A partial map of the barley genome incorporating restriction fragment length polymorphism, polymerase chain reaction, isozyme, and morphological marker loci

Nine low copy number genomic DNA clones, a ribosomal sequence, and seven cDNA clones were found to identify polymorphisms in cultivated barley (Hordeum vulgare L.). An F2 population consisting of 100 plants was produced from a cross between a high-yielding two-rowed feed barley cultivar and a genetic marker stock homozygous for nine recessive and one dominant morphological marker genes. Through the use of these 10 well-distributed marker genes, five previously mapped isozyme loci, and two storage-protein loci, the approximate recombinational location for each of 17 restriction fragment length polymorphism loci was estimated. One clone, pMSU21, identified variation that appeared to be the result of a small insertion-deletion event that differentiated two-rowed and six-rowed genotypes. This difference was characterized, and one allele was sequenced. Oligonucleotide primers that flanked the insertion-deletion event were synthesized, and DNA samples from the F2 population were subjected to polymerase chain reaction sequence amplification. The variation identified by this technique was determined to be allelic to the variation identified using pMSU21 in Southern blot analysis. Maps of previously undescribed informative clones are included.     

IL-17E, a novel proinflammatory ligand for the IL-17 receptor homolog IL-17Rh1

We report identification of interleukin (IL)-17E, a novel member of the IL-17 family of cytokines. IL-17E is a ligand for the recently identified protein termed EVI27/IL-17BR, which we term IL-17 receptor homolog 1 (IL-17Rh1) in light of the multiple reported ligand-receptor relationships. Murine EVI27 was identified through its location at a common site of retroviral integration in BXH2 murine myeloid leukemias. IL-17Rh1 shows highest level expression in kidney with moderate expression in multiple other organs, whereas IL-17E mRNA was detected at very low levels in several peripheral tissues. IL-17E induces activation of NF-kappaB and stimulates production of the proinflammatory chemokine IL-8.     

Isolation and characterization of a novel conus peptide with apparent antinociceptive activity

Cone snails are tropical marine mollusks that envenomate prey with a complex mixture of neuropharmacologically active compounds. We report the discovery and biochemical characterization of a structurally unique peptide isolated from the venom of Conus marmoreus. The new peptide, mr10a, potently increased withdrawal latency in a hot plate assay (a test of analgesia) at intrathecal doses that do not produce motor impairment as measured by rotarod test. The sequence of mr10a is NGVCCGYKLCHOC, where O is 4-trans-hydroxyproline. This sequence is highly divergent from all other known conotoxins. Analysis of a cDNA clone encoding the toxin, however, indicates that it is a member of the recently described T-superfamily. Total chemical synthesis of the three possible disulfide arrangements of mr10a was achieved, and elution studies indicate that the native form has a disulfide connectivity of Cys1-Cys4 and Cys2-Cys3. This disulfide linkage is unprecedented among conotoxins and defines a new family of Conus peptides.     

The spasmodic peptide defines a new conotoxin superfamily

We purified and characterized a peptide from the venom of Conus textile that makes normal mice assume the phenotype of a well-known mutant, the spasmodic mouse. This "spasmodic" peptide has 27 amino acids, including two gamma-carboxyglutamate (Gla) residues. A cDNA clone encoding the precursor for the peptide was identified; a gamma-carboxylation recognition signal sequence (gamma-CRS) is present in the -1 --> -20 region of the peptide precursor. Both the gamma-CRS and the position of the Gla residues in the mature toxin are notably different from other Gla-containing conopeptides. The spasmodic peptide has a novel disulfide framework and distinct signal sequence which together define a new P-superfamily of conopeptides. A cDNA encoding another member of the P-superfamily was identified from a different species, Conus gloriamaris.     

Diet composition,feed preferences and mouth morphology of early stage silver therapon (Leiopotherapon plumbeus,Kner 1864) larvae reared in outdoor tanks

This study examined the diet composition, feeding preferences, and mouth morphology of the silver therapon (Leiopotherapon plumbeus, Kner 1864) larvae under captive conditions. Larvae were reared in outdoor tanks (4 m³) with natural food grown 2 weeks prior to start of larval rearing. Food preference was measured by the Chesson's electivity index (α_i). Gut content analysis of larvae sampled between 5 and 25 days after hatching (DAH) showed the dominance in the diet by zooplankton, mainly copepod nauplii, cladocerans and insect larvae. Small fish larvae (5–9 DAH; 3.32–6.29 mm standard length) preferred cladocerans, ciliates and copepod nauplii; whereas older larvae (12–25 DAH; 5.45–19.26 mm standard length) preferred insect larvae over cladocerans and adult insects. The mouth gape size at 5 DAH was 359 μm and increased to 3.75 mm at 40 DAH when body size grew at an average rate of 0.59 mm d^?1. The standard length (SL) of L. plumbeus larvae was strongly associated with mouth size (r² = 0.98, P < 0.05), indicating a progressive increase of ingested prey size of the fish larvae. These results clarified the early life feeding ecology of this species, which is essential in developing effective hatchery techniques.     

Definition of the M-conotoxin superfamily: characterization of novel peptides from molluscivorous Conus venoms

Most of the >50,000 different pharmacologically active peptides in Conus venoms belong to a small number of gene superfamilies. In this work, the M-conotoxin superfamily is defined using both biochemical and molecular criteria. Novel excitatory peptides purified from the venoms of the molluscivorous species Conus textile and Conus marmoreus all have a characteristic pattern of Cys residues previously found in the mu-, kappaM-, and psi-conotoxins (CC-C-C-CC). The new peptides are smaller (12-19 amino acids) than the mu-, kappaM-, and psi-conotoxins (22-24 amino acids). One peptide, mr3a, was chemically synthesized in a biologically active form. Analysis of the disulfide bridges of a natural peptide tx3c from C. textile and synthetic peptide mr3a from C. marmoreus showed a novel pattern of disulfide connectivity, different from that previously established for the mu- and psi-conotoxins. Thus, these peptides belong to a new group of structurally and pharmacologically distinct conotoxins that are particularly prominent in the venoms of mollusc-hunting Conus species. Analysis of cDNA clones encoding the novel peptides as well as those encoding mu-, kappaM-, and psi-conotoxins revealed highly conserved amino acid residues in the precursor sequences; this conservation in both amino acid sequence and in the Cys pattern defines a gene superfamily, designated the M-conotoxin superfamily. The peptides characterized can be provisionally assigned to four distinct groups within the M-superfamily based on sequence similarity within and divergence between each group. A notable feature of the superfamily is that two distinct structural frameworks have been generated by changing the disulfide connectivity on an otherwise conserved Cys pattern.     

Conformational lability in serine protease active sites: structures of hepatocyte growth factor activator (HGFA) alone and with the inhibitory domain from HGFA inhibitor-1B

Hepatocyte growth factor activator (HGFA) is a serine protease that converts hepatocyte growth factor (HGF) into its active form. When activated HGF binds its cognate receptor Met, cellular signals lead to cell growth, differentiation, and migration, activities which promote tissue regeneration in liver, kidney and skin. Intervention in the conversion of HGF to its active form has the potential to provide therapeutic benefit where HGF/Met activity is associated with tumorigenesis. To help identify ways to moderate HGF/Met effects, we have determined the molecular structure of the protease domain of HGFA. The structure we determined, at 2.7 A resolution, with no pseudo-substrate or inhibitor bound is characterized by an unconventional conformation of key residues in the enzyme active site. In order to find whether this apparently non-enzymatically competent arrangement would persist in the presence of a strongly-interacting inhibitor, we also have determined, at 2.6 A resolution, the X-ray structure of HGFA complexed with the first Kunitz domain (KD1) from the physiological inhibitor hepatocyte growth factor activator inhibitor 1B (HAI-1B). In this complex we observe a rearranged substrate binding cleft that closely mirrors the cleft of other serine proteases, suggesting an extreme conformational dynamism. We also characterize the inhibition of 16 serine proteases by KD1, finding that the previously reported enzyme specificity of the intact extracellular region of HAI-1B resides in KD1 alone. We find that HGFA, matriptase, hepsin, plasma kallikrein and trypsin are potently inhibited, and use the complex structure to rationalize the structural basis of these results.     

Competition between Decapping Complex Formation and Ubiquitin-Mediated Proteasomal Degradation Controls Human Dcp2 Decapping Activity

mRNA decapping is a central step in eukaryotic mRNA decay that simultaneously shuts down translation initiation and activates mRNA degradation. A major complex responsible for decapping consists of the decapping enzyme Dcp2 in association with decapping enhancers. An important question is how the activity and accumulation of Dcp2 are regulated at the cellular level to ensure the specificity and fidelity of the Dcp2 decapping complex. Here, we show that human Dcp2 levels and activity are controlled by a competition between decapping complex assembly and Dcp2 degradation. This is mediated by a regulatory domain in the Dcp2 C terminus, which, on the one hand, promotes Dcp2 activation via decapping complex formation mediated by the decapping enhancer Hedls and, on the other hand, targets Dcp2 for ubiquitin-mediated proteasomal degradation in the absence of Hedls association. This competition between Dcp2 activation and degradation restricts the accumulation and activity of uncomplexed Dcp2, which may be important for preventing uncontrolled decapping or for regulating Dcp2 levels and activity according to cellular needs.     

Anti-angiogenic activity of Gracilaria coronopifolia J.G. Agardh extract by lowering the levels of trace metals (iron,zinc and copper) in duck chorioallantoic membrane and in vitro activation of AMP-kinase

Molecular Biology Reports - AMP-activated protein kinase (AMPK) is an intracellular energy sensor important in metabolic regulation, cell growth, and survival. However, the specific role of AMPK...     

Cutting edge: novel human dendritic cell- and monocyte-attracting chemokine-like protein identified by fold recognition methods

Chemokines play an important role in the immune system by regulating cell trafficking in homeostasis and inflammation. In this study, we report the identification and characterization of a novel cytokine-like protein, DMC (dendritic cell and monocyte chemokine-like protein), which attracts dendritic cells and monocytes. The key to the identification of this putative new chemokine was the application of threading techniques to its uncharacterized sequence. Based on our studies, DMC is predicted to have an IL-8-like chemokine fold and to be structurally and functionally related to CXCL8 and CXCL14. Consistent with our predictions, DMC induces migration of monocytes and immature dendritic cells. Expression studies show that DMC is constitutively expressed in lung, suggesting a potential role for DMC in recruiting monocytes and dendritic cells from blood into lung parenchyma.