Binding of the erlin1/2 complex to the third intralumenal loop of IP3R1 triggers its ubiquitin-proteasomal degradation

Long-term activation of inositol 1,4,5-trisphosphate receptors (IP₃Rs) leads to their degradation by the ubiquitin–proteasome pathway. The first and rate-limiting step in this process is thought to be the association of conformationally active IP₃Rs with the erlin1/2 complex, an endoplasmic reticulum–located oligomer of erlin1 and erlin2 that recruits the E3 ubiquitin ligase RNF170, but the molecular determinants of this interaction remain unknown. Here, through mutation of IP₃R1, we show that the erlin1/2 complex interacts with the IP₃R1 intralumenal loop 3 (IL3), the loop between transmembrane (TM) helices 5 and 6, and in particular, with a region close to TM5, since mutation of amino acids D-2471 and R-2472 can specifically block erlin1/2 complex association. Surprisingly, we found that additional mutations in IL3 immediately adjacent to TM5 (e.g., D2465N) almost completely abolish IP₃R1 Ca²⁺ channel activity, indicating that the integrity of this region is critical to IP₃R1 function. Finally, we demonstrate that inhibition of the ubiquitin-activating enzyme UBE1 by the small-molecule inhibitor TAK-243 completely blocked IP₃R1 ubiquitination and degradation without altering erlin1/2 complex association, confirming that association of the erlin1/2 complex is the primary event that initiates IP₃R1 processing and that IP₃R1 ubiquitination mediates IP₃R1 degradation. Overall, these data localize the erlin1/2 complex–binding site on IP₃R1 to IL3 and show that the region immediately adjacent to TM5 is key to the events that facilitate channel opening.     

Biotechnological challenges of bioartificial kidney engineering

With the world-wide increase of patients with renal failure, the development of functional renal replacement therapies have gained significant interest and novel technologies are rapidly evolving. Currently used renal replacement therapies insufficiently remove accumulating waste products, resulting in the uremic syndrome. A more preferred treatment option is kidney transplantation, but the shortage of donor organs and the increasing number of patients waiting for a transplant warrant the development of novel technologies. The bioartificial kidney (BAK) is such promising biotechnological approach to replace essential renal functions together with the active secretion of waste products. The development of the BAK requires a multidisciplinary approach and evolves at the intersection of regenerative medicine and renal replacement therapy. Here we provide a concise review embracing a compact historical overview of bioartificial kidney development and highlighting the current state-of-the-art, including implementation of living-membranes and the relevance of extracellular matrices. We focus further on the choice of relevant renal epithelial cell lines versus the use of stem cells and co-cultures that need to be implemented in a suitable device. Moreover, the future of the BAK in regenerative nephrology is discussed.     

RELATIONSHIP BETWEEN CELL CYCLE AND LIGHT‐LIMITED GROWTH RATE IN OCEANIC PROCHLOROCOCCUS (MIT9312) AND SYNECHOCOCCUS (WH8103) (CYANOBACTERIA)1

The relationships between growth rate, cell‐cycle parameters, and cell size were examined in two unicellular cyanobacteria representative of open‐ocean environments: Prochlorococcus (strain MIT9312) and Synechococcus (strain WH8103). Chromosome replication time, C, was constrained to a fairly narrow range of values (～4–6 h) in both species and did not appear to vary with growth rate. In contrast, the pre‐ and post‐DNA replication periods, B and D, respectively, decreased with increasing growth rate from maxima of ～30 and 10–20 h to minima of ～4–6 and 2–3 h, respectively. The combined duration of the chromosome replication and postreplication periods (C+D), a quantity often used in the estimation of Prochlorococcus in situ growth rates, varied ～2.4‐fold over the range of growth rates examined. This finding suggests that assumptions of invariant C+D may adversely influence Prochlorococcus growth rate estimates. In both strains, cell mass was the greatest in slowly growing cells and decreased 2‐ to 3‐fold over the range of growth rates examined here. Estimated cell mass at the start of replication appeared to decrease with increasing growth rate, indicating that the initiation of chromosome replication in Prochlorococcus and Synechococcus is not a simple function of cell biomass, as suggested previously. Taken together, our results reflect a notable degree of similarity between oceanic Synechococcus and Prochlorococcus strains with respect to their growth‐rate‐specific cell‐cycle characteristics.     

鸡传染性喉气管炎病毒gB基因的克隆及其在耻垢分枝杆菌中的表达

以ILTV基因组为模板 ,利用PCR特异扩增出gB基因 ,定向克隆到中间质粒载体pY_α ,构建了中间质粒pY_α_gB。然后以中间质粒pY_α_gB为模板 ,扩增出含有人结核分枝杆菌启动子hsp70基因和堪萨斯分枝杆菌α信号肽基因的hsp_α_gB片段 ,回收补平后与穿梭表达载体pRR3平端连接 ,从而构建大肠杆菌_分枝杆菌穿梭表达质粒pR_α_gB。再将其电转化至耻垢分枝杆菌M .smegmatismc2 15 5 ,ELISA检测表明重组菌株M .smegmatismc2 15 5 (pR_α_gB)的表达产物具有很好的反应原性。Westernblot检测说明gB基因在分枝杆菌中获得了表达并具有良好的免疫原性。鸡胚中和试验结果表明该重组菌株可以中和 1个剂量EID50 的ILTV强毒 ,能够保护SPF鸡胚抵抗强毒攻击     

Identification of Coffee Genes Expressed During Systemic Acquired Resistance and Incompatible Interaction with Hemileia vastatrix

Coffee genes associated with systemic acquired resistance (SAR) and incompatible reaction against coffee leaf rust inoculation were identified by suppression subtractive hybridization. Analysis of 384 clones of each of the subtracted cDNA libraries identified genes involved in oxidative burst/apoptosis/hypersensitive response, synthesis of antimicrobial proteins, synthesis and transport of antimicrobial metabolites, signal perception and transduction, metabolism of lipids, regulated protein degradation and cell maintenance and development. Induction of distinct sets of genes in the two resistance responses was observed. A wide range of genes involved in defence responses described in other plant species was also found in coffee plants. Semi-quantitative and quantitative RT-PCR analysis of seven selected genes showed differences in their expression profile within 72 h after treatment. Full-length cDNA sequences of two β-1,3-glucanases, one induced during SAR and the other in the incompatible reaction, were obtained by 5' and 3' RACE and the sequence data suggest different properties and cellular localization of the encoded proteins.     

Detection of Nitrosomonas spp. by polymerase chain reaction

Abstract A unique genomic DNA fragment was isolated from Nitrosomonas europaea ATCC 19718. Based on the sequence of this fragment, oligonucleotide primers for polymerase chain reaction amplification were prepared which amplify sequences of 775 and 658 bp. The predicted DNA fragments were both amplified from the genome of N. europaea and a Nitrosomonas spp. isolated from a local oxidation pond. The primers failed to amplify DNA from the genomes of the ammonia oxidiser Nitrosolobous multiformis , the nitrite oxidiser Nitrococcus mobilis as well as from the genomes of other unrelated heterotrophic bacteria. These DNA sequences could be amplified from 0.01 ng of N. europaea genomic DNA or from 100 intact cells, and it was possible to detect Nitrosomonas DNA in a DNA mixture extracted from water samples drawn from a local oxidation pond.     

Signs of stabilisation and stable coexistence

Many empirical studies motivated by an interest in stable coexistence have quantified negative density dependence, negative frequency dependence, or negative plant–soil feedback, but the links between these empirical results and ecological theory are not straightforward. Here, we relate these analyses to theoretical conditions for stabilisation and stable coexistence in classical competition models. By stabilisation, we mean an excess of intraspecific competition relative to interspecific competition that inherently slows or even prevents competitive exclusion. We show that most, though not all, tests demonstrating negative density dependence, negative frequency dependence, and negative plant–soil feedback constitute sufficient conditions for stabilisation of two‐species interactions if applied to data for per capita population growth rates of pairs of species, but none are necessary or sufficient conditions for stable coexistence of two species. Potential inferences are even more limited when communities involve more than two species, and when performance is measured at a single life stage or vital rate. We then discuss two approaches that enable stronger tests for stable coexistence‐invasibility experiments and model parameterisation. The model parameterisation approach can be applied to typical density‐dependence, frequency‐dependence, and plant–soil feedback data sets, and generally enables better links with mechanisms and greater insights, as demonstrated by recent studies.     

CH/π interactions of π-system of acetylacetonato chelate ring: Comparison of CH/π interactions of Ni(II)-acetylacetonato chelate and benzene rings

The calculations have been done for CH/π interaction with π-system of Ni(II)-acetylacetonato chelate ring. The results show that there is an attractive electrostatic interaction, while dispersion component is a major source of attractive interacting energies. The interaction was compared with CH/π interaction between two benzene rings. The comparison shows that two interactions are quite similar, enabling to estimate the energy of CH/π interaction with π-system of Ni(II)-acetylacetonato chelate ring to be about 10.5 kJ/mol. The results indicate that CH/π interactions of chelate ring in various molecular systems can be as important as CH/π interactions of phenyl ring.