尖孢镰孢一新硝酸盐营养突变类型与营养体亲和性

从73个尖孢镰孢(Fusarium oxysporum)不同专化型菌株上获得684个硝酸盐营养突变株(nit mutant)。作相关氮源利用试验及亚硝酸反应后,鉴定出一新硝酸盐营养突变类型:亚硝酸盐还原酶结构基因类型,命名为nit8,占总突变株的6.7%。同时被鉴别的还有nit1、nit3和Nit M三种突变类型,它们分别占突变株总数的81.0%,3.8%和8.5%。此外,首次引入一种亚硝酸反应在这类研究中的应用,还提出了互补指数概念与公式来表示nit突变株营养体之间亲和的能力。     

The dprA gene is required for natural transformation of Helicobacter pylori

Genetic recombination in Helicobacter pylori is believed to be involved in host adaptation of this gastric pathogen and uptake of DNA by natural transformation can result in changes in virulence factors as well as antigenic variation. To elucidate the mechanisms involved in natural transformation we tested two genes with homology to known competence genes (dprA and traG) for their role in this process. Insertion mutants in these genes were constructed in two different H. pylori strains and their competence by natural transformation was compared to the wild-type. Mutation of the traG homolog did not reduce competence. Mutation of the dprA gene, however, severely impaired natural transformation both with plasmid and chromosomal DNA. Our data indicate that dprA and comB3 are essential parts of a common pathway for chromosomal and plasmid transformation.     

Exaggerated root respiration accounts for growth retardation in a starchless mutant of Arabidopsis thaliana

The knock‐out mutation of plastidial phosphoglucomutase (pgm) causes a starchless phenotype in Arabidopsis thaliana, and results in a severe growth reduction of plants cultivated under diurnal conditions. It has been speculated that high soluble sugar levels accumulating during the light phase in leaf mesophyll might cause a reduction of photosynthetic activity or that shortage of reduced carbon during the night is the reason for the slow biomass gain of pgm. Separate simultaneous measurements of leaf net photosynthesis and root respiration demonstrate that photosynthetic activity per unit fresh weight is not reduced in pgm, whereas root respiration is strongly elevated. Comparison with a mutant defective in the dominating vacuolar invertase (AtβFruct4) revealed that high sucrose concentration in the cytosol, but not in the vacuole, of leaf cells is responsible for elevated assimilate transport to the root. Increased sugar supply to the root, as observed in pgm mutants, forces substantial respiratory losses. Because root respiration accounts for 80% of total plant respiration under long‐day conditions, this gives rise to retarded biomass formation. In contrast, reduced vacuolar invertase activity leads to reduced net photosynthesis in the shoot and lowered root respiration, and affords an increased root/shoot ratio. The results demonstrate that roots have very limited capacity for carbon storage but exert rigid control of supply for their maintenance metabolism.     

Charge states rather than propensity for beta-structure determine enhanced fibrillogenesis in wild-type Alzheimer's beta-amyloid peptide compared to E22Q Dutch mutant

The activity of the Alzheimer's amyloid beta-peptide is a sensitive function of the peptide's sequence. Increased fibril elongation rate of the E22Q Dutch mutant of the Alzheimer's amyloid beta-peptide relative to that of the wild-type peptide has been observed. The increased activity has been attributed to a larger propensity for the formation of beta structure in the monomeric E22Q mutant peptide in solution relative to the WT peptide. That hypothesis is tested using four nanosecond timescale simulations of the WT and Dutch mutant forms of the Abeta(10-35)-peptide in aqueous solution. The simulation results indicate that the propensity for formation of beta-structure is no greater in the E22Q mutant peptide than in the WT peptide. A significant measure of "flickering" of helical structure in the central hydrophobic cluster region of both the WT and mutant peptides is observed. The simulation results argue against the hypothesis that the Dutch mutation leads to a higher probability of formation of beta-structure in the monomeric peptide in aqueous solution. We propose that the greater stability of the solvated WT peptide relative to the E22Q mutant peptide leads to decreased fibril elongation rate in the former. Stability difference is due to the differing charge state of the two peptides. The other proposal leads to the prediction that the fibril elongation rates for the WT and the mutant E22Q should be similar under acid conditions.     

Single amino acid substitution in HIV-1 integrase catalytic core causes a dramatic shift in inhibitor selectivity

HIV-1 integrase (IN) mediates the insertion of viral cDNA into the cell genome, a vital process for replication. This step is catalyzed by two separate DNA reaction events, termed 3'-processing and strand transfer. Here, we show that six inhibitors from five structurally different classes of compounds display a selectivity shift towards preferential strand transfer inhibition over the 3'-processing activity of IN when a single serine is substituted at position C130. Even though IN utilizes the same active site for both reactions, this finding suggests a distinct conformational dissimilarity in the mechanistic details of each IN catalytic event.     

Phosphorylation of NHERF1 S279 and S301 differentially regulates breast cancer cell phenotype and metastatic organotropism

Metastatic cancer cells are highly plastic for the expression of different tumor phenotype hallmarks and organotropism. This plasticity is highly regulated but the dynamics of the signaling processes orchestrating the shift from one cell phenotype and metastatic organ pattern to another are still largely unknown. The scaffolding protein NHERF1 has been shown to regulate the expression of different neoplastic phenotypes through its PDZ domains, which forms the mechanistic basis for metastatic organotropism. This reprogramming activity was postulated to be dependent on its differential phosphorylation patterns. Here, we show that NHERF1 phosphorylation on S279/S301 dictates several tumor phenotypes such as in vivo invasion, NHE1-mediated matrix digestion, growth and vasculogenic mimicry. Remarkably, injecting mice with cells having differential NHERF1 expression and phosphorylation drove a shift from the predominantly lung colonization (WT NHERF1) to predominately bone colonization (double S279A/S301A mutant), indicating that NHERF1 phosphorylation also acts as a signaling switch in metastatic organotropism.     

Mechanism of regulating the expression of λN gene by ribosomal protein at translational level

In ribosomal protein S12 mutant or L24 mutant the expression of λN gene was depressed at translational level. To study its mechanism the λN gene region of λN -lacZ gene fusion was trimmed from its 5′ end to 3′ end with DNA exonuclease III (DNA cxoIII) in order to alter the TIR (translational initiation region) and the ding region of λN gene. After DNA sequencing 23 species of different λN-lacZ fused genes were obtained. The β-galactosidase activities of these deletants in ribosomal protein mutant were compared with that in wild type strain. The result indicated that (i) S12 mutant could affect 305 subunit’s binding to the TIR of λN gene messenger and cause the difficulty in forming 30s initiation complex and then decrease the efficiency of translational initiation; (ii) in S12 mutant the coding region of λN gene alw affected the expression λN gene; (iii) in L24 mutant the inhibition of λN gene expression was not related to translational initiation and the 5′ end of the coding region of λN gene, but related to the 3′ end of λN gene. Project supported by the National Natural Science Foundation of China (Grant Nos. 39480014, 39570162) and Chinese Academy of Sciences.     

The dosage effect of the wildtype GBSS allele is linear for GBSS activity but not for amylose content: absence of amylose has a distinct influence on the physico-chemical properties of starch

A gene-dosage population was obtained by crossing two genotypes that were duplex for the GBSS allele. Nulliplex, simplex, duplex or triplex/quadruplex plants could be identified by monitoring the segregation of red and blue microspores after staining with iodine. GBSS activity was significantly different for all groups and showed an almost linear dosage effect for the wildtype GBSS gene. A dosage effect was found for amylose content that was not linear. The amylose content was similar for both the duplex and triplex/quadruplex group. Within the simplex group, differences in amylose content were found, which might be due to a different genetic background. There was no linear correlation between GBSS activity and amylose content. A certain level of GBSS activity led to a maximum amount of amylose, and further increase in GBSS activity did not result in a further increase in amylose content. The presence of one or more wildtype GBSS allele(s), and therefore the presence of amylose in the starch granules, had a great influence on the physico-chemical properties of the starch suspensions.     

拟南芥耐盐突变体stg2的筛选

采取在高盐平板上萌发的方法,对一个雌激素诱导激活型拟南芥突变体库进行了耐盐突变体的筛选,最终得到了2株稳定的耐盐突变体。本文中对其中的一株耐盐突变体,命名为stg2(salt tolerance during germination 2),进行了研究。遗传实验表明它的耐盐特性是受雌激素诱导的,是功能获得型的耐盐突变体。本实验中还探讨了stg2突变体的筛选过程及耐盐生理特点。     

Heterooligomeric complexes formed by human small heat shock proteins HspB1 (Hsp27) and HspB6 (Hsp20)

Formation of heterooligomeric complexes of human small heat shock proteins (sHsp) HspB6 (Hsp20) and HspB1 (Hsp27) was analyzed by means of native gel electrophoresis, analytical ultracentrifugation, chemical cross-linking and size-exclusion chromatography. HspB6 and HspB1 form at least two different complexes with apparent molecular masses 100–150 and 250–300 kDa, and formation of heterooligomeric complexes is temperature dependent. These complexes are highly mobile, easily exchange their subunits and are interconvertible. The stoichiometry of HspB1 and HspB6 in both complexes is close to 1/1 and smaller complexes are predominantly formed at low, whereas larger complexes are predominantly formed at high protein concentration. Formation of heterooligomeric complexes does not affect the chaperone-like activity of HspB1 and HspB6 if insulin or skeletal muscle F-actin was used as model protein substrates. After formation of heterooligomeric complexes the wild type HspB1 inhibits the rate of phosphorylation of HspB6 by cAMP-dependent protein kinase. The 3D mutant mimicking phosphorylation of HspB1 also forms heterooligomeric complexes with HspB6, but is ineffective in inhibition of HspB6 phosphorylation. Inside of heterooligomeric complexes HspB6 inhibits phosphorylation of HspB1 by MAPKAP2 kinase. Thus, in heterooligomeric complexes HspB6 and HspB1 mutually affect the structure of each other and formation of heterooligomeric complexes might influence diverse processes depending on small heat shock proteins.