Pin1 inhibitors: Pitfalls,progress and cellular pharmacology

Compelling data supports the hypothesis that Pin1 inhibitors will be useful for the therapy of cancer: Pin1 deficient mice resist the induction of breast cancers normally evoked by expression of MMTV-driven Ras or Erb2 alleles. While Pin1 poses challenges for drug discovery, several groups have identified potent antagonists by structure based drug design, significant progress has been made designing peptidic inhibitors and a number of natural products have been found that blockade Pin1, notably epigallocatchechin gallate (EGCG), a major flavonoid in green tea. Here we critically discuss the modes of action and likely specificity of these compounds, concluding that a suitable chemical biology tool for probing the function of Pin1 has yet to be found. We conclude by outlining some open questions regarding the target validation of Pin1 and the prospects for identification of improved inhibitors in the future.     

The phylogeny of charadriiform birds (shorebirds and allies) – reassessing the conflict between morphology and molecules

Multiple molecular analyses provide a congruent and well‐supported phylogeny of the charadriiform family‐level taxa, which conflicts with previous hypotheses based on osteological data. In order to revise the latter and to identify new characters of phylogenetic significance, skeletons of most charadriiform family‐level taxa were examined and 49 characters analysed. Tree topology was sensitive to outgroup choice, but the result of the analysis rooted with Columbidae (doves and pigeons) recovered a monophyletic Scolopaci, Charadrii, and nonturnicid Lari. With regard to the inclusion of Alcidae and Glareolidae in the Lari, the results of the present study are also in better concordance with the new molecular phylogenies than previous analyses of morphological data. Furthermore, for the first time an apomorphy of a clade including Thinocoridae, Pedionomidae, Rostratulidae, and Jacanidae was identified. Inclusion of Turnicidae in the Lari could not be supported, but there is no strong morphological evidence for an alternative placement. Pluvianus shares derived osteological features with the Burhinidae, and its position in the molecular analyses likewise cannot be corroborated with morphological data. Based on the topology of the molecular consensus tree, the ancestral state of selected characters is reconstructed. It is finally noted that recent calibrations of molecular analyses, which indicate an origin of extant charadriiform lineages in the Cretaceous, are based on incorrectly identified fossils. © 2011 The Linnean Society of London, Zoological Journal of the Linnean Society, 2011, 161 , 916–934.     

Perspectives of taste reception

Summary Ion: solute cotransporters frequency are incapable of achieving equilibrium between the solute accumulation and the transmembrane difference of the electrochemical potential of the ion. The presence of uncoupled flows of ion and solutes (leaks) is often advanced as an explanation. Here an alternative is discussed. The net accumulation of solute may be so slow that equilibrium can never be attained at finite times (e.g., several hours). Cotransporters may exhibit strong product inhibition, and the net influx of solute approaches zero far from equilibrium. The inherent slowness of net transport under these conditions is termed catalytic inefficiency. The likelihood that galactoside: H⁺ cotransport inEscherichia coli, hexose: H⁺ cotransport inChlorella vulgaris, andd-glucose: Na⁺ cotransport in brush-border membranes exhibit catalytic inefficiency is examined. The existence of strong product inhibition complicates the determination of the stoichiometry of cotransport and the characterization of chemically modified or mutant cotransporters.     

Structural and functional analysis of the single-strand origin of replication from the lactococcal plasmid pWV01

The single-strand origin (SSO) of the rolling-circle (RC), broad-host-range lactococcal plasmid pWVO1 was functionally characterized. The activity of this SSO in the conversion of single-stranded DNA to double-stranded DNA was tested both in vivo and in vitro. In addition, the effect of this SSO on plasmid maintenance was determined. The functional pWVO1 SSO comprises a 250 by region, containing two inverted repeats (IRs). The activity of each IR was tested, separately and in combination, in a plasmid derivative that was otherwise completely devoid of structures that might function as SSO. One of the IRs (IR 1) showed some homology with other previously described SSOs of the SSO_A type, as well as with the conversion signal of the Escherichia coli phage X174. This IR was shown to have a partial, RNA polymerise-independent activity in complementary strand synthesis, both in vivo and in vitro. The second IR, which had no activity of its own, was required for full SSO activity, both in vivo and in vitro. The conversion of single-stranded DNA to the double-stranded form by the complete SSO was only partly sensitive to inhibition by rifampicin, indicating the existence of an RNA polymerase-independent pathway for this event. The results suggest that the pWVO1 SSO can be activated by two different routes: an RNA polymerise-dependent one (requiring the entire SSO), and an RNA polymerase-independent one (requiring only IR I).     

链霉菌质粒的复制与进化

近年来, 随着大质粒提取和检测技术的发展, 尤其是高通量DNA测序技术的应用, 使得链霉菌大的环型质粒和线型质粒的研究取得了较快的进展。相比于研究透彻的细菌Theta型复制的质粒, 链霉菌Theta型质粒在复制区的结构、复制蛋白和调控蛋白作用的分子机理等方面具有多样性和新颖性。新鉴定的许多线型质粒的中心复制区表明中心复制的起始可以靠近端粒, 一个质粒也可以有2个以上的复制区。新分离的端粒序列显示端粒“折返”不是必需的, 而形成二级结构对于端粒复制是重要的。链霉菌环型和线型质粒的测序分析显示它们之间存在亲缘关系。环型质粒可以与噬菌体共整合, 实验证明它们在一定条件下可以相互转换。这些研究结果表明, 链霉菌环型、线型质粒和噬菌体从结构到功能到进化具有多样性、新颖性和亲缘关系。     

Metabolism in 1,3‐propanediol fed‐batch fermentation by a D‐lactate deficient mutant of Klebsiella pneumoniae

Klebsiella pneumoniae HR526, a new isolated 1,3‐propanediol (1,3‐PD) producer, exhibited great productivity. However, the accumulation of lactate in the late‐exponential phase remained an obstacle of 1,3‐PD industrial scale production. Hereby, mutants lacking D ‐lactate pathway were constructed by knocking out the ldhA gene encoding fermentative D ‐lactate dehydrogenase (LDH) of HR526. The mutant K. pneumoniae LDH526 with the lowest LDH activity was studied in aerobic fed‐batch fermentation. In experiments using pure glycerol as feedstock, the 1,3‐PD concentrations, conversion, and productivity increased from 95.39 g L^?1, 0.48 and 1.98 g L^?1 h^?1 to 102. 06 g L^?1, 0.52 mol mol^?1 and 2.13 g L^?1 h^?1, respectively. The diol (1,3‐PD and 2,3‐butanediol) conversion increased from 0.55 mol mol^?1 to a maximum of 0.65 mol mol^?1. Lactate would not accumulate until 1,3‐PD exceeded 84 g L^?1, and the final lactate concentration decreased dramatically from more than 40 g L^?1 to <3 g L^?1. Enzymic measurements showed LDH activity decreased by 89–98% during fed‐batch fermentation, and other related enzyme activities were not affected. NADH/NAD⁺ enhanced more than 50% in the late‐exponential phase as the D ‐lactate pathway was cut off, which might be the main reason for the change of final metabolites concentrations. The ability to utilize crude glycerol from biodiesel process and great genetic stability demonstrated that K. pnemoniae LDH526 was valuable for 1,3‐PD industrial production. Biotechnol. Bioeng. 2009; 104: 965–972. © 2009 Wiley Periodicals, Inc.     

Structural dynamics of the mitochondrial ADP/ATP carrier revealed by molecular dynamics simulation studies

The mitochondrial adenosine diphosphate/adenosine triphosphate (ADP/ATP) carrier has been recently crystallized in complex with its specific inhibitor carboxyatractyloside (CATR). In the crystal structure, the six-transmembrane helix bundle that defines the nucleotide translocation pathway is closed on the matrix side due to sharp kinks in the odd-numbered helices. The closed conformation is further sealed by the loops protruding into the matrix that interact through an intricate network of charge-pairs. To gain insight into its structural dynamics we performed molecular dynamics (MD) simulation studies of the ADP/ATP carrier with and without its cocrystallized inhibitor. The two trajectories sampled a conformational space around two different configurations characterized by distinct salt-bridge networks with a significant shift from inter- to intrarepeat bonding on the matrix side in the absence of CATR. Analysis of the geometrical parameters defining the transmembrane helices showed that even-numbered helices can undergo a face rotation, whereas odd-numbered helices can undergo a change in the wobble angle with a conserved proline acting as molecular hinge. Our results provide new information on the dynamical properties of the ADP/ATP carrier and for the first time yield a detailed picture of a stable carrier conformation in absence of the inhibitor.     

Biogenesis of Respiratory Complex I

Proteins specifically involved in the biogenesis of respiratory complex I in eukaryotes have been characterized. The complex I intermediate associated proteins CIA30 and CIA84 are tightly bound to an assembly intermediate of the membrane arm. Like chaperones, they are involved in multiple rounds of membrane arm assembly without being part of the mature structure. Two biosynthetic subunits of eukaryotic complex I have been characterized. The acyl carrier subunit is needed for proper assembly of the peripheral arm as well as the membrane arm of complex I. It may interact with enzymes of a mitochondrial fatty acid synthetase. The 39/40-kDa subunit appears to be an isomerase with a tightly bound NADPH. It is related to a protein family of reductases/isomerases. Both subunits have been discussed to be involved in the synthesis of a postulated, novel, high-potential redox group.     

Knockdown of SLC34A2 inhibits cell proliferation,metastasis, and elevates chemosensitivity in glioma

Solute carrier 34 A2 (SLC34A2) is a member of SLC34 family that is a group of phosphate transporters. SLC34A2 has been reported to play critical roles in tumorigenesis and progression. However, the researches about the biological roles of SLC34A2 in glioma have not yet been reported. In this study, we analyzed the expression patterns of SLC34A2 in clinical glioma tumor tissues and cell lines. The results demonstrated that SLC34A2 was generally overexpressed in both glioma tissues and cell lines. To further investigate the roles of SLC34A2 in glioma, lentivirus containing specific SLC34A2 short hairpin RNA (sh-SLC34A2) was used to infect glioma cell lines U251 and U87 for the knockdown of SLC34A2. The following studies proved that SLC34A2 knockdown exhibited suppressive effects on cell proliferation and migration/invasion. SLC34A2 knockdown also inhibited epithelial-mesenchymal transition (EMT) phenotype, as evidenced by the increased E-cadherin expression, and the decreased N-cadherin and fibronectin expressions. Besides, knockdown of SLC34A2 enhanced the temozolomide (TMZ) sensitivity of U251 and U87 cells. In vivo tumorigenicity assay demonstrated that SLC34A2 knockdown inhibited tumor growth. Moreover, SLC34A2 knockdown suppressed the activation of epidermal growth factor receptor (EGFR)/PI3K/AKT signaling pathway in U87 cells. GW2974 (EGFR inhibitor) increased SLC34A2 knockdown-inhibited cell proliferation, migration/invasion, as well as enhanced SLC34A2 knockdown-increased the TMZ sensitivity of glioma cells. These findings suggested that SLC34A2 might be a new potential therapeutic target for the therapy of glioma patients.