A new δ chain variant hemoglobin A2-Corfu or α2δ2 116 Arg→Cys (G18), detected by δ-globin gene analysis in a Greek family

Summary Hemoglobin A₂-Corfu or ₂₂116 ArgCys (G18) is a new chain variant detected in a family of Greek descent by direct sequencing of amplified DNA.     

Functional expression of the epithelial Ca(2+) channels (TRPV5 and TRPV6) requires association of the S100A10-annexin 2 complex

TRPV5 and TRPV6 constitute the Ca(2+) influx pathway in a variety of epithelial cells. Here, we identified S100A10 as the first auxiliary protein of these epithelial Ca(2+) channels using yeast two-hybrid and GST pull-down assays. This S100 protein forms a heterotetrameric complex with annexin 2 and associates specifically with the conserved sequence VATTV located in the C-terminal tail of TRPV5 and TRPV6. Of these five amino acids, the first threonine plays a crucial role since the corresponding mutants (TRPV5 T599A and TRPV6 T600A) exhibited a diminished capacity to bind S100A10, were redistributed to a subplasma membrane area and did not display channel activity. Using GST pull-down and co-immunoprecipitation assays we demonstrated that annexin 2 is part of the TRPV5-S100A10 complex. Furthermore, the S100A10-annexin 2 pair colocalizes with the Ca(2+) channels in TRPV5-expressing renal tubules and TRPV6-expressing duodenal cells. Importantly, downregulation of annexin 2 using annexin 2-specific small interfering RNA inhibited TRPV5 and TRPV6-mediated currents in transfected HEK293 cells. In conclusion, the S100A10-annexin 2 complex plays a crucial role in routing of TRPV5 and TRPV6 to plasma membrane.     

The association between male infertility and sperm disomy: Evidence for variation in disomy levels among individuals and a correlation between particular semen parameters and disomy of specific chromosome pairs

Background  

The association between infertility and sperm disomy is well documented. Results vary but most report that men with severely compromised semen parameters have a significantly elevated proportion of disomic sperm. The relationship between individual semen parameters and segregation of specific chromosome pairs is however less well reported as is the variation of disomy levels in individual men.     

Relationships between nitrogen,dry matter accumulation and glucosinolates in <Emphasis Type="Italic">Eruca sativa</Emphasis> Mills. The applicability of the critical NO<Subscript>3</Subscript>-N levels approach

Background and Aims

Rocket salad (Eruca sativa Mills) is one of the major leafy vegetables produced worldwide and has been characterized as a rich source of chemoprotective glucosinolates (GSL). The relationship between N fertilization and the resulting plant biomass and N status with GSL quantity and quality in rocket leaves was examined.

Methods

A pot experiment was conducted, applying ten different N-rates and destructive sampling was carried out 15, 30 and 45 days after transplanting (DAT). The Mitscherlich equation was used to establish NO₃-N critical levels at each growth stage and as an indicator of N demand for relative maximum dry matter accumulation and glucosinolate content and composition was determined.

Results

Glucosinolate content was significantly influenced by N rate, growth stage and their interaction. Different GSL types showed dissimilar responses to N fertilization: aliphatic GSLs were significantly reduced under increased N rates whereas indole GSL showed the reverse. Under excess N fertilization (>1.04 g/plant), dry matter accumulation remained constant, NO₃-N was significantly increased and total GSL content was significantly reduced, factors that could lead to an anticipated product quality decline.

Conclusions

The application of the critical NO₃-N level approach used to identify optimal N fertilization rates for plant growth could serve as means to obtain optimized GSL content in the edible plant parts.

     

Role of the hydrophobic and charged residues in the 218–226 region of apoA-I in the biogenesis of HDL

We investigated the significance of hydrophobic and charged residues 218–226 on the structure and functions of apoA-I and their contribution to the biogenesis of HDL. Adenovirus-mediated gene transfer of apoA-I[L218A/L219A/V221A/L222A] in apoA-I^−/− mice decreased plasma cholesterol and apoA-I levels to 15% of wild-type (WT) control mice and generated pre-β- and α4-HDL particles. In apoA-I^−/− × apoE^−/− mice, the same mutant formed few discoidal and pre-β-HDL particles that could not be converted to mature α-HDL particles by excess LCAT. Expression of the apoA-I[E223A/K226A] mutant in apoA-I^−/− mice caused lesser but discrete alterations in the HDL phenotype. The apoA-I[218–222] and apoA-I[E223A/K226A] mutants had 20% and normal capacity, respectively, to promote ABCA1-mediated cholesterol efflux. Both mutants had ∼65% of normal capacity to activate LCAT in vitro. Biophysical analyses suggested that both mutants affected in a distinct manner the structural integrity and plasticity of apoA-I that is necessary for normal functions. We conclude that the alteration of the hydrophobic 218–222 residues of apoA-I disrupts apoA-I/ABCA1 interactions and promotes the generation of defective pre-β particles that fail to mature into α-HDL subpopulations, thus resulting in low plasma apoA-I and HDL. Alterations of the charged 223, 226 residues caused milder but discrete changes in HDL phenotype.     

Differential Regulation of Phenylethanolamine N-Methyltransferase Expression in Two Distinct Subpopulations of Bovine Chromaffin Cells

Abstract: Chromaffin cells were isolated from bovine adrenal glands and fractionated into two distinct subpopulations by density gradient centrifugation on Percoll. Cells in the more dense fraction stored epinephrine (E) as their predominant catecholamine (81% of total catecholamines), contained high levels of phenylethanolamine N-methyltransferase (PNMT) activity, and exhibited intense PNMT immunoreactivity. This population of chromaffin cells was termed the E-rich cell population. Cells in the less dense fraction, the norepinephrine (NE)-rich cell population, stored predominantly NE (75% of total catecholamines). Although the NE-rich cells had only 3% as much PNMT activity as did the E-rich cells, 20% of the NE-rich cells were PNMT immunoreactive. This suggested that the PNMT-positive cells in the NE-rich cell cultures contained less PNMT per cell than did E-rich cells and may not be typical adrenergic cells. The regulation of PNMT mRNA levels and PNMT activity in primary cultures of E-rich and NE-rich cells was compared. At the time the cells were isolated, PNMT mRNA levels in NE-rich cells were ～20% of those in E-rich cells; within 48 h in culture, PNMT mRNA in both populations declined to almost undetectable levels. Treatment with dexamethasone increased PNMT mRNA levels and PNMT activity in both populations. In E-rich cells, dexamethasone restored PNMT mRNA to the level seen in freshly isolated cells and increased PNMT activity twofold. In NE-rich cells, dexamethasone increased PNMT mRNA to levels twice those found in freshly isolated cells and increased PNMT activity sixfold. Cycloheximide blocked the effects of dexamethasone on PNMT mRNA expression in NE-rich cells but had little effect in E-rich cells. Angiotensin II, forskolin, and phorbol 12,13-dibutyrate elicited large increases in PNMT mRNA levels in E-rich cells but had no effect in NE-rich cells. Our data suggest that PNMT expression is regulated differently in the two chromaffin cell subpopulations.