of cranial sutures as well as by calculation of an overall suture fusion rating for the skull.
Breeding ecology and bias in offspring sex ratio in little grassbirds (Megalurus gramineus)
Authors: McIntosh Rebecca R. , Kats Romke , Berg Mathew , Komdeur Jan , Elgar Mark A.
Published: December 2003

Little grassbirds (Megalurus gramineus) are small, sexually monomorphic passerines that live in reed beds, lignum swamps and salt marshes in southern Australia. The breeding biology and patterns of sex allocation of the little grassbird were investigated over a single breeding season. Our observations of this species in the Edithvale Wetland Reserve revealed a highly male-biased population sex ratio, with some breeding territories containing several additional males. Nevertheless, there was little compelling evidence that little grassbirds breed cooperatively. The growth rates of male and female nestlings were similar and, as predicted by theory, there was no overall primary sex ratio bias. However, the primary sex ratio was female-biased early in the breeding season and became increasingly male-biased later in the breeding season.
Myoglobin forms amyloid fibrils by association of unfolded polypeptide segments
Authors: Fändrich Marcus , Forge Vincent , Buder Katrin , Kittler Marlis , Dobson Christopher M. , Diekmann Stephan
Published: December 2003

Observations that β-sheet proteins form amyloid fibrils under at least partially denaturing conditions has raised questions as to whether these fibrils assemble by docking of preformed β-structure or by association of unfolded polypeptide segments. By using α-helical protein apomyoglobin, we show that the ease of fibril assembly correlates with the extent of denaturation. By contrast, monomeric β-sheet intermediates could not be observed under the conditions of fibril formation. These data suggest that amyloid fibril formation from apomyoglobin depends on disordered polypeptide segments and conditions that are selectively unfavorable to folding. However, it is inevitable that such conditions often stabilize protein folding intermediates.
Markov chain Monte Carlo without likelihoods
Authors: Marjoram Paul , Molitor John , Plagnol Vincent , Tavaré Simon
Published: December 2003

Many stochastic simulation approaches for generating observations from a posterior distribution depend on knowing a likelihood function. However, for many complex probability models, such likelihoods are either impossible or computationally prohibitive to obtain. Here we present a Markov chain Monte Carlo method for generating observations from a posterior distribution without the use of likelihoods. It can also be used in frequentist applications, in particular for maximum-likelihood estimation. The approach is illustrated by an example of ancestral inference in population genetics. A number of open problems are highlighted in the discussion.
Marker succession during the development of keratinocytes from cultured human embryonic stem cells
Authors: Green Howard , Easley Karen , Iuchi Shiro
Published: December 2003

Human embryonic stem cells injected into
scid
mice produce nodules containing differentiated somatic tissues. From the trypsinized cells of such a nodule, we have recovered keratinocytes that can be grown in cell culture. The method of recovery is sensitive enough to detect small numbers of keratinocytes formed in the nodule, but for purposes of analysis, it is preferable to study the development of the entire keratinocyte lineage in culture. The principle of our analysis is the successive appearance of markers, including transcription factors with considerable specificity for the keratinocyte (p63 and basonuclin) and differentiation markers characteristic of its final state (keratin 14 and involucrin). We have determined the order of marker succession during the time- and migration-dependent development of keratinocytes from single embryoid bodies in cell culture. Of the markers we have examined, p63 was the earliest to appear in the keratinocyte lineage. The successive accumulation of later markers provides increasing certainty of emergence of the definitive keratinocyte.
A role in vacuolar arginine transport for yeast Btn1p and for human CLN3, the protein defective in Batten disease
Authors: Kim Yoojin , Ramirez-Montealegre Denia , Pearce David A.
Published: December 2003

In
Saccharomyces cerevisiae
, transport of arginine into the vacuole has previously been shown to be facilitated by a putative H
+
/arginine antiport. We confirm that transport of arginine into isolated yeast vacuoles requires ATP and we demonstrate a requirement for a functional vacuolar H
+
-ATPase. We previously reported that deletion of
BTN1
(
btn1
-Δ), an ortholog of the human Batten disease gene
CLN3
, resulted in a decrease in vacuolar pH during early growth. We report that this altered vacuolar pH in
btn1
-Δ strains underlies a lack of arginine transport into the vacuole, which results in a depletion of endogenous vacuolar arginine levels. This arginine transport defect in
btn1
-Δ is complemented by expression of either
BTN1
or the human
CLN3
gene and strongly suggests a function for transport of, or regulation of the transport of, basic amino acids into the vacuole or lysosome for yeast Btn1p, and human CLN3 protein, respectively. We propose that defective transport at the lysosomal membrane caused by an absence of functional CLN3 is the primary biochemical defect that results in Batten disease.
Cap analysis gene expression for high-throughput analysis of transcriptional starting point and identification of promoter usage
Authors: Shiraki Toshiyuki , Kondo Shinji , Katayama Shintaro , Waki Kazunori , Kasukawa Takeya , Kawaji Hideya , Kodzius Rimantas , Watahiki Akira , Nakamura Mari , Arakawa Takahiro , Fukuda Shiro , Sasaki Daisuke , Podhajska Anna , Harbers Matthias , Kawai Jun , Carninci Piero , Hayashizaki Yoshihide
Published: December 2003

We introduce cap analysis gene expression (CAGE), which is based on preparation and sequencing of concatamers of DNA tags deriving from the initial 20 nucleotides from 5′ end mRNAs. CAGE allows high-throughout gene expression analysis and the profiling of transcriptional start points (TSP), including promoter usage analysis. By analyzing four libraries (brain, cortex, hippocampus, and cerebellum), we redefined more accurately the TSPs of 11-27% of the analyzed transcriptional units that were hit. The frequency of CAGE tags correlates well with results from other analyses, such as serial analysis of gene expression, and furthermore maps the TSPs more accurately, including in tissue-specific cases. The high-throughput nature of this technology paves the way for understanding gene networks via correlation of promoter usage and gene transcriptional factor expression.
A tenascin-C aptamer identified by tumor cell SELEX: Systematic evolution of ligands by exponential enrichment
Authors: Daniels Dion A. , Chen Hang , Hicke Brian J. , Swiderek Kristine M. , Gold Larry
Published: December 2003

The targeting of molecular repertoires to complex systems rather than biochemically pure entities is an accessible approach that can identify proteins of biological interest. We have probed antigens presented by a monolayer of tumor cells for their ability to interact with a pool of aptamers. A glioblastoma-derived cell line, U251, was used as the target for systematic evolution of ligands by exponential enrichment by using a single-stranded DNA library. We isolated specifically interacting oligonucleotides, and biochemical strategies were used to identify the protein target for one of the aptamers. Here we characterize the interaction of the DNA aptamer, GBI-10, with tenascin-C, an extracellular protein found in the tumor matrix. Tenascin-C is believed to be involved in both embryogenesis and oncogenesis pathways. Systematic evolution of ligands by exponential enrichment appears to be a successful strategy for the
a priori
identification of targets of biological interest within complex systems.
Targeted deletion of apoptosis signal-regulating kinase 1 attenuates left ventricular remodeling
Authors: Yamaguchi Osamu , Higuchi Yoshiharu , Hirotani Shinichi , Kashiwase Kazunori , Nakayama Hiroyuki , Hikoso Shungo , Takeda Toshihiro , Watanabe Tetsuya , Asahi Michio , Taniike Masayuki , Matsumura Yasushi , Tsujimoto Ikuko , Hongo Kenichi , Kusakari Yoichiro , Kurihara Satoshi , Nishida Kazuhiko , Ichijo Hidenori , Hori Masatsugu , Otsu Kinya
Published: December 2003

Left ventricular remodeling that occurs after myocardial infarction (MI) and pressure overload is generally accepted as a determinant of the clinical course of heart failure. The molecular mechanism of this process, however, remains to be elucidated. Apoptosis signal-regulating kinase 1 (ASK1) is a mitogen-activated protein kinase kinase kinase that plays an important role in stress-induced apoptosis. We used ASK1 knockout mice (ASK
-/-
) to test the hypothesis that ASK1 is involved in development of left ventricular remodeling. ASK
-/-
hearts showed no morphological or histological defects. Echocardiography and cardiac catheterization revealed normal global structure and function. Left ventricular structural and functional remodeling were determined 4 weeks after coronary artery ligation or thoracic transverse aortic constriction (TAC). ASK
-/-
had significantly smaller increases in left ventricular end-diastolic and end-systolic ventricular dimensions and smaller decreases in fractional shortening in both experimental models compared with WT mice. The number of terminal deoxynucleotidyl transferase biotin-dUDP nick end-labeling-positive myocytes after MI or TAC was decreased in ASK
-/-
compared with that in WT mice. Overexpression of a constitutively active mutant of ASK1 induced apoptosis in isolated rat neonatal cardiomyocytes, whereas neonatal ASK
-/-
cardiomyocytes were resistant to H
2
O
2
-induced apoptosis. An
in vitro
kinase assay showed increased ASK1 activity in heart after MI or TAC in WT mice. Thus, ASK1 plays an important role in regulating left ventricular remodeling by promoting apoptosis.
Epstein–Barr virus latent membrane protein 1 activation of NF-κB through IRAK1 and TRAF6
Authors: Luftig Micah , Prinarakis Efthimios , Yasui Teruhito , Tsichritzis Theodore , Cahir-McFarland Ellen , Inoue Jun-Ichiro , Nakano Hiroyasu , Mak Tak Wah , Yeh Wen-Chen , Li Xiaoxia , Akira Shizuo , Suzuki Nobutaka , Suzuki Shinobu , Mosialos George , Kieff Elliott
Published: December 2003

Epstein–Barr virus latent membrane protein 1 (LMP1) activation of NF-κB is critical for Epstein–Barr virus-infected B lymphocyte survival. LMP1 activates the IκB kinase complex and NF-κB through two cytoplasmic signaling domains that engage tumor necrosis factor receptor-associated factor (TRAF)1/2/3/5 or TRADD and RIP. We now use cells lacking expression of TRAF2, TRAF5, TRAF6, IKKα, IKKβ, IKKγ, TAB2, IL-1 receptor-associated kinase (IRAK)1, or IRAK4 to assess their roles in LMP1-mediated NF-κB activation. LMP1-induced RelA nuclear translocation was similar in IKKα knockout (KO) and WT murine embryo fibroblasts (MEFs) but substantially deficient in IKKβ KO MEFs. NF-κB-dependent promoter responses were also substantially deficient in IKKβ KO MEFs but were hyperactive in IKKα KO MEFs. More surprisingly, NF-κB responses were near normal in TRAF2 and TRAF5 double-KO MEFs, IKKγ KO MEFs, TAB2 KO MEFs, and IRAK4 KO MEFs but were highly deficient in TRAF6 KO MEFs and IRAK1 KO HEK293 cells. Consistent with the importance of TRAF6, LMP1-induced NF-κB activation in HEK293 cells was inhibited by expression of dominant-negative TAB2 and Ubc13 alleles. These data extend a role for IKKα in IKKβ regulation, identify an unusual IKKβ-dependent and IKKγ-independent NF-κB activation, and indicate that IRAK1 and TRAF6 are essential for LMP1-induced NF-κB activation.
Arabidopsis CHL27, located in both envelope and thylakoid membranes, is required for the synthesis of protochlorophyllide
Authors: Tottey Stephen , Block Maryse A. , Allen Michael , Westergren Tomas , Albrieux Catherine , Scheller Henrik V. , Merchant Sabeeha , Jensen Poul Erik
Published: December 2003

CHL27, the
Arabidopsis
homologue to
Chlamydomonas
Crd1, a plastid-localized putative diiron protein, is required for the synthesis of protochlorophyllide and therefore is a candidate subunit of the aerobic cyclase in chlorophyll biosynthesis. δ-Aminolevulinic acid-fed antisense
Arabidopsis
plants with reduced amounts of Crd1/CHL27 accumulate Mg-protoporphyrin IX monomethyl ester, the substrate of the cyclase reaction. Mutant plants have chlorotic leaves with reduced abundance of all chlorophyll proteins. Fractionation of
Arabidopsis
chloroplast membranes shows that Crd1/CHL27 is equally distributed on a membrane-weight basis in the thylakoid and inner-envelope membranes.
Crystal structures that suggest late development of genetic code components for differentiating aromatic side chains
Authors: Yang Xiang-Lei , Otero Francella J. , Skene Robert J. , McRee Duncan E. , Schimmel Paul , Ribas de Pouplana Lluís
Published: December 2003

Early forms of the genetic code likely generated “statistical” proteins, with similar side chains occupying the same sequence positions at different ratios. In this scenario, groups of related side chains were treated by aminoacyl-tRNA synthetases as a single molecular species until a discrimination mechanism developed that could separate them. The aromatic amino acids tryptophan, tyrosine, and phenylalanine likely constituted one of these groups. A crystal structure of human tryptophanyl-tRNA synthetase was solved at 2.1 Å with a tryptophanyl-adenylate bound at the active site. A cocrystal structure of an active fragment of human tyrosyl-tRNA synthetase with its cognate amino acid analog was also solved at 1.6 Å. The two structures enabled active site identifications and provided the information for structure-based sequence alignments of ≈45 orthologs of each enzyme. Two critical positions shared by all tyrosyl-tRNA synthetases and tryptophanyl-tRNA synthetases for amino acid discrimination were identified. The variations at these two positions and phylogenetic analyses based on the structural information suggest that, in contrast to many other amino acids, discrimination of tyrosine from tryptophan occurred late in the development of the genetic code.
Trans-editing of mischarged tRNAs
Authors: Ahel Ivan , Korencic Dragana , Ibba Michael , Söll Dieter
Published: December 2003

Aminoacyl-tRNA synthetases (aaRSs) are multidomain proteins that specifically attach amino acids to their cognate tRNAs. Their most conserved, and presumably evolutionarily oldest, domains are the catalytic cores, which activate amino acids and transfer them to the 3′ ends of tRNAs. Additional domains appended to or inserted in the body of aaRSs increase efficiency and specificity of the aminoacylation process, either by providing additional tRNA contacts, or by hydrolyzing noncognate amino acid products (cis
-
editing). Here, we report specific tRNA-dependent trans
-
editing by aaRS-like proteins that reciprocate the editing domains of aaRSs, but not the remainder of the corresponding enzyme. A freestanding homologue of the prolyl-tRNA synthetase-editing domain, the PrdX protein from
Clostridium sticklandii
, efficiently and specifically hydrolyzes Ala-tRNA
Pro
. Similarly, autonomous alanyl-tRNA synthetase-editing domain homologues (AlaX proteins) from
Methanosarcina barkeri
and
Sulfolobus solfataricus
hydrolyze Ser-tRNA
Ala
and Gly-tRNA
Ala
substrates. The discovery of autonomous editing proteins efficient in hydrolyzing misacylated products provides a direct link between ancestral aaRSs consisting solely of the catalytic core and extant enzymes to which functionally independent modules are appended.
Climatic suitability for malaria transmission in Africa, 1911–1995
Authors: Small Jennifer , Goetz Scott J. , Hay Simon I.
Published: December 2003

Time series analysis of a climate-driven model of malaria transmission shows limited evidence for an increase in suitability during the last century across Africa. Outside areas where climate was always or never suitable, <17% of the continent showed significant trends in malaria transmission. Of these areas, 5.7% showed positive deterministic trends, 6.1% had negative deterministic trends, and 5.1% exhibited stochastic trends. In areas with positive trends, precipitation, rather than temperature, was the primary forcing variable. This analysis highlights the need to examine the relationship between climate and malaria more closely and to fully consider nonclimatic factors as drivers of increased malaria transmission across the continent.
Redox-coupled proton translocation in biological systems: Proton shuttling in cytochrome
c
oxidase
Authors: Namslauer Andreas , Pawate Ashtamurthy S. , Gennis Robert B. , Brzezinski Peter
Published: December 2003

In the respiratory chain free energy is conserved by linking the chemical reduction of dioxygen to the electrogenic translocation of protons across a membrane. Cytochrome
c
oxidase (C
c
O) is one of the sites where this linkage occurs. Although intensively studied, the molecular mechanism of proton pumping by this enzyme remains unknown. Here, we present data from an investigation of a mutant C
c
O from
Rhodobacter sphaeroides
[Asn-139 → Asp, ND(I-139)] in which proton pumping is completely uncoupled from the catalytic turnover (i.e., reduction of O
2
). However, in this mutant C
c
O, the rate by which O
2
is reduced to H
2
O is even slightly higher than that of the wild-type C
c
O. The data indicate that the disabling of the proton pump is a result of a perturbation of E(I-286), which is located 20 Å from N(I-139) and is an internal proton donor to the catalytic site, located in the membrane-spanning part of C
c
O. The mutation results in raising the effective pK
a
of E(I-286) by 1.6 pH units. An explanation of how the mutation uncouples catalytic turnover from proton pumping is offered, which suggests a mechanism by which C
c
O pumps protons.
Rapid protein fold determination using unassigned NMR data
Authors: Meiler Jens , Baker David
Published: December 2003

Experimental structure determination by x-ray crystallography and NMR spectroscopy is slow and time-consuming compared with the rate at which new protein sequences are being identified. NMR spectroscopy has the advantage of rapidly providing the structurally relevant information in the form of unassigned chemical shifts (CSs), intensities of NOESY crosspeaks [nuclear Overhauser effects (NOEs)], and residual dipolar couplings (RDCs), but use of these data are limited by the time and effort needed to assign individual resonances to specific atoms. Here, we develop a method for generating low-resolution protein structures by using unassigned NMR data that relies on thede novoprotein structure prediction algorithm,rosetta[Simons, K. T., Kooperberg, C., Huang, E. & Baker, D. (1997)J. Mol. Biol.268, 209–225] and a Monte Carlo procedure that searches for the assignment of resonances to atoms that produces the best fit of the experimental NMR data to a candidate 3D structure. A large ensemble of models is generated from sequence information alone by usingrosetta, an optimal assignment is identified for each model, and the models are then ranked based on their fit with the NMR data assuming the identified assignments. The method was tested on nine protein sequences between 56 and 140 amino acids and published CS, NOE, and RDC data. The procedure yielded models with rms deviations between 3 and 6 Å, and, in four of the nine cases, the partial assignments obtained by the method could be used to refine the structures to high resolution (0.6–1.8 Å) by repeated cycles of structure generation guided by the partial assignments, followed by reassignment using the newly generated models.
Differential survival of solitary and aggregated bacterial cells promotes aggregate formation on leaf surfaces
Authors: Monier J.-M. , Lindow S. E.
Published: December 2003

The survival of individual
Pseudomonas syringae
cells was determined on bean leaf surfaces maintained under humid conditions or periodically exposed to desiccation stress. Cells of
P. syringae
strain B728a harboring a GFP marker gene were visualized by epifluorescence microscopy, either directly
in situ
or after recovery from leaves, and dead cells were identified as those that were stained with propidium iodide in such populations. Under moist, conducive conditions on plants, the proportion of total live cells was always high, irrespective of their aggregated state. In contrast, the proportion of the total cells that remained alive on leaves that were periodically exposed to desiccation stress decreased through time and was only ≈15% after 5 days. However, the fraction of cells in large aggregates that were alive on such plants in both condition was much higher than more solitary cells. Immediately after inoculation, cells were randomly distributed over the leaf surface and no aggregates were observed. However, a very aggregated pattern of colonization was apparent within 7 days, and >90% of the living cells were located in aggregates of 100 cells or more. Our results strongly suggest that, although conducive conditions favor aggregate formation, such cells are much more capable of tolerating environmental stresses, and the preferential survival of cells in aggregates promotes a highly clustered spatial distribution of bacteria on leaf surfaces.
Vacuolar sorting receptor for seed storage proteins in
Arabidopsis thaliana
Authors: Shimada Tomoo , Fuji Kentaro , Tamura Kentaro , Kondo Maki , Nishimura Mikio , Hara-Nishimura Ikuko
Published: December 2003

The seeds of higher plants accumulate large quantities of storage protein. During seed maturation, storage protein precursors synthesized on rough endoplasmic reticulum are sorted to protein storage vacuoles, where they are converted into the mature forms and accumulated. Previous attempts to determine the sorting machinery for storage proteins have not been successful. Here we show that a type I membrane protein, AtVSR1/AtELP, of
Arabidopsis
functions as a sorting receptor for storage proteins. The
atvsr1
mutant missorts storage proteins by secreting them from cells, resulting in an enlarged and electron-dense extracellular space in the seeds. The
atvsr1
seeds have distorted cells and smaller protein storage vacuoles than do WT seeds, and
atvsr1
seeds abnormally accumulate the precursors of two major storage proteins, 12S globulin and 2S albumin, together with the mature forms of these proteins. AtVSR1 was found to bind to the C-terminal peptide of 12S globulin in a Ca
2+
-dependent manner. These findings demonstrate a receptor-mediated transport of seed storage proteins to protein storage vacuoles in higher plants.
Electroosmotic enhancement of the binding of a neutral molecule to a transmembrane pore
Authors: Gu Li-Qun , Cheley Stephen , Bayley Hagan
Published: December 2003

The flux of solvent water coupled to the transit of ions through protein pores is considerable. The effect of this electroosmotic solvent flow on the binding of a neutral molecule [β-cyclodextrin (βCD)] to sites within the staphylococcal α-hemolysin pore was investigated. Mutant α-hemolysin pores were used to which βCD can bind from either entrance and through which the direction of water flow can be controlled by choosing the charge selectivity of the pore and the polarity of the applied potential. The
K
d
values for βCD for individual mutant pores varied by >100-fold with the applied potential over a range of –120 to +120 mV. In all cases, the signs of the changes in binding free energy and the influence of potential on the association and dissociation rate constants for βCD were consistent with an electroosmotic effect.
The compositional adjustment of amino acid substitution matrices
Authors: Yu Yi-Kuo , Wootton John C. , Altschul Stephen F.
Published: December 2003

Amino acid substitution matrices are central to protein-comparison methods. In most commonly used matrices, the substitution scores take a log-odds form, involving the ratio of “target” to “background” frequencies derived from large, carefully curated sets of protein alignments. However, such matrices often are used to compare protein sequences with amino acid compositions that differ markedly from the background frequencies used for the construction of the matrices. Of course, the target frequencies should be adjusted in such cases, but the lack of an appropriate way to do this has been a long-standing problem. This article shows that if one demands consistency between target and background frequencies, then a log-odds substitution matrix implies a unique set of target and background frequencies as well as a unique scale. Standard substitution matrices therefore are truly appropriate only for the comparison of proteins with standard amino acid composition. Accordingly, we present and evaluate a rationale for transforming the target frequencies implicit in a standard matrix to frequencies appropriate for a nonstandard context. This rationale yields asymmetric matrices for the comparison of proteins with divergent compositions. Earlier approaches are unable to deal with this case in a fully consistent manner. Composition-specific substitution matrix adjustment is shown to be of utility for comparing compositionally biased proteins, including those of organisms with nucleotide-biased, and therefore codon-biased, genomes or isochores.
Changing identities: tRNA duplication and remolding within animal mitochondrial genomes
Authors: Rawlings Timothy A. , Collins Timothy M. , Bieler Rüdiger
Published: December 2003

Although the majority of metazoan mitochondrial genomes (mtDNAs) contain the same 37 genes, including 22 encoding transfer RNAs (tRNAs), the recognition of orthologs is not always straightforward. Here we demonstrate that inferring tRNA orthologs among taxa by using anticodon triplets and deduced secondary structure can be misleading: through a process of tRNA duplication and mutation in the anticodon triplet, remolded leucine (L
UUR
) tRNA genes have repeatedly taken over the role of isoaccepting L
CUN
leucine tRNAs within metazoan mtDNA. In the present work, data from within the gastropods and a broad survey of metazoan mtDNA suggest that tRNA leucine duplication and remolding events have occurred independently at least seven times within three major animal lineages. In all cases where the mechanism of gene remolding can be inferred with confidence, the direction is the same: from L
UUR
to L
CUN
. Gene remolding and its apparent asymmetry have significant implications for the use of mitochondrial tRNA gene orders as phylogenetic markers. Remolding complicates the identification of orthologs and can result in convergence in gene order. Careful sequence-based analysis of tRNAs can help to recognize this homoplasy, improving gene-order-based phylogenetic hypotheses and underscoring the importance of careful homology assessment. tRNA remolding also provides an additional mechanism by which gene order changes can occur within mtDNA: through the changing identity of tRNA genes themselves. Recognition of these remolding events can lead to new interpretations of gene order changes, as well as the discovery of phylogenetically relevant gene dynamics that are hidden at the level of gene order alone.
Structural and functional characterization of the VirB5 protein from the type IV secretion system encoded by the conjugative plasmid pKM101
Authors: Yeo Hye-Jeong , Yuan Qing , Beck Moriah R. , Baron Christian , Waksman Gabriel
Published: December 2003

Type IV secretion systems mediate intercellular transfer of macro-molecules via a mechanism ancestrally related to that of bacterial conjugation machineries. TraC of the IncN plasmid pKM101 belongs to the VirB5 family of proteins, an essential component of most type IV secretion systems. Here, we present the structure of TraC. VirB5/TraC is a single domain protein, which consists of a three helix bundle and a loose globular appendage. Structure-based site-directed mutagenesis followed by functional studies indicates that VirB5 proteins participate in protein-protein interactions important for pilus assembly and function.
Transcriptome analysis of chlamydial growth during IFN-γ-mediated persistence and reactivation
Authors: Belland Robert J. , Nelson David E. , Virok Dezso , Crane Deborah D. , Hogan Daniel , Sturdevant Daniel , Beatty Wandy L. , Caldwell Harlan D.
Published: December 2003

Chlamydia trachomatisis an obligatory intracellular prokaryotic parasite that causes a spectrum of clinically important chronic inflammatory diseases of humans. Persistent infection may play a role in the pathophysiology of chlamydial disease. Here we describe the chlamydial transcriptome in anin vitromodel of IFN-γ-mediated persistence and reactivation from persistence. Tryptophan utilization, DNA repair and recombination, phospholipid utilization, protein translation, and general stress genes were up-regulated during persistence. Down-regulated genes included chlamydial late genes and genes involved in proteolysis, peptide transport, and cell division. Persistence was characterized by altered but active biosynthetic processes and continued replication of the chromosome. On removal of IFN-γ, chlamydiae rapidly reentered the normal developmental cycle and reversed transcriptional changes associated with cytokine treatment. The coordinated transcriptional response to IFN-γ implies that a chlamydial response stimulon has evolved to control the transition between acute and persistent growth of the pathogen. In contrast to the paradigm of persistence as a general stress response, our findings suggest that persistence is an alternative life cycle used by chlamydiae to avoid the host immune response.
Fear of novelty in infant rats predicts adult corticosterone dynamics and an early death
Authors: Cavigelli S. A. , McClintock M. K.
Published: December 2003

Individuals who are fearful of novelty have a larger hypothalamic-pituitary-adrenal axis response than do nonfearful individuals. We hypothesized that a fearful behavioral style emerging early in life would be associated with life-long altered adrenal activity. Because there is ample physiological evidence both costs and benefits of adrenal activation, we determined whether such a stable emotional-neuroendocrine trait was associated with differential morbidity and mortality. To conduct such lifespan work, we studied a relatively short-lived mammal: the Norway rat. We first established that an animal's hesitation or willingness to explore a novel environment (“neophobia” and “neophilia,” respectively) is an identifiable and stable behavioral trait in young-adult males and that neophobia, compared with neophilia, was associated with a greater glucocorticoid response to novelty. Second, we were able to detect behavioral differences among infant rats within a family, and this behavioral disposition at infancy predicted the magnitude of the glucocorticoid response in late middle age. Males identified as neophobic during infancy died sooner than their less fearful brothers. Although both types of males died with similar pathologies (tumors), neophobic males were 60% more likely to die at any point in time. This lifespan study identifies an emotional trait in infancy that predicts an early death and an associated neuroendocrine trait in adulthood that is a potential mechanism underlying the relationship between behavioral style and longevity.
IκBα/IκBε deficiency reveals that a critical NF-κB dosage is required for lymphocyte survival
Authors: Goudeau Bertrand , Huetz François , Samson Sandrine , Di Santo James P. , Cumano Ana , Beg Amer , Israël Alain , Mémet Sylvie
Published: December 2003

In most cells, the NF-κB transcription factor is sequestered in the cytoplasm by interaction with inhibitory proteins, the IκBs. Here, we show that combined IκBα/IκBε deficiency in mice leads to neonatal death, elevated κB binding activity, overexpression of NF-κB target genes, and disruption of lymphocyte production. In IκBα/IκBε-deficient fetuses, B220+IgM+B cells and single-positive T cells die by apoptosis. In adults, IκBα-/-IκBε-/-reconstituted chimeras exhibit a nearly complete absence of T and B cells that is not rescued by cotransfer with wild-type bone marrow. These findings demonstrate that IκBs tightly control NF-κB activityin vivoand that increased NF-κB activity intrinsically impairs lymphocyte survival. Because reduction or rise of NF-κB activity leads to similar dysfunction, they also reveal that only a narrow window of NF-κB activity is tolerated by lymphocytes.
Structure of Mth11/
Mth
Rpp29, an essential protein subunit of archaeal and eukaryotic RNase P
Authors: Boomershine William P. , McElroy Craig A. , Tsai Hsin-Yue , Wilson Ross C. , Gopalan Venkat , Foster Mark P.
Published: December 2003

We have determined the solution structure of Mth11 (
Mth
Rpp29), an essential subunit of the RNase P enzyme from the archaebacterium
Methanothermobacter thermoautotrophicus (Mth
). RNase P is a ubiquitous ribonucleoprotein enzyme primarily responsible for cleaving the 5′ leader sequence during maturation of tRNAs in all three domains of life. In eubacteria, this enzyme is made up of two subunits: a large RNA (≈120 kDa) responsible for mediating catalysis, and a small protein cofactor (≈15 kDa) that modulates substrate recognition and is required for efficient
in vivo
catalysis. In contrast, multiple proteins are associated with eukaryotic and archaeal RNase P, and these proteins exhibit no recognizable homology to the conserved bacterial protein subunit. In reconstitution experiments with recombinantly expressed and purified protein subunits, we found that
Mth
Rpp29, a homolog of the Rpp29 protein subunit from eukaryotic RNase P, is an essential protein component of the archaeal holoenzyme. Consistent with its role in mediating protein–RNA interactions, we report that
Mth
Rpp29 is a member of the oligonucleotide/oligosaccharide binding fold family. In addition to a structured β-barrel core, it possesses unstructured N- and C-terminal extensions bearing several highly conserved amino acid residues. To identify possible RNA contacts in the protein–RNA complex, we examined the interaction of the 11-kDa protein with the full 100-kDa
Mth
RNA subunit by using NMR chemical shift perturbation. Our findings represent a critical step toward a structural model of the RNase P holoenzyme from archaebacteria and higher organisms.