Molecular-genetic characterisation of a new nematode resistance gene in wheat

Bread wheat lines introgressed with Aegilops ventricosa chromosomes were evaluated for their resistance to the Australian cereal cyst nematode (CCN, Heterodera avenae) pathotype Ha13. Higher levels of resistance relative to the phenotype of the Cre1 CCN resistance gene in wheat were found in the donor Ae. ventricosa parental lines and chromosome-5N^v substitution or addition lines. The newly identified resistance to pathotype Ha13 on chromosome 5N^v, designated, Cre6, was shown to be independent of the Ae. ventricosa-derived Cre2 gene, effective against several European pathotypes. Another Ae. ventricosa derived gene, Cre5, showed partial resistance to pathotype Ha13. Inhibition of Ha13 female nematode reproduction was ranked in the order Cre6 >Cre1 >CreF≥Cre5. Cre6 was inherited as a single dominant locus. Gene sequences encoding nucleotide-binding sites and leucine-rich repeats (NBS-LRR) from the Cre3 CCN-pathotype Ha13 resistance locus were used as probes to isolate related sequences from one of the donor Ae. ventricosa parents. Related sequences from Ae. ventricosa (71–73% similarity at the amino-acid level to the Cre3-derived sequences) of chromosome 5N^v origin were identified and served as diagnostic molecular markers for the presence of 5N^v. CCN-susceptible plants, found as variants in some of the purported chromosome 5N^v lines, were also found to be missing the diagnostic 5N^v RFLP markers assayed by the NBS-LRR probe. An alloplasmic chromosome-5N^v addition line with Ae. ventricosa cytoplasm in the wheat cultivar, Moisson, background was particularly variable, with 43% CCN-susceptible plants and a corresponding loss of the diagnostic chromosome-5 molecular markers. Received: 26 June 2000 / Accepted: 15 July 2000     

Transplantation of insulin-secreting cells differentiated from human adipose tissue-derived stem cells into type 2 diabetes mice

Currently, there are limited ways to preserve or recover insulin secretory capacity in human pancreas. We evaluated the efficacy of cell therapy using insulin-secreting cells differentiated from human eyelid adipose tissue-derived stem cells (hEAs) into type 2 diabetes mice. After differentiating hEAs into insulin-secreting cells (hEA-ISCs) in vitro, cells were transplanted into a type 2 diabetes mouse model. Serum levels of glucose, insulin and c-peptide were measured, and changes of metabolism and inflammation were assessed in mice that received undifferentiated hEAs (UDC group), differentiated hEA-ISCs (DC group), or sham operation (sham group). Human gene expression and immunohistochemical analysis were done. DC group mice showed improved glucose level, and survival up to 60 days compared to those of UDC and sham group. Significantly increased levels of human insulin and c-peptide were detected in sera of DC mice. RT-PCR and immunohistochemical analysis showed human gene expression and the presence of human cells in kidneys of DC mice. When compared to sham mice, DC mice exhibited lower levels of IL-6, triglyceride and free fatty acids as the control mice. Transplantation of hEA-ISCs lowered blood glucose level in type 2 diabetes mice by increasing circulating insulin level, and ameliorating metabolic parameters including IL-6.     

Proliferating cell nuclear antigen (PCNA) as a proliferative marker during embryonic and adult zebrafish hematopoiesis

We investigated the expression of proliferative cell nuclear antigen (PCNA) in zebrafish to delineate the proliferative hematopoietic component during adult and embryonic hematopoiesis. Immunostaining for PCNA and enhanced green fluorescence protein (eGFP) was performed in wild-type and fli1-eGFP (endothelial marker) and gata1-eGFP (erythroid cell marker) transgenic fish. Expression of PCNA mRNA was examined in wild-type and chordin morphant embryos. In adult zebrafish kidney, the renal tubules are surrounded by endothelial cells and it is separated into hematopoietic and excretory compartments. PCNA was expressed in hematopoietic progenitor cells but not in mature neutrophils, eosinophils or erythroid cells. Some PCNA+ cells are scattered in the hematopoietic compartment of the kidney while others are closely associated with renal tubular cells. PCNA was also expressed in spermatogonial stem cells and intestine crypts, consistent with its role in cell proliferation and DNA synthesis. In embryos, PCNA is expressed in the brain, spinal cord and intermediate cell mass (ICM) at 24 h-post fertilization. In chordin morphants, PCNA is significantly upregulated in the expanded ICM. Therefore, PCNA can be used to mark cell proliferation in zebrafish hematopoietic tissues and to identify a population of progenitor cells whose significance would have to be further investigated.     

Cytochemical investigation of genic male-sterility in Chinese cabbage

A genic male sterile Chinese cabbage, Brassica campestris L. ssp. chinensis Makino, was examined using cytological and cytochemical methods to characterize the process of pollen abortion in this plant. Thick sections of both fertile and sterile anthers at different developmental stages were stained using Toluidine Blue O, Periodic Acid-Schiff’s (PAS) reaction and Sudan Black B to detect cytochemical changes that may occur in the distribution of insoluble polysaccharide and lipid storage bodies. Pollen abortion in sterile anthers occurs at an early stage of microspore development. During early microspore development, reductions in the number of starch grains in the connective tissue of fertile anthers coincide with the accumulation of starch grains in cells of the anther wall. In the late microspore stage, a large vacuole forms in the microspore, and tapetal cells synthesize and accumulate lipid droplets. The cellular organization of tapetal cells in sterile anthers appears similar to that in fertile anthers, except for the absence of lipid droplets in cells of sterile anthers and diffusely labeled tapetal polysaccharides, suggesting defects in nutrient storage. Supported by National Natural Science Foundation of CHINA (30170060)     

Production of biohydrogen by recombinant expression of [NiFe]-hydrogenase 1 in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Background  

Hydrogenases catalyze reversible reaction between hydrogen (H₂) and proton. Inactivation of hydrogenase by exposure to oxygen is a critical limitation in biohydrogen production since strict anaerobic conditions are required. While [FeFe]-hydrogenases are irreversibly inactivated by oxygen, it was known that [NiFe]-hydrogenases are generally more tolerant to oxygen. The physiological function of [NiFe]-hydrogenase 1 is still ambiguous. We herein investigated the H₂ production potential of [NiFe]-hydrogenase 1 of Escherichia coli in vivo and in vitro. The hya A and hya B genes corresponding to the small and large subunits of [NiFe]-hydrogenase 1 core enzyme, respectively, were expressed in BL21, an E. coli strain without H₂ producing ability.     

Engagement of ubiquitination and de-ubiquitination at rostral ventrolateral medulla in experimental brain death

Background

Whereas brain death is a vitally important clinical phenomenon, our contemporary understanding on its underlying cellular mechanisms remains elusive. This study evaluated whether the ubiquitin-proteasome system (UPS) in the rostral ventrolateral medulla (RVLM), a neural substrate that our laboratory identified previously to be intimately related to brain death, is engaged in this fatal process.

Methods

We performed proteomics, Western Blot, real-time PCR, ELISA and pharmacological experiments in conjunction with a clinically relevant experimental endotoxemia model of brain death based on intravenous administration of Escherichia coli lipopolysaccharide in adult male Sprague–Dawley rats.

Results

Proteomics, Western blot and enzyme activity analyses demonstrated that polyubiquitination was preserved and de-ubiquitination by ubiquitin C-terminal hydrolase isozyme-L1 (UCH-L1) was sustained, alongside increased monoubiquitin availability or proteasome activity in RVLM over the course of experimental endotoxemia. However, real-time PCR revealed no significant alteration in proteasome subunit alpha type-1, ubiquitin or UCH-L1 at mRNA level. Functionally, whereas microinjection into the bilateral RVLM of proteasome inhibitors (lactacystin or proteasome inhibitor II) potentiated survival, an inhibitor of ubiquitin-recycling (ubiquitin aldehyde) or an UCH-L1 inhibitor exacerbated mortality.

Conclusions

We proposed previously that the progression towards brain death entails a tug-of-war between pro-death and pro-life programs in RVLM. It is conceivable that ubiquitination or de-ubiquitination in RVLM participate in brain death by regulating the degradation of the proteins involved in those programs.     

Phytoplankton primary productivity seasonality and changes in a small African lake,Lake Hora-Kilole,Ethiopia

The seasonality of primary productivity by phytoplankton in relation to physico-chemical and biological variables was studied in Lake Hora-Kilole from August 2007 to May 2008. In 1989, the Mojo River was temporarily diverted to flow into the lake, which substantially changed its physico-chemical conditions and the composition of the phytoplankton. Primary productivity was controlled primarily by soluble reactive phosphorus (SRP), ammonia (NH₃), temperature and euphotic depth (Z_eu). The light-saturated rate of photosynthesis (A_max) varied from 370 to 3 843?mg O₂ m^?3 h^?1 with the maximum value corresponding to the seasonal maximum of phytoplankton biomass. Compared to the period before the diversion of the river, A_max was reduced by more than ninety-fold in early 1990s and by less than five-fold in 2007 and 2008. Similarly, average phytoplankton chlorophyll a was reduced by more than 2.5 × in the early 1990s and to less than 50% in 2007 and 2008. This highlights the importance of the diversion river water on the physico-chemical and biological environment of the lake.