Absence ofGABRA1 Ala322Asp mutation in juvenile myoclonic epilepsy families from India

An Ala322Asp mutation in theGABRA1 gene was recently reported to be responsible for causing the autosomal dominant (AD) form of juvenile myoclonic epilepsy (JME) in a French-Canadian family. To study if JME families from India exhibiting the AD mode of inheritance carry the Ala322Asp mutation, we examined 35 unrelated JME-affected individuals from such families for the Ala322Asp mutation inGABRA1. Ala322Asp mutation was not observed in any of these JME-affected individuals, suggesting that this mutation is unlikely to be a predominant mutation involved in causation of epilepsy. To evaluate the possibility of other mutation(s) in and aroundGABRA1 that may predispose to JME, we compared the allele frequencies at two marker loci, D5S2118 and D5S422, flankingGABRA1, in probands and 100 matched population controls. One of the allele frequencies at D5S422 shows a significant difference between the cases and controls (χ² = 11.44, d.f. = 1,P = 0.0007), suggesting genetic association between JME and genes located in the proximity of the DNA marker.     

Detection of gene duplications and block duplications in eukaryotic genomes

Several eukaryotic genomes have been completely sequenced and this provides an opportunity to investigate the extent and characteristics (e.g., single gene duplication, block duplication, etc.) of gene duplication in a genome. Detecting duplicate genes in a genome, however, is not a simple problem because of several complications such as domain shuffling, the existence of isoforms derived from alternative splicing, and annotational errors in the databases. We describe a method for overcoming these difficulties and the extents of gene duplication in the genomes of Drosophila melanogaster, Caenorhabditis elegans, and yeast inferred from this method. We also describe a method for detecting block duplications in a genome. Application of this method showed that block duplication is a common phenomenon in both yeast and nematode. The patterns of block duplication in the two species are, however, markedly different. Yeast shows much more extensive block duplication than nematode, with some chromosomes having more than 40% of the duplications derived from block duplications. Moreover, in yeast the majority of block duplications occurred between chromosomes, while in nematode most block duplications occurred within chromosomes.     

Non‐angiosperm phytochromes and the evolution of vascular plants

The phytochromes, a class of plant light‐sensing pigments, are a gene family with a long, complex evolutionary history. Angiosperms each have five or more phytochromes (designated A to E in Arabidopsis ) with distinct functions as light receptors and only moderate sequence identities for different types within a species. The long‐term challenge taken up here is to trace the origin and function of the various motifs within the angiosperm phytochromes through gymnosperm phytochromes (types N, O and P) and lower plant phytochromes, sometimes reaching even to bacterial progenitor molecules. Particularly intriguing are the findings of homology of a C‐terminal region of phytochromes with bacterial transmitter modules and of a large N‐terminal region with a protein encoded by a gene from the cyanobacterum Synechocystis . Phylogenetic analysis helps to answer general questions such as the times of divergence of mono‐ and dicotyledons, of groups of gymnosperms or of ferns. Phytochrome sequences suggest (1) that mono‐ and dicotyledons became separated 150‐200 million years earlier than indicated by the fossil record and (2) that Ginkgo and Cycas have been separated unexpectedly late from the lineage giving rise to the Pinidae. (3) The status of Psilotum as a close relative of the primeval vascular plants is not supported. Phytochrome gene sequences additionally reveal that (4) moss and fern phytochromes have erratically acquired C‐termini which, though kinase‐like, are different from the common ones and that (5) introns have been lost, gained or shifted in position from algae to angiosperms. Phytochromes promise to be a rich source of phylogenetic information into the future as more sequences and functional data emerge, not least from studies of lower plants.     

Response to three seasons of elevated ozone in the progeny of healthy and unhealthy Norway spruce trees from a plantation with the 'top dying' syndrome

We tested a hypothesis that elevated ozone was an eliciting or contributing factor in outbreaks of the 'top dying' (or 'subtop dying') syndrome in Norway spruce ( Picea abies ). Progeny were used from open-pollinated trees within a stand with the 'top dying' syndrome. The mother trees were classified in relation to the expression of the 'top dying' syndrome, and progeny from the healthiest and least healthy thirds of the population were exposed to high and low concentrations of ozone for three seasons. Elevated ozone did not affect height growth of the trees. It did not measurably affect net photosynthesis, stomatal conductance or instantaneous water use efficiency. Chlorophyll and carotenoid contents were also not significantly affected by ozone concentration. In the first year, instantaneous water use efficiency was lower in the progeny of the unhealthy mother trees than in the healthy mother trees. Furthermore, the unhealthy mother trees tended to produce longer annual shoots and showed more winter damage at the end of the experiment. None of these parameters were related to ozone concentration in the atmosphere. These results do not support a hypothesis that elevated ozone is a significant contributory factor or an eliciting factor in the development of the syndrome.     

Evolution of murine α1-proteinase inhibitors: Gene amplification and reactive center divergence

The organization and sequence of genes encoding the ₁-proteinase inhibitor (₁PI), a major serine proteinase inhibitor of the mammalian bloodstream, have been compared in several species, including murine rodents (genus Mus). Analysis of gene copy number indicates that amplification of ₁PI genes occurred at some time during evolution of the Mus genus, leading to fixation of a family of about three to five genes in several existing species (e.g., M. domesticus and M. saxicola), and only a single gene in others (e.g., M. caroli). A phylogeny for the various mammalian ₁PI mRNAs was constructed based upon synonymous substitutions within coding regions. The mRNAs in different murine species diverged from a common ancestor before the formation of the first species lineages of the Mus genus, i.e., about 10–13 million years ago. Thus, ₁PI gene amplification must have occurred prior to Mus speciation; gene families were retained in some, but not all, murine species. The reactive center region of the ₁PI polypeptide, which determines target protease specificity, has diverged rapidly during evolution of the Mus species, but not during evolution of other mammalian species included in the analysis. It is likely that this accelerated evolution of the reactive center, which has been noted previously for serine proteinase inhibitors, was driven by some sort of a positive Darwinian selection that was exerted in a taxon-specific manner. We suggest that evolution of ₁PI genes of murine rodents has been characterized by both modification of gene copy number and rapid reactive center divergence. These processes may have resulted in a broadened repertoire of proteinase inhibitors that was evolutionarily advantageous during Mus speciation.Correspondence to: F.G. Berger     

Comparative amino acid sequence analysis of Thermotoga maritima β-glucosidase (BglA) deduced from the nucleotide sequence of the gene indicates distant relationship between β-glucosidases of the BGA family and other families of β-1,4-glycosyl hydrolases

The primary structure of the bglA gene region encoding a -glucosidase of Thermotoga maritima strain MSB8 was determined. The bglA gene has the potential to code for a polypeptide of 446 amino acids with a predicted molecular mass of 51545 Da. The T, maritima -glucosidase (BglA) was overexpressed in E. coli at a level comprising approximately 15–20% of soluble cellular protein. Based on its amino acid sequence, as deduced from the nucleotide sequence of the gene, BglA can be classified as a broad-specificity -glucosidase and as a member of the -glucosidase family BGA, in agreement with the results of enzymatic characterization of the recombinant protein. Comparative sequence analysis revealed distant amino acid sequence similarities between BGA family -glucosidases, a -xylosidase, -1,4-glycanases of the enzyme family F (mostly xylanases), and other families of -1,4-glycosyl hydrolases. This result indicates that BGA -glucosidases may comprise one enzyme family within a large enzyme order of retaining -glycosyl hydrolases, and that the members of these enzyme groups may be inter-related at the level of active site architecture and perhaps even on the level of overall three-dimensional fold.     

Evolutionary genetics ofDrosophila esterases

Over 30 carboxylester hydrolases have been identified inD. melanogaster. Most are classified as acetyl, carboxyl or cholinesterases. Sequence similarities among most of the carboxyl and all the cholinesterases so far characterised fromD. melanogaster and other eukaryotes justify recognition of a carboxyl/cholinesterase multigene family. This family shows minimal sequence similarities with other esterases but crystallographic data for a few non-drosophilid enzymes show that the family shares a distinctive overall structure with some other carboxyl and aryl esterases, so they are all put in one superfamily of / hydrolases. Fifteen esterase genes have been mapped inD. melanogaster and twelve are clustered at two chromosomal sites. The constitution of each cluster varies acrossDrosophila species but two carboxyl esterases in one cluster are sufficiently conserved that their homologues can be identified among enzymes conferring insecticide resistance in other Diptera. Sequence differences between two other esterases, the EST6 carboxyl esterase and acetylcholinesterase, have been interpreted against the consensus super-secondary structure for the carboxyl/cholinesterase multigene family; their sequence differences are widely dispersed across the structure and include substantial divergence in substrate binding sites and the active site gorge. This also applies when EST6 is compared across species where differences in its expression indicate a difference in function. However, comparisons within and among species where EST6 expression is conserved show that many aspects of the predicted super-secondary structure are tightly conserved. Two notable exceptions are a pair of polymorphisms in the substrate binding site of the enzyme inD. melanogaster. These polymorphisms are associated with differences in substrate interactions_vitro} and demographic data indicate that the alternative forms are not selectively equivalentin vivo.     

Evolution of EF-hand calcium-modulated proteins. V. The genes encoding EF-hand proteins are not clustered in mammalian genomes

Summary The chromosomal assignments of genes belonging to the EF-hand family which have a common origin are compiled in this article. So far data are available from 27 human gene loci belonging to 6 subfamilies and 8 murine loci belonging to 4 subfamilies. Chromosomal localization has been obtained by somatic-cell hybrid analysis using the Southern blot technique or PCR amplification, metaphase spread in situ hybridization, or isolation of the particular genes from chromosome-specific libraries. Except for genes of the S-100 alpha proteins which are grouped on human chromosome 1q12-25 and mouse chromosome 3, no linkage has been found for genes encoding EF-hand proteins, indicating absence of selective pressure for maintaining chromosomal clustering. Six of these genes map to known syntenic groups conserved in the human and mouse genomes. This suggests that chromosomal translocations occurred before divergence of these species. The possible significance of chromosomal positioning with respect to nearby located known genes and genetic disease loci is discussed.     

Biological meaning,statistical significance,and classification of local spatial similarities in nonhomologous proteins

We have completed an exhaustive search for the common spatial arrangements of backbone fragments (SARFs) in nonhomologous proteins. This type of local structural similarity, incorporating short fragments of backbone atoms, arranged not necessarily in the same order along the polypeptide chain, appears to be important for protein function and stability. To estimate the statistical significance of the similarities, we have introduced a similarity score. We present several locally similar structures, with a large similarity score, which have not yet been reported. On the basis of the results of pairwise comparison, we have performed hierarchical cluster analysis of protein structures. Our analysis is not limited by comparison of single chains but also includes complex molecules consisting of several subunits. The SARFs with backbone fragments from different polypeptide chains provide a stable interaction between subunits in protein molecules. In many cases the active site of enzymes is located at the same position relative to the common SARFs, implying a function of the certain SARFs as a universal interface of the protein-substrate interaction.