Partial mispairing and crossing-over between β0 and δ genes as the origin of the δ β0 thalassemia gene

Summary Two double heterozygous ⁰/⁰ thalassemic sibs of Mexican descent were studied. The father had a ⁰/⁰ genotype, while the mother, one sib and several maternal relatives were ⁰/⁰ heterozygotes. Parental consanguinity and an apparently low frequency of thalassemia among Mexicans suggested a possible common origin of both ⁰ and ⁰ genes. A hypothesis to explain such a possibility is proposed on the basis of a partial mispairing between ⁰ and genes followed by a crossing-over which would results in a ⁰ recombinant gene. This hypothesis could also be extended to explain the 22 gluala, 22 alaglu and 116 arghis Hb variants as recombinants from double crossing-over between and mispaired genes for which the name interstitial-Lepore is proposed.     

Total Contents of 《Natural Product Research and Development》in 2 0 0 1

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Properties of sodium and potassium channels of the squid giant axon far below 0°C

Summary Squid giant axon could be excited in concentrated glycerol solutions containing normal concentrations of electrolytes, when osmolalities of solutions inside and outside the axon were matched. These glycerol solutions did not freeze at the temperature as low as –19°C. The nerve excitation in these solutions were observed at this low temperature. The excitation process at this low temperature was slowed down and time constants of the excitation kinetics were several hundredfold larger than those in normal seawater at 10°C, under which temperature the squid habituated. The temperature coefficients for the electrophysiological membrane parameters under this condition were larger than those in normal seawater above 0°C. The Q₁₀ value for the conduction velocity was 2.0 and that of the duration of the action potential was around 8.5. The time course of the membrane currents was also slowed with the Q₁₀ value of around 5 and the magnitude decreased with the Q₁₀ value of around 2 as the temperature was lowered. The Q₁₀ values for the kinetics of the on process of the Na-channel were around 4.5 and were almost the same as those of the off process of the Na-channel in the wide range of the temperature below 0°C. The Q₁₀ value of the on process of K-channel was around 6.5 and was larger than those for Na-channel. The Q₁₀ values increased gradually as the temperature was lowered.     

Sperm ultrastructure in the diatoms Melosira and Thalassiosira and the significance of the 9 + 0 configuration

The most complete account to date of the ultrastructure of flagellate cells in diatoms is given for the sperm of Thalassiosira lacustris and Melosira moniliformis var. octogona, based on serial sections. The sperm are uniflagellate, with no trace of a second basal body, and possess a 9?+?0 axoneme. The significance of the 9?+?0 configuration is discussed: lack of the central pair microtubules and radial spokes does not compromise the mastigoneme-bearing flagellum’s capacity to perform planar beats and thrust reversal and may perhaps be related to sensory/secretory function of the sperm flagellum during plasmogamy. The basal bodies of diatoms are confirmed to contain doublets rather than triplets, which may correlate with the absence of some centriolar proteins found in most cells producing active flagella. Whereas Melosira possesses a normal cartwheel structure in the long basal body, no such structure is present in Thalassiosira, which instead possesses ‘intercalary fibres’ linking the basal body doublets. No transitional helices or transitional plates are present in either species studied. Cones of microtubules are associated with the basal body and partially enclose the nucleus in M. moniliformis and T. lacustris. They do not appear to be true microtubular roots and may arise through transformation of the meiosis II spindle. A close association between cone microtubules and tubules containing mastigonemes may indicate a function in intracellular mastigoneme transport. No correlation can yet be detected between methods of spermatogenesis and phylogeny in diatoms, contrary to previous suggestions.     

Positions of the cytoplasmic end of BK α S0 helix relative to S1–S6 and of β1 TM1 and TM2 relative to S0–S6

The large-conductance, voltage- and Ca²⁺-gated K⁺ (BK) channel consists of four α subunits, which form a voltage- and Ca²⁺-gated channel, and up to four modulatory β subunits. The β1 subunit is expressed in smooth muscle, where it slows BK channel kinetics and shifts the conductance–voltage (G-V) curve to the left at [Ca²⁺] > 2 µM. In addition to the six transmembrane (TM) helices, S1–S6, conserved in all voltage-dependent K⁺ channels, BK α has a unique seventh TM helix, S0, which may contribute to the unusual rightward shift in the G-V curve of BK α in the absence of β1 and to a leftward shift in its presence. Such a role is supported by the close proximity of S0 to S3 and S4 in the voltage-sensing domain. Furthermore, on the extracellular side of the membrane, one of the two TM helices of β1, TM2, is adjacent to S0. We have now analyzed induced disulfide bond formation between substituted Cys residues on the cytoplasmic side of the membrane. There, in contrast, S0 is closest to the S2–S3 loop, from which position it is displaced on the addition of β1. The cytoplasmic ends of β1 TM1 and TM2 are adjacent and are located between the S2–S3 loop of one α subunit and S1 of a neighboring α subunit and are not adjacent to S0; i.e., S0 and TM2 have different trajectories through the membrane. In the absence of β1, 70% of disulfide bonding of W43C (S0) and L175C (S2–S3) has no effect on V₅₀ for activation, implying that the cytoplasmic end of S0 and the S2–S3 loop move in concert, if at all, during activation. Otherwise, linking them together in one state would obstruct the transition to the other state, which would certainly change V₅₀.     

Thermal Stress Induced Aggregation of Aquaporin 0 (AQP0) and Protection by α-Crystallin via Its Chaperone Function

Aquaporin 0 (AQP0) formerly known as membrane intrinsic protein (MIP), is expressed exclusively in the lens during terminal differentiation of fiber cells. AQP0 plays an important role not only in the regulation of water content but also in cell-to-cell adhesion of the lens fiber cells. We have investigated the thermal stress-induced structural alterations of detergent (octyl glucoside)-solubilized calf lens AQP0. The results show an increase in the amount of AQP0 that aggregated as the temperature increased from 40°C to 65°C. α-Crystallin, molecular chaperone abundantly present in the eye lens, completely prevented the AQP0 aggregation at a 1∶1 (weight/weight) ratio. Since α-crystallin consists of two gene products namely αA- and αB-crystallins, we have tested the recombinant proteins on their ability to prevent thermal-stress induced AQP0 aggregation. In contrast to the general observation made with other target proteins, αA-crystallin exhibited better chaperone-like activity towards AQP0 compared to αB-crystallin. Neither post-translational modifications (glycation) nor C-terminus truncation of AQP0 have any appreciable effect on its thermal aggregation properties. α-Crystallin offers similar protection against thermal aggregation as in the case of the unmodified AQP0, suggesting that αcrystallin may bind to either intracellular loops or other residues of AQP0 that become exposed during thermal stress. Far-UV circular dichroism studies indicated a loss of αhelical structures when AQP0 was subjected to temperatures above 45°C, and the presence of α-crystallin stabilized these secondary structures. We report here, for the first time, that α-crystallin protects AQP0 from thermal aggregation. Since stress-induced structural perturbations of AQP0 may affect the integrity of the lens, presence of the molecular chaperone, α-crystallin (particularly αA-crystallin) in close proximity to the lens membrane is physiologically relevant.     

Composition and biomass of summer metazoan plankton in the 0–200 m layer of the Atlantic sector of the Antarctic

Composition of the metazoan plankton was studied during R.V. Dmitry Mendeleev cruise 43 (February to April, 1989) in the Atlantic sector of the Southern Ocean. Samples were collected from ten stations at six locations. Four of the locations were in open oceanic waters along the 15° W longitude. Two others were in the Bransfield Strait and in inshore waters near Elephant Island. At three locations at 15° W sampling was conducted twice or thrice. At all stations three different sampling gears were used to collect different size groups of Zooplankton: series of hauls were performed by 2001 water-bottle, mesoplankton net and macroplankton trawl for depths from 200 m to the surface. The average biomass of Zooplankton in open oceanic waters was 20.55 g · m^–2 wet weight. Copepoda Calanoida dominated composing 54.8% of the total plankton, followed by Euphausiacea (19.8%), Ctenophora (9.7%) and Copepoda Cyclopoida (7.2%). Biomass of any other taxonomic group was less than 1g·m^–2. The relative biomass of Calanoida had a tendency to decrease southward along 15°W from 86.1 to 68.1% in February and from 81.8 to 23.6% in March–April. The relative biomass of Euphausiacea increased in the same manner from 2.3 to 17.8% in February and from 3.7 to 41.6% in March–April. The average biomass of calanoids from February to March–April decreased from 77.3 to 31.2% and that of euphausiids increased from 6.2 to 33.8%. The contribution of copepods and euphausiids to the production of the plankton community in the Antarctic is discussed.     

Rates of chilling to 0°C: implications for the survival of microorganisms and relationship with membrane fluidity modifications

The effects of slow chilling (2°C min⁻¹) and rapid chilling (2,000°C min⁻¹) were investigated on the survival and membrane fluidity of Escherichia coli, of Bacillus subtilis, and of Saccharomyces cerevisiae. Cell death was found to be dependent on the physiological state of cell cultures and on the rate of temperature downshift. Slow temperature decrease allowed cell stabilization, whereas the rapid chilling induced an immediate loss of viability of up to more than 90 and 70% for the exponentially growing cells of E. coli and B. subtilis, respectively. To relate the results of viability with changes in membrane physical state, membrane anisotropy variation was monitored during thermal stress using the fluorescence probe 1,6-diphenyl-1,3,5-hexatriene. No variation in the membrane fluidity of all the three microorganisms was found after the slow chilling. It is interesting to note that fluorescence measurements showed an irreversible rigidification of the membrane of exponentially growing cells of E. coli and B. subtilis after the instantaneous cold shock, which was not observed with S. cerevisiae. This irreversible effect of the rapid cold shock on the membrane correlated well with high rates of cell inactivation. Thus, membrane alteration seems to be the principal cause of the cold shock injury.     

Phase studies of model biomembranes: Macroscopic coexistence of Lα + Lβ, with light-induced coexistence of Lα + Lo Phases

Phase diagrams of 3-component lipid bilayer mixtures containing cholesterol reveal major differences among the different types of lipids. Here we report that mixtures of cholesterol together with POPC and a high-melting temperature PC or sphingomyelin show different phase behavior from similar mixtures that contain DOPC or di-phytanoyl-PC instead of POPC. In particular, only one region of macroscopic phase coexistence occurs with POPC, a region of coexisting liquid disordered and solid phases, {Lα + Lβ}. Fluorescence microscopy imaging is useful for these studies, but is subject to artifactual light-induced domain formation, as reported by Ayuyan and Cohen [A.G. Ayuyan, F.S. Cohen, Lipid peroxides promote large rafts: Effects of excitation of probes in fluorescence microscopy and electrochemical reactions during vesicle formation, Biophys. J. 91 (2006) 2172-2183.]. This artifact can be attenuated by decreased illumination and low dye concentration. The use of the free radical scavenger n-propyl gallate can reduce the artifact, but this molecule enters the bilayer and itself perturbs the phase behavior. We suggest that the light-induced domain separation artifact might actually arise from pre-existing lipid clusters that are induced to coalesce, and therefore indicates highly nonrandom mixing of the lipid components.     

Simultaneous net accumulation of both K+ and Na+ by lymphocytes at 0°C

Human lymphocytes at 0°C in low Na⁺ medium accumulate both K⁺ and Na⁺ to levels higher than in the external medium. This is not due to an impermeable compartment or a Donnan equilibrium, and is incompatible with the membrane Na⁺-pump concept. In contrast, it supports prior evidence that ion exchange in lymphocytes is mediated by adsorption onto and desorption from fixed anionic sites within the cell. Additional aspects of ion and water contents of cells in low Na⁺ medium are described and are explained by this concept.     

Square-planar copper(II) halide complexes of tridentate ligands with π-π stacking interactions and alternating short and long Cu ? Cu distances

The preparation of a new tridentate N₂O-donor ligand N-(2-pyridylmethyl)-3-methoxysalicylaldiminato (HL) is described, together with the corresponding copper(II) complexes [Cu(L)X] (X = Cl⁻, Br⁻). The compounds were characterized by elemental analysis, spectral, magnetic and crystallographic studies. In both compounds, the local molecular structure of the Cu(II) ion involves a square-planar CuN₂OX chromophore, consisting of a deprotonated phenolate oxygen, an imine nitrogen, the pyridine nitrogen and X. In the solid state, π-π stacking interactions are dominantly present, involving the pyridine and phenolate rings of neighboring molecules, which lead to a one-dimensional arrangement with alternating short and long Cu ? Cu distances of [3.720, 4.599 Å] for the bromo complex and of [3.698, 4.696 Å] for the chloro complex. The temperature-dependent magnetic measurements and EPR data of polycrystalline samples, as well as of frozen solutions in CHCl₃ show that there is no observable exchange interaction between the Cu ions. The EPR parameters (g_∥, A_∥) agree with a perfect planar geometry, just as found in the X-ray analysis.     

Carbonyl complexes of Rh(I), Ir(I) and Mo(0) containing substituted derivatives of 1,10-phenanthroline and 2,2′-bipyridine

The A₁ symmetry v_CO of the carbonyl complexes [Mo(chel)(CO)₄], [M(chel)(CO)₂] [PF₆] (M=Rh, Ir; chel=bipy, phen and substituted derivatives) are used for determining the electron donor-acceptor properties of the title ligands. The steric hindrance of the methyl groups in positions 2 and 9 of the phenanthroline favours the formation of Rh(I) and Ir(I) pentacoordinated derivatives.     

Molecular Evolution and Nucleotide Sequences of the Maize Plastid Genes for the α Subunit of Cf1 (atpA) and the Proteolipid Subunit of Cf0 (atpH)

The nucleotide sequences of the maize plastid genes for the alpha subunit of CF1 (atpA) and the proteolipid subunit of CF0 (atpH) are presented. The evolution of these genes among higher plants is characterized by a transition mutation bias of about 2:1 and by rates of synonymous and nonsynonymous substitution which are much lower than similar rates for genes from other sources. This is consistent with the notion that the plastid genome is evolving conservatively in primary sequence. Yet, the mode and tempo of sequence evolution of these and other plastid-encoded coupling factor genes are not the same. In particular, higher rates of nonsynonymous substitution in atpE (the gene for the epsilon subunit of CF1) and higher rates of synonymous substitution in atpH in the dicot vs. monocot lineages of higher plants indicate that these sequences are likely subject to different evolutionary constraints in these two lineages. The 5'- and 3'-transcribed flanking regions of atpA and atpH from maize, wheat and tobacco are conserved in size, but contain few putative regulatory elements which are conserved either in their spatial arrangement or sequence complexity. However, these regions likely contain variable numbers of "species-specific" regulatory elements. The present studies thus suggest that the plastid genome is not a passive participant in an evolutionary process governed by a more rapidly changing, readily adaptive, nuclear compartment, but that novel strategies for the coordinate expression of genes in the plastid genome may arise through rapid evolution of the flanking sequences of these genes.     

Key aromatic residues at subsites + 2 and + 3 of glycoside hydrolase family 31 α-glucosidase contribute to recognition of long-chain substrates

Glycoside hydrolase family 31 α-glucosidases (31AGs) show various specificities for maltooligosaccharides according to chain length. Aspergillus niger α-glucosidase (ANG) is specific for short-chain substrates with the highest k_cat/K_m for maltotriose, while sugar beet α-glucosidase (SBG) prefers long-chain substrates and soluble starch. Multiple sequence alignment of 31AGs indicated a high degree of diversity at the long loop (N-loop), which forms one wall of the active pocket. Mutations of Phe236 in the N-loop of SBG (F236A/S) decreased k_cat/K_m values for substrates longer than maltose. Providing a phenylalanine residue at a similar position in ANG (T228F) altered the k_cat/K_m values for maltooligosaccharides compared with wild-type ANG, i.e., the mutant enzyme showed the highest k_cat/K_m value for maltotetraose. Subsite affinity analysis indicated that modification of subsite affinities at + 2 and + 3 caused alterations of substrate specificity in the mutant enzymes. These results indicated that the aromatic residue in the N-loop contributes to determining the chain-length specificity of 31AGs.     

Synthesis and Cytotoxicity of О- and N-Acyl Derivatives of Azepanobetulin

Russian Journal of Bioorganic Chemistry - A five-stage synthesis of azepanobetulin from betulin with a total yield of 47% has been carried out. The acylation of azepanobetulin with anhydrides or...     

Cardiovascular causes of sudden unexpected death in children and adolescents (0–17 years)

Background

Little is known about the causes of unexpected death in minors (0–17 years). In young adults an important cause is cardiovascular disease, with primary arrhythmogenic disorders, atherosclerotic events, cardiomyopathies and myocarditis as main contributors. The aim of this autopsy study was to determine the contribution of cardiovascular disease to unexpected death in minors.

Methods and results

In the Netherlands, systematic investigation of all cases of unexplained death in minors was compulsory in a nationwide governmental project during a 15-month period. Autopsies were performed according to a standardised protocol (autopsy rate 85%). A cardiovascular cause of death was found in 13/56 cases (23%). In the group <1 year, the main cardiovascular causes were various congenital defects (n?=?3) and myocarditis (n?=?2). In the 1–9 year group, no cardiovascular causes were found. In the 10–14 year group, hypertrophic cardiomyopathy (n?=?1) and ruptured ascending aortic aneurysm (n?=?1) were among the observed cardiovascular causes. In 14/56 (25%) cases autopsy revealed no structural abnormalities that could explain the sudden death, mostly in the group <1 year.

Conclusion

This national cohort with a high autopsy rate reveals a high incidence (23%) of cardiovascular diseases as the pathological substrate of sudden unexpected death in children. Another high percentage of minors (25%) showed no structural abnormalities, with the possibility of a genetic arrhythmia. These findings underline the importance of systematic autopsy in sudden death in minors, with implications for cardiogenetic screening of relatives.

     

Rectification of the water permeability in COS-7 cells at 22, 10 and 0°C

The osmotic and permeability parameters of a cell membrane are essential physico-chemical properties of a cell and particularly important with respect to cell volume changes and the regulation thereof. Here, we report the hydraulic conductivity, L(p), the non-osmotic volume, V(b), and the Arrhenius activation energy, E(a), of mammalian COS-7 cells. The ratio of V(b) to the isotonic cell volume, V(c iso), was 0.29. E(a), the activation energy required for the permeation of water through the cell membrane, was 10,700, and 12,000 cal/mol under hyper- and hypotonic conditions, respectively. Average values for L(p) were calculated from swell/shrink curves by using an integrated equation for L(p). The curves represented the volume changes of 358 individually measured cells, placed into solutions of nonpermeating solutes of 157 or 602 mOsm/kg (at 0, 10 or 22°C) and imaged over time. L(p) estimates for all six combinations of osmolality and temperature were calculated, resulting in values of 0.11, 0.21, and 0.10 μm/min/atm for exosmotic flow and 0.79, 1.73 and 1.87 μm/min/atm for endosmotic flow (at 0, 10 and 22°C, respectively). The unexpected finding of several fold higher L(p) values for endosmotic flow indicates highly asymmetric membrane permeability for water in COS-7. This phenomenon is known as rectification and has mainly been reported for plant cell, but only rarely for animal cells. Although the mechanism underlying the strong rectification found in COS-7 cells is yet unknown, it is a phenomenon of biological interest and has important practical consequences, for instance, in the development of optimal cryopreservation.