Taxonomic complexity and breeding system transitions: conservation genetics of the Epipactis leptochila complex (Orchidaceae)

The genus Epipactis contains a problematical complex of autogamous taxa among which species limits are difficult to define. Different authors have treated these plants in different ways, some recognizing the different taxa as distinct species, others considering them as minor intraspecific variants. These contrasting treatments have a direct impact on the conservation resources and status such plants command; 'endemic orchid species' are perceived as having high conservation value, 'localized minor variants' are not. We used allozyme and chloroplast restriction fragment length polymorphism (RFLP) and sequencing analyses to investigate patterns of population genetic structure underlying the taxonomic complexity in this group. Populations of E. dunensis, E. leptochila and E. muelleri were homozygous and uniform for all loci studied here. There were, however, fixed genetic differences among these taxa. Comparisons with published data from the putative progenitor species for the autogamous taxa (the widespread, allogamous E. helleborine) suggest iterative origins of autogamy, rather than the self-pollinating taxa all being merely mutational variants of a single autogamous lineage.     

Infection by a Hematodinium-like parasitic dinoflagellate causes Pink Crab Disease (PCD) in the edible crab Cancer pagurus

The edible crab (Cancer pagurus) supports a large and valuable fishery in UK waters. Much of the catch is transported live to continental Europe in specially designed live-well ('vivier') vehicles. During the winter of 2000/2001, many trap-caught crabs from Guernsey, Channel Islands, UK, were reportedly moribund and pink in colour. These crabs generally died before and during vivier transportation. We provide histological, immunological, and molecular evidence that this condition is associated with infection by a Hematodinium-like dinoflagellate parasite similar to that previously reported in C. pagurus and to an infection causing seasonal mass mortalities of the Norway lobster (Nephrops norvegicus). Pathologically, every altered host bore the infection, which was characterised by very large numbers of plasmodial and vegetative stages in the haemolymph and depletion of reserve cells in the hepatopancreas. Due to the hyperpigmentation of the carapace and appendages, we have called this infection 'Pink Crab Disease' (PCD). Similar Hematodinium infections cause 'Bitter Crab Disease' in tanner and snow crabs, which has had a negative effect on their marketability. At present, little is known about the seasonality, transmission, and market impact of this infection in C. pagurus.     

The ENTH domain

The epsin NH2-terminal homology (ENTH) domain is a membrane interacting module composed by a superhelix of alpha-helices. It is present at the NH2-terminus of proteins that often contain consensus sequences for binding to clathrin coat components and their accessory factors, and therefore function as endocytic adaptors. ENTH domain containing proteins have additional roles in signaling and actin regulation and may have yet other actions in the nucleus. The ENTH domain is structurally similar to the VHS domain. These domains define two families of adaptor proteins which function in membrane traffic and whose interaction with membranes is regulated, in part, by phosphoinositides.     

Mutational bisection of the mitochondrial DNA stability and amino acid biosynthetic functions of ilv5p of budding yeast

Ilv5p is a bifunctional yeast mitochondrial enzyme required for branched chain amino acid biosynthesis and for the stability of mitochondrial DNA (mtDNA) and its parsing into nucleoids. The latter occurs when the general amino acid control (GAC) pathway is activated. We have isolated ilv5 mutants that lack either the enzymatic (a(-)D(+)) or the mtDNA stability function (a(+)D(-)) of the protein. The affected residues in these two mutant classes cluster differently when mapped to the 3-D structure of the spinach ortholog of Ilv5p. a(-)D(+) mutations map to conserved internal domains known to be important for substrate and cofactor binding, whereas the a(+)D(-) mutations map to a C-terminal region on the surface of the protein. The a(+)D(-) mutants also have a temperature-sensitive phenotype when grown on a glycerol medium, which correlates with their degree of mtDNA instability. Analysis of an a(+)D(-) mutant with a strong mtDNA instability phenotype shows that it is also unable to parse mtDNA into nucleoids when activated by the GAC pathway. Finally, the wild-type Escherichia coli ortholog of Ilv5p behaves like a(+)D(-) mutants when expressed and targeted to mitochondria in ilv5Delta yeast cells, suggesting that yeast Ilv5p acquired its mtDNA function after the endosymbiotic event.     

The X-ray crystal structure of human gamma S-crystallin C-terminal domain.

gammaS-crystallin is a major human lens protein found in the outer region of the eye lens, where the refractive index is low. Because crystallins are not renewed they acquire post-translational modifications that may perturb stability and solubility. In common with other members of the betagamma-crystallin superfamily, gammaS-crystallin comprises two similar beta-sheet domains. The crystal structure of the C-terminal domain of human gammaS-crystallin has been solved at 2.4 A resolution. The structure shows that in the in vitro expressed protein, the buried cysteines remain reduced. The backbone conformation of the "tyrosine corner" differs from that of other betagamma-crystallins because of deviation from the consensus sequence. The two C-terminal domains in the asymmetric unit are organized about a slightly distorted 2-fold axis to form a dimer with similar geometry to full-length two-domain family members. Two glutamines found in lattice contacts may be important for short range interactions in the lens. An asparagine known to be deamidated in human cataract is located in a highly ordered structural region.     

Structure of the mammalian CoA transferase from pig heart

Ketoacidosis affects patients who are deficient in the enzyme activity of succinyl-CoA:3-ketoacid CoA transferase (SCOT), since SCOT catalyses the activation of acetoacetate in the metabolism of ketone bodies. Thus far, structure/function analysis of the mammalian enzyme has been predicted based on the three-dimensional structure of a CoA transferase determined from an anaerobic bacterium that utilizes its enzyme for glutamate fermentation. To better interpret clinical data, we have determined the structure of a mammalian CoA transferase from pig heart by X-ray crystallography to 2.5 A resolution. Instrumental to the structure determination were selenomethionine substitution and the use of argon during purification and crystallization. Although pig heart SCOT adopts an alpha/beta protein fold, resembling the overall fold of the bacterial CoA transferase, several loops near the active site of pig heart SCOT follow different paths than the corresponding loops in the bacterial enzyme, accounting for differences in substrate specificities. Two missense mutations found associated with SCOT of ketoacidosis patients were mapped to a location in the structure that might disrupt the stabilization of the amino-terminal strand and thereby interfere with the proper folding of the protein into a functional enzyme.     

IGF-I and IGFBP-3 transport in the rat heart

Specific binding of IGF-binding protein (IGFBP)-3 was shown to be present in the isolated, beating rat heart. The uptake of perfused (125)I-labeled IGF-I in the beating heart was decreased to 9% by blocking IGF-I binding sites with the IGF-I analog Long R(3) (LR(3)) IGF-I. When LR(3) was perfused with complexes of (125)I-IGF-I. IGFBP-3, uptake of (125)I-IGF-I was decreased to 41%, which was significantly greater than LR(3) and (125)I-IGF-I (41 vs. 9%). These data suggest that both microvessel IGF-I and IGFBP-3 binding sites contribute to the transport of IGF-I in the perfused rat heart. This also suggests a novel and plausible mechanism whereby circulating IGFs reach sites of IGF bioactivity.