FANCJ/BRIP1 recruitment and regulation of FANCD2 in DNA damage responses

FANCJ/BRIP1 encodes a helicase that has been implicated in the maintenance of genomic stability. Here, to better understand FANCJ function in DNA damage responses, we have examined the regulation of its cellular localization. FANCJ nuclear foci assemble spontaneously during S phase and are induced by various stresses. FANCJ foci colocalize with the replication fork following treatment with hydroxyurea, but not spontaneously. Using FANCJ mutants, we find that FANCJ helicase activity and the capacity to bind BRCA1 are both involved in FANCJ recruitment. Given similarities to the recruitment of another Fanconi anemia protein, FANCD2, we tested for colocalization of FANCJ and FANCD2. Importantly, these proteins show substantial colocalization, and FANCJ promotes the assembly of FANCD2 nuclear foci. This process is linked to the proper localization of FANCJ itself since both FANCJ and FANCD2 nuclear foci are compromised by FANCJ mutants that abrogate its helicase activity or interaction with BRCA1. Our results suggest that FANCJ is recruited in response to replication stress and that FANCJ/BRIP1 may serve to link FANCD2 to BRCA1.     

DOA1/UFD3 plays a role in sorting ubiquitinated membrane proteins into multivesicular bodies

Ubiquitin (Ub) is a sorting signal that targets integral membrane proteins to the interior of the vacuole/lysosome by directing them into lumenal vesicles of multivesicular bodies (MVBs). The Vps27-Hse1 complex, which is homologous to the Hrs-STAM complex in mammalian cells, serves as a Ub-sorting receptor at the surface of early endosomes. We have found that Hse1 interacts with Doa1/Ufd3. Doa1 is known to interact with Cdc48/p97 and Ub and is required for maintaining Ub levels. We find that the Hse1 Src homology 3 domain binds directly to the central PFU domain of Doa1. Mutations in Doa1 that block Hse1 binding but not Ub binding do not alter Ub levels but do result in the missorting of the MVB cargo GFP-Cps1. Loss of Doa1 also causes a synthetic growth defect when combined with loss of Vps27. Unlike the loss of Doa1 alone, the doa1Delta vps27Delta double mutant phenotype is not suppressed by Ub overexpression, demonstrating that the effect is not due to indirect consequence of lowered Ub levels. Loss of Doa1 results in a defect in the accumulation of GFP-Ub within yeast vacuoles, implying that there is a reduction in the flux of ubiquitinated membrane proteins through the MVB pathway. This defect was also reflected by an inability to properly sort Vph1-GFP-Ub, a modified subunit of the multiprotein vacuolar ATPase complex, which carries an in-frame fusion of Ub as an MVB sorting signal. These results reveal novel roles for Doa1 in helping to process ubiquitinated membrane proteins for sorting into MVBs.     

黄连木五倍子的生态研究与利用

一、产地自然概况黄连木五倍子最早发现于河南省南部信阳市郊贤山脚下(东经114°0′,,北纬32°10′)。该地为低山丘陵,海拔90一320m,南部紧连鸡公山厂傍依南湾水库和狮河。     

Improved eicosapentaenoic acid production in Pythium splendens RBB-5 based on metabolic regulation analysis

Applied Microbiology and Biotechnology - Eicosapentaenoic acid (EPA) is an essential polyunsaturated fatty acid for human beings. At present, the production of commercially available long-chain...     

Mapping and validation of a new QTL for adult-plant resistance to powdery mildew in Chinese elite bread wheat line Zhou8425B

Key message

Four QTLs for adult-plant resistance to powdery mildew were mapped in the Zhou8425B/Chinese Spring population, and a new QTL on chromosome 3B was validated in 103 wheat cultivars derived from Zhou8425B.

Abstract

Zhou8425B is an elite wheat (Triticum aestivum L.) line widely used as a parent in Chinese wheat breeding programs. Identification of genes for adult-plant resistance (APR) to powdery mildew in Zhou8425B is of high importance for continued controlling the disease. In the current study, the high-density Illumina iSelect 90K single-nucleotide polymorphism (SNP) array was used to map quantitative trait loci (QTL) for APR to powdery mildew in 244 recombinant inbred lines derived from the cross Zhou8425B/Chinese Spring. Inclusive composite interval mapping identified QTL on chromosomes 1B, 3B, 4B, and 7D, designated as QPm.caas-1BL.1, QPm.caas-3BS, QPm.caas-4BL.2, and QPm.caas-7DS, respectively. Resistance alleles at the QPm.caas-1BL.1, QPm.caas-3BS, and QPm.caas-4BL.2 loci were contributed by Zhou8425B, whereas that at QPm.caas-7DS was from Chinese Spring. QPm.caas-3BS, likely to be a new APR gene for powdery mildew resistance, was detected in all four environments. One SNP marker closely linked to QPm.caas-3BS was transferred into a semi-thermal asymmetric reverse PCR (STARP) marker and tested on 103 commercial wheat cultivars derived from Zhou8425B. Cultivars with the resistance allele at the QPm.caas-3BS locus had averaged maximum disease severity reduced by 5.3%. This STARP marker can be used for marker-assisted selection in improvement of the level of powdery mildew resistance in wheat breeding.

     

Revealing the Inhibitory Effect of Ginseng on Mitochondrial Respiration through Synaptosomal Proteomics

Ginseng, the active ingredients of which are ginsenosides, is the most popular herbal medicine and has potential merit in the treatment of cerebral disorders. To better understand the function of Ginseng in the cerebral system, we examined changes in the protein expression profiles of synaptosomes extracted from the cerebral cortical and hippocampal tissues of rats administered a high or low dose of Ginseng for 2 weeks. More than 5000 proteins belonging to synaptosomes were simultaneously identified and quantitated by an approach combining tandem mass tags with 2D liquid chromatography‐mass spectrometry (LC‐MS). Regarding differentially expressed proteins, downregulated proteins were much more highly induced than upregulators in the cerebral cortical and hippocampal synaptosomes, regardless of the dose of Ginseng. Bioinformatic analysis indicated the majority of the altered proteins to be located in the mitochondria, directly or indirectly affecting mitochondrial oxidative respiration. Further functional experiments using the substrate‐uncoupler inhibitor titration approach confirmed that three representative ginsenosides were able to inhibit oxidative phosphorylation in mitochondria. Our results demonstrate that Ginseng can regulate the function of mitochondria and alter the energy metabolism of cells, which may be useful for the treatment of central nervous disorders.     

Statistical analysis of the Bacterial Carbohydrate Structure Data Base (BCSDB): Characteristics and diversity of bacterial carbohydrates in comparison with mammalian glycans

Background  

There are considerable differences between bacterial and mammalian glycans. In contrast to most eukaryotic carbohydrates, bacterial glycans are often composed of repeating units with diverse functions ranging from structural reinforcement to adhesion, colonization and camouflage. Since bacterial glycans are typically displayed at the cell surface, they can interact with the environment and, therefore, have significant biomedical importance.     

Cells scaffold complex for Intervertebral disc Anulus Fibrosus tissue engineering: in vitro culture and product analysis

The study was designed to investigate feasibility of tissue culture in vitro utilizing static culture method. Annulus fibrosus cells obtained from spine of rabbits were cultured. Results showed that fibrous tissue infiltration could be detected in shallow layer. With extended time, tissue infiltration depth increased, but there were still a large amount of holes in central part. Fibrous tissue infiltration was detected in the control side products and inner infiltration wasn't obvious. Hydroxyproline content of the control side products gradually increased with extended culture time. Hydroxyproline content of the control side products in the third and fourth month was significantly higher than that in the first month, but lower than those of the experimental side products and normal annulus fibrosus cells. DNA content of the control side products in the third and fourth month was significantly increased compared to the first month. DNA content of the control side products at each phase point was significantly lower than that of the experimental side and normal annulus fibrosus cells. Furthermore, there was lower expression levels of the type I, II collagen mRNA and protein in the experimental side scaffolds compared to the control side product. This study demonstrates the successful formation of Intervertebral disc Anulus Fibrosus in vitro by static culture method.     

GABAergic disinhibition facilitates polysynaptic excitatory transmission in rat anterior cingulate cortex

Various studies implicate the anterior cingulate cortex (ACC) in processing pain. Combining whole-cell patch clamp recordings in rat ACC slices and a formalin-induced conditioned place avoidance (F-CPA) behavioral model, the present study was to address the effect of GABA(A) receptors on excitatory transmission to ACC layer V neurons and its possible functional significance related to pain. Removal of GABA(A) inhibition by bicuculline (10 microM) induced a novel long-lasting response in layer V neurons, which could be blocked by high divalent extracellular solution and was sensitive to relatively higher rate stimuli. Co-application of NMDA receptor antagonist APV (50 microM) and non-NMDA receptor antagonist DNQX (10 microM) completely blocked the responses. Enhancement of inhibition by intra-ACC microinjection of muscimol abolished the acquisition of F-CPA without affecting formalin-induced acute nociceptive responses. These results suggest that GABA(A) inhibition may be involved in pain-related aversion by modulating glutamate-mediated excitatory transmission in the ACC.