Inhibition of BMPs by follistatin is required for FGF3 expression and segmental patterning of the hindbrain

A network of molecular interactions is required in the developing vertebrate hindbrain for the formation and anterior-posterior patterning of the rhombomeres. FGF signaling is required in this network to upregulate the expression of the Krox20 and Kreisler segmentation genes, but little is known of how FGF gene expression is regulated in the hindbrain. We show that the dynamic expression of FGF3 in chick hindbrain segments and boundaries is similar to that of the BMP antagonist, follistatin. Consistent with a regulatory relationship between BMP signaling and FGF3 expression, we find that an increase in BMP activity due to blocking of follistatin translation by morpholino antisense oligonucleotides or overexpression of BMP results in strong inhibition of FGF3 expression. Conversely, addition of follistatin leads to an increase in the level of FGF3 expression. Furthermore, the segmental inhibition of BMP activity by follistatin is required for the expression of Krox20, Hoxb1 and EphA4 in the hindbrain. In addition, we show that the maintenance of FGF3 gene expression requires FGF activity, suggestive of an autoregulatory loop. These results reveal an antagonistic relationship between BMP activity and FGF3 expression that is required for correct segmental gene expression in the chick hindbrain, in which follistatin enables FGF3 expression by inhibiting BMP activity.     

Metal ions‐binding T4 lysozyme as an intramolecular protein purification tag compatible with X‐ray crystallography

Phage T4 lysozyme is a well folded and highly soluble protein that is widely used as an insertion tag to improve solubility and crystallization properties of poorly behaved recombinant proteins. It has been used in the fusion protein strategy to facilitate crystallization of various proteins including multiple G protein‐coupled receptors, lipid kinases, or sterol binding proteins. Here, we present a structural and biochemical characterization of its novel, metal ions‐binding mutant (mbT4L). We demonstrate that mbT4L can be used as a purification tag in the immobilized‐metal affinity chromatography and that, in many respects, it is superior to the conventional hexahistidine tag. In addition, structural characterization of mbT4L suggests that mbT4L can be used as a purification tag compatible with X‐ray crystallography.     

Around the world in 10 million years: biogeography of the nearly cosmopolitan true toads (Anura: Bufonidae)

Aim The species‐rich family of true toads (Anura: Bufonidae) has been the focus of several earlier studies investigating the biogeography of geographically widespread taxa. Herein, we employ newly developed Bayesian divergence estimate methods to investigate the biogeographical history of this group. Resulting age estimates are used to test several key temporal hypotheses including that the origin of the bufonid clade pre‐dates Gondwanan vicariance (~105 million years ago, Ma). Area cladograms are also invoked to investigate the geographical origin of the family. Location Worldwide, except the Australia–New Guinea plate, Madagascar and the Antarctic. Methods A phylogenetic hypothesis of the relationships among true toads was derived from analysis of 2521 bp of DNA data including fragments from three mitochondrial (12S, tRNA^val, 16S) and two nuclear (RAG‐1, CXCR‐4) genes. Analysis of multiple, unlinked loci with a Bayesian method for estimating divergence times allowed us to address the timing and biogeographical history of Bufonidae. Resulting divergence estimates permitted the investigation of alternative vicariance/dispersal scenarios that have been proposed for true toads. Results Our area cladogram resulting from phylogenetic analysis of DNA data supports a South American origin for Bufonidae. Divergence estimates indicate that the family originated earlier than had been suggested previously (78–99 Ma). The age of the enigmatic Caribbean clade was dated to the late Palaeocene–early Eocene. A return of bufonids to the New World in the Eocene was followed by rapid diversification and secondary expansion into South America by the early Oligocene (Rupelian). Main conclusions The South American origin of Bufonidae in the Upper Cretaceous was followed by relatively rapid expansion and radiation around the globe, ending with a return to the Americas via a Eurasian/North American land bridge in the Eocene. Though the exact route of this dispersal (Beringia or North Atlantic) remains unclear, an argument is made for the less frequently invoked North Atlantic connection. The origin of the enigmatic Caribbean lineage was found to be consistent with colonization following the bolide impact at the K/T boundary. These findings provide the first, firm foundation for understanding true toad divergence times and their truly remarkable and global radiation.     

A new global biome reconstruction and data-model comparison for the Middle Pliocene

Aim To produce a robust, comprehensive global biome reconstruction for the Middle Pliocene (c. 3.6–2.6 Ma), which is based on an internally consistent palaeobotanical data set and a state‐of‐the‐art coupled climate–vegetation model. The reconstruction gives a more rigorous picture of climate and environmental change during the Middle Pliocene and provides a new boundary condition for future general circulation model (GCM) studies. Location Global. Methods Compilation of Middle Pliocene vegetation data from 202 marine and terrestrial sites into the comprehensive GIS data base TEVIS (Tertiary Environmental Information System). Translation into an internally consistent classification scheme using 28 biomes. Comparison and synthesis of vegetation reconstruction from palaeodata with the outputs of the mechanistically based BIOME4 model forced by climatology derived from the HadAM3 GCM. Results The model results compare favourably with available palaeodata and highlight the importance of employing vegetation–climate feedbacks and the anomaly method in biome models. Both the vegetation reconstruction from palaeobotanical data and the BIOME4 prediction indicate a general warmer and moister climate for the Middle Pliocene. Evergreen taiga as well as temperate forest and grassland shifted northward, resulting in much reduced tundra vegetation. Warm‐temperate forests (with subtropical taxa) spread in mid and eastern Europe and tropical savannas and woodland expanded in Africa and Australia at the expense of deserts. Discrepancies which occurred between data reconstruction and model simulation can be related to: (1) poor spatial model resolution and data coverage; (2) uncertainties in delimiting biomes using climate parameters; or (3) uncertainties in model physics and/or geological boundary conditions. Main conclusions The new global biome reconstruction combines vegetation reconstruction from palaeobotanical proxies with model simulations. It is an important contribution to the further understanding of climate and vegetation changes during the Middle Pliocene warm interval and will enhance our knowledge about how vegetation may change in the future.     

Activation of cap-independent translation by variant eukaryotic initiation factor 4G in vivo

Protein synthesis is tightly controlled by assembly of an intricate ribonucleoprotein complex at the m⁷GTP-cap on eukaryotic mRNAs. Ensuing linear scanning of the 5′ untranslated region (UTR) is believed to transfer the preinitiation complex to the initiation codon. Eukaryotic mRNAs are characterized by significant 5′ UTR heterogeneity, raising the possibility of differential control of translation initiation rate at individual mRNAs. Curiously, many mRNAs with unconventional, highly structured 5′ UTRs encode proteins with central biological roles in growth control, metabolism, or stress response. The 5′ UTRs of such mRNAs may influence protein synthesis rate in multiple ways, but most significantly they have been implicated in mediating alternative means of translation initiation. Cap-independent initiation bypasses strict control over the formation of initiation intermediates at the m⁷GTP cap. However, the molecular mechanisms that favor alternative means of ribosome recruitment are not understood. Here we provide evidence that eukaryotic initiation factor (eIF) 4G controls cap-independent translation initiation at the c-myc and vascular endothelial growth factor (VEGF) 5′ UTRs in vivo. Cap-independent translation was investigated in tetracycline-inducible cell lines expressing either full-length eIF4G or a C-terminal fragment (Ct) lacking interaction with eIF4E and poly(A) binding protein. Expression of Ct, but not intact eIF4G, potently stimulated cap-independent initiation at the c-myc/VEGF 5′ UTRs. In vitro RNA-binding assays suggest that stimulation of cap-independent translation initiation by Ct is due to direct association with the c-myc/VEGF 5′ UTR, enabling 43S preinitiation complex recruitment. Our work demonstrates that variant translation initiation factors enable unconventional translation initiation at mRNA subsets with distinct structural features.     

Genetics of leucine aminopeptidase in apple

Summary Six zones of LAP activity were detected in apples, some of them tissue specific. Genetic studies in four of them revealed the presence of four genes LAP-1, LAP-2, LAP-3 and LAP-4 with 4, 5, 4 and 4 alleles respectively including two null alleles. There were no big differences in allelic frequency within cultivars, selections, rootstocks and Malus species. Close linkage was found between LAP-2 and resistance to mildew derived from White Angel.     

宁南退耕还草区近地面湍流通量观测初报

根据 2 0 0 2～ 2 0 0 3年小气候考察资料 ,分别分析了四季晴天条件下宁南退耕还草区净辐射、感热、潜热和土壤热通量比例和变化特征。结果表明 ,近地层各通量有较规则的日变化。白天 ,感热通量占净辐射的 5 0 %以上 ,一般达 80 %～ 10 0 %。只有夏季全天和秋季白天的潜热通量为正 ,在春季的早晨和秋季的傍晚潜热通量为负。冬春季节土壤的热量收支大于夏秋季节。     

Acetylation of TUG Protein Promotes the Accumulation of GLUT4 Glucose Transporters in an Insulin-responsive Intracellular Compartment

Insulin causes the exocytic translocation of GLUT4 glucose transporters to stimulate glucose uptake in fat and muscle. Previous results support a model in which TUG traps GLUT4 in intracellular, insulin-responsive vesicles termed GLUT4 storage vesicles (GSVs). Insulin triggers TUG cleavage to release the GSVs; GLUT4 then recycles through endosomes during ongoing insulin exposure. The TUG C terminus binds a GSV anchoring site comprising Golgin-160 and possibly other proteins. Here, we report that the TUG C terminus is acetylated. The TUG C-terminal peptide bound the Golgin-160-associated protein, ACBD3 (acyl-CoA-binding domain-containing 3), and acetylation reduced binding of TUG to ACBD3 but not to Golgin-160. Mutation of the acetylated residues impaired insulin-responsive GLUT4 trafficking in 3T3-L1 adipocytes. ACBD3 overexpression enhanced the translocation of GSV cargos, GLUT4 and insulin-regulated aminopeptidase (IRAP), and ACBD3 was required for intracellular retention of these cargos in unstimulated cells. Sirtuin 2 (SIRT2), a NAD⁺-dependent deacetylase, bound TUG and deacetylated the TUG peptide. SIRT2 overexpression reduced TUG acetylation and redistributed GLUT4 and IRAP to the plasma membrane in 3T3-L1 adipocytes. Mutation of the acetylated residues in TUG abrogated these effects. In mice, SIRT2 deletion increased TUG acetylation and proteolytic processing. During glucose tolerance tests, glucose disposal was enhanced in SIRT2 knock-out mice, compared with wild type controls, without any effect on insulin concentrations. Together, these data support a model in which TUG acetylation modulates its interaction with Golgi matrix proteins and is regulated by SIRT2. Moreover, acetylation of TUG enhances its function to trap GSVs within unstimulated cells and enhances insulin-stimulated glucose uptake.