A multiprotein nuclear complex connects Fanconi anemia and Bloom syndrome

Bloom syndrome (BS) is a genetic disorder associated with dwarfism, immunodeficiency, reduced fertility, and an elevated risk of cancer. To investigate the mechanism of this disease, we isolated from human HeLa extracts three complexes containing the helicase defective in BS, BLM. Interestingly, one of the complexes, termed BRAFT, also contains five of the Fanconi anemia (FA) complementation group proteins (FA proteins). FA resembles BS in genomic instability and cancer predisposition, but most of its gene products have no known biochemical activity, and the molecular pathogenesis of the disease is poorly understood. BRAFT displays a DNA-unwinding activity, which requires the presence of BLM because complexes isolated from BLM-deficient cells lack such an activity. The complex also contains topoisomerase IIIalpha and replication protein A, proteins that are known to interact with BLM and could facilitate unwinding of DNA. We show that BLM complexes isolated from an FA cell line have a lower molecular mass. Our study provides the first biochemical characterization of a multiprotein FA complex and suggests a connection between the BLM and FA pathways of genomic maintenance. The findings that FA proteins are part of a DNA-unwinding complex imply that FA proteins may participate in DNA repair.     

Inhibition of nicotinoprotein (NAD+-containing) alcohol dehydrogenase by trans-4-(N,N-dimethylamino)-cinnamaldehyde binding to the active site

Ethanol oxidation by nicotinoprotein alcohol dehydrogenase (np-ADH) from the bacterium Amycolatopsis methanolica is inhibited by trans-4-(N,N-dimethylamino)-cinnamaldehyde through direct binding to the catalytic zinc ion in a substrate-like geometry. This binding is accompanied by a characteristic red shift of the aldehyde absorbance from 398 nm to 467 nm. Np-ADH is structurally related to mammalian ADH class I, and a model of np-ADH shows how the cinnamaldehyde derivative can be accommodated in the active site of the nicotinoprotein, correlating the structural and enzymological data.     

Differential cytokine expression in myocytes and non-myocytes after myocardial infarction in rats

The proinflammatory cytokines interleukin (IL)-1 and IL-6 are increased after acute myocardial infarction (MI). Moreover, serum IL-6 level is elevated after MI, but has also been associated with heart failure. In the present study, heart function was monitored in a rat model of chronic MI. Cytokine expression in the infarcted and non-infarcted myocardium as well as in hearts of sham-operated controls was measured by the ribonuclease-protection assay. To identify the cells contributing to the increased cytokine expression, we further analyzed myocytes and non-myocytes isolated in the acute phase as well as during congestive heart failure (CHF) after MI. There was a strong induction in cytokine expression in the myocytes of the infarct area 6 h after MI. In the non-infarcted myocardium, cytokine expression increased only slightly in the non-myocytes after 6 h. This was not different from sham-operated controls and may, therefore, be induced by stress and catecholamines. In CHF, however, cytokine expression level in myocytes was normal. It increased slightly but significantly in the non-myocytes 4 and 8 weeks after MI. In conclusion, we suggest that pro-inflammatory cytokines, produced by the ischemic myocytes may be involved in the initiation of wound healing of the necrotic area, whereas the effect of pro-inflammatory cytokines in CHF, if any, seems not to be crucial.     

Lack of enhanced spinal regeneration in Nogo-deficient mice

The failure of regeneration of severed axons in the adult mammalian central nervous system is thought to be due partly to the presence of endogenous inhibitors of axon regeneration. The nogo gene encodes three proteins (Nogo-A, -B, and -C) that have been proposed to contribute to this inhibition. To determine whether deletion of nogo enhances regenerative ability, we generated two lines of mutant mice, one lacking Nogo-A and -B but not -C (Nogo-A/B mutant), and one deficient in all three isoforms (Nogo-A/B/C mutant). Although Nogo-A/B-deficient myelin has reduced inhibitory activity in a neurite outgrowth assay in vitro, tracing of corticospinal tract fibers after dorsal hemisection of the spinal cord did not reveal an obvious increase in regeneration or sprouting of these fibers in either mouse line, suggesting that elimination of Nogo alone is not sufficient to induce extensive axon regeneration.     

Persistent use of false myeloma cell lines

Multiple myeloma (MM) is a neoplasm of a terminally differentiated B-cell. Human myeloma cell lines were shown to be suitable model systems for use in various fields of the biological sciences. Within the last 20 years more than 100 cell lines have been established. So-called 'myeloma cell lines' have been previously reported and are still widely used which are in reality Epstein-Barr virus (EBV)-positive B-lymphoblastoid cell lines. The presence of the EBV-genome in residual normal B-cells provides them with a selective growth advantage after explantation. Cell lines represent an extremely important resource for research in a variety of fields and disciplines. As the cell lines are used as in vitro model systems in lieu of primary material, it is crucial that the cells in the culture flasks faithfully correspond to the purported objects of study. On closer examination, the use of false cell lines may be seen to invalidate a significant percentage of scientific work, or at least cast doubts on the relevance of these in vitro results to the cell type or tumor in vivo. Ultimately, use of cross-contaminated cell lines is a waste of human and material resources. Henceforth, it should be mandatory to prove the proper derivation of each new cell line by comparing DNA fingerprints or karyotypes of the patient's primary cells and the cultured cells. The availability of well characterized and authenticated bona fide MM cell lines is of great importance for the study of the biology, etiology and treatment of the disease.     

Simultaneous measurement of protein one-bond and two-bond nitrogen-carbon coupling constants using an internally referenced quantitative J-correlated [(15)N,(1)H]-TROSY-HNC experiment

A quantitative J-correlation pulse sequence is described that allows simultaneous determination of one-bond and two-bond nitrogen-carbon coupling constants for protonated or deuterated proteins. Coupling constants are calculated from volume ratios between cross peaks and reference axial peaks observed in a single 3D spectrum. Accurate backbone ¹ J _NC, ¹ J _NC, and ² J _NC coupling constants are obtained for the two [¹⁵N;¹³C]-labeled, medium-sized proteins flavodoxin and xylanase and for the [²H;¹⁵N;¹³C]-labeled, large protein DFPase. A dependence of one-bond and two-bond J _NC values on protein backbone torsion angles is readily apparent, in agreement with previously found correlations. In addition, the experiment is performed on isotropic as well as aligned protein to measure associated ¹⁵N-¹³C residual dipolar couplings.     

The circling behavior of the deafblind LEW-<Emphasis Type="Italic">ci2</Emphasis> rat is linked to a segment of RNO10 containing <Emphasis Type="Italic">Myo15</Emphasis> and <Emphasis Type="Italic">Kcnj12</Emphasis>

The LEW/Ztm-ci2 rat is an autosomal recessive mutant that displays circling behavior, deafness, progressive retinopathy, locomotor hyperactivity, ataxia, and opisthotonus. We performed a genome-wide scan of a (LEW/Ztm-ci2 × BN/Ztm) F₁ × LEW/Ztm-ci2 backcross population with anonymous microsatellite markers to analyze the genetics of this mutant rat. This linkage analysis demonstrated a very strong association of RNO10 SSLP markers to the phenotype with a core region in the central part of the chromosome. The knowledge of genes mapping to this part of the rat genome and their linkage to SSLP markers is still poor. We developed SSLP markers closely linked to genes, which might be responsible for the mutant phenotype by using the growing amount of rat-specific DNA sequences available at World Wide Web databases. Application of this method facilitated the search for candidate genes for the phenotype of the LEW-ci2 rat. We were able to map Myo15 and its neighboring genes, Znf179 and Aldh3a1, to the region of interest and Myo1c to a more distal location on RNO10. Further rat BAC clones were used to create a physical map of the region of interest. This map revealed the position of further genes. Among those is Kcnj12. Owing to their localization on RNO10 and their involvement in a similar pathology in human and mouse, Myo15 and Kcnj12 can be regarded as candidate genes for the deafblind phenotype of the LEW-ci2 rat.     

Arbuscular mycorrhizal fungi-parasite-host interaction for the control of<Emphasis Type="Italic"> Striga hermonthica</Emphasis> (Del.) Benth. in sorghum [<Emphasis Type="Italic">Sorghum bicolor</Emphasis> (L.) Moench]

Five Glomus species (G. intraradices, G. albidum, G. mosseae, G. fasciculatum, and G. etunicatum) were compared against a check [without arbuscular mycorrhizal (AM) fungi, plus Striga] and control (without AM fungi or Striga) treatments for the control of Striga in a tolerant sorghum variety (War-wara bashi) in an experiment carried out in 12-cm-diameter clay pots. The experiment was carried out in a controlled growth chamber. G. mosseae significantly reduced the number of Striga emerging per plant, increased plant growth, shoot and total dry matter yield of sorghum, did not affect the root dry matter compared with the other AM fungi species, but had a comparable effect to the control treatment. All the AM fungi except G. mosseae, and also the Striga-infested treatment, increased the root:shoot ratio compared to the control treatment. The percent reduction (62%) of Striga emergence after G. mosseae inoculation resulted in about a 30% increase in total dry matter yield of sorghum over the control, while the total loss in dry matter yield of sorghum due to Striga infestation was 36%. Root colonization of sorghum by AM fungi was highest for G. mosseae (44%) followed by G. intraradices (24%) and G. albidum (23%) then G. fasciculatum (18%), with the lowest recorded for G. etunicatum (14%). No colonization of Striga roots was observed. The potential of AM fungi to reduce or to compensate for Striga infestation could be important for soil management, especially in the tropics, and for the reduction of Striga-resistant varieties of sorghum which are mycorrhiza-responsive.     

A thin-film electrochemical study of the "blue" copper proteins,auracyanin A and auracyanin B,from the photosynthetic bacterium <Emphasis Type="Italic">Chloroflexus</Emphasis><Emphasis Type="Italic">aurantiacus</Emphasis>: the reduction potential as a function of pH

The reversible formal potentials of auracyanin A and auracyanin B, two closely related "blue" copper proteins from the photosynthetic bacterium Chloroflexus aurantiacus, have been determined by protein film voltammetry in the range 4相似文献