Data sets,partitions, and characters: philosophies and procedures for analyzing multiple data sets

We compared four approaches for analyzing three data sets derived from staphylinoid beetles, a superfamily whose known species diversity is roughly comparable to that of vertebrates. One data set is derived from adult morphology and the two molecular data sets are from 12S ribosomal RNA and cytochrome b mitochondrial DNA. We found that taxonomic congruence following conditional data combination, herein called compatible evidence (CE), resolved more nodes compatible with an initial conservative hypothesis than did total evidence (TE), conditional data combination (CDC), or taxonomic congruence (TC). CE sets a base of nodes obtained by CDC analysis and then investigates what further agreement may arise in a universe where these nodes are accepted as given. We suggest that CE75-75 may be appropriate for future studies that aim to both generate a well-corroborated tree and investigate conflicts between data sets, partitions, and characters. CE75-75 is a 75% bootstrap consensus CDC tree followed by combinable-component consensus of a 75% bootstrap consensus of each homogeneous set of partitions having hierarchical structure.     

Acceptor specificity of the human leukocyte alpha3 fucosyltransferase: role of FucT-VII in the generation of selectin ligands

The alpha3 fucosyltransferase, FucT-VII, is one of the key glycosyltransferases involved in the biosynthesis of the sialyl Lewis X (sLex) antigen on human leukocytes. The sialyl Lewis X antigen (NeuAcalpha(2-3)Galbeta(1-4)[Fucalpha(1-3)]GlcNAc-R) is an essential component of the recruitment of leukocytes to sites of inflammation, mediating the primary interaction between circulating leukocytes and activated endothelium. In order to characterize the enzymatic properties of the leukocyte alpha3 fucosyltransferase FucT-VII, the enzyme has been expressed in Trichoplusia ni insect cells. The enzyme is capable of synthesizing both sLexand sialyl-dimeric-Lexstructures in vitro , from 3'-sialyl-lacNAc and VIM-2 structures, respectively, with only low levels of fucose transfer observed to neutral or 3'-sulfated acceptors. Studies using fucosylated NeuAcalpha(2-3)-(Galbeta(1- 4)GlcNAc)3-Me acceptors demonstrate that FucT-VII is able to synthesize both di-fucosylated and tri-fucosylated structures from mono- fucosylated precursors, but preferentially fucosylates the distal GlcNAc within a polylactosamine chain. Furthermore, the rate of fucosylation of the internal GlcNAc residues is reduced once fucose has been added to the distal GlcNAc. These results indicate that FucT-VII is capable of generating complex selectin ligands, in vitro , however the order of fucose addition to the lactosamine chain affects the rate of selectin ligand synthesis.      

A Radiation Hybrid Map of the BRCA1 Region

A locus on chromosome 17q, designated “BRCA1,” has been identified as a predisposition gene for breast cancer. A panel of chromosome 17–specific radiation-reduced somatic cell hybrid clones has been assembled for high-resolution mapping of chromosome 17. A series of 35 markers, known to span the BRCA1 locus, were tested against this hybrid panel by PCR assays. Statistical analysis of these data yields a BRCA1 radiation hybrid map at a density sufficient to initiate YAC cloning and pulsed-field gel electrophoretic mapping of the candidate region. In addition, many of the markers reveal genetic polymorphisms and may be tested in breast cancer families and in loss-of-heterozygosity studies of sporadic breast cancers to better define the BRCA1 gene candidate region.     

Accelerated breakdown of reticulocyte protein formed under conditions of amino acid depletion.

1. Labile protein is formed when rat or rabbit reticulocytes are incubated in medium deficient in individual amino acids, especially histidine, valine or alanine. The fraction of unstable protein is increased to about 35% of the total protein synthesized when the histidinyl-tRNA-charging inhibitor, histidinol, is added to histidine-deficient media. 2. The molecular weights of the labile proteins measured by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis in the presence of urea are less than haemoglobin and probably represent prematurely terminated haemoglobin chains. 3. Although protein synthesis is always lower under conditions that produce labile protein, inhibition of protein synthesis by fluoride or cycloheximide does not give an effect similar to amino acid depletion. 4. The synthesis of protein in deficient medium does not alter the degradation rate of pre-existing protein in reticulocytes and is thus unrelated to the stringent response in bacteria. 5. We propose that amino acid-deficient medium leads to a decreased charging of the appropriate tRNA, a concomitant decrease in protein synthesis and the degradation of nascent peptides.     

Metabolism of glucose, fructose and lactate in vivo in chronically cannulated foetuses and in suckling lambs.

1. Chronically cannulated sheep foetuses and suckling lambs were injected with 14C-labelled glucose, fructose or lactate, and sequential blood samples taken under conditions of minimal stress and without anaesthesia. 2. Gluconeogenesis from lactate was not detectable in foetal sheep, but the pathway was active in suckling lambs. 3. Fructose utilization rates were low in foetal sheep, with no measurable conversion into glucose or lactate. 4. The high rates of irreversible loss of both glucose and lactate in the foetus were decreased in suckling lambs. Radioactivity from labelled glucose entered both the lactate and fructose pools in foetal sheep, and entered the lactate pool in suckling lambs. 5. A model is proposed in which carbon flow between glucose, fructose and lactate has been quantified in foetal sheep.     

Freeze-fracture of the normal and pathological megakaryocyte lineage in chronic megakaryocytic-granulocytic myelosis

Freeze-fracture and thin sections were performed on human bone marrow of chronic megakaryocytic-granulocytic myelosis (CMGM) to study the three-dimensional fine structure and maturation of normal and atypical megakaryocytes and thrombocytes. In the many normally maturing megakaryocytes the development of the demarcation membrane system (DMS) was best investigated by comparison of thin sections with freeze-fracture replicas. The DMS shows no connections with the Golgi apparatus or rough-surfaced endoplasmic reticulum, but originates from tubular infoldings of the plasma membrane. These infoldings are always in continuity with the extracellular space and form an intracellular membranous pool by branching and coalescing of flattened tubules from which finally the perforated cisternae of the DMS arise. Freeze-fracture of the normal thrombocytes confirms earlier findings. The abnormal giant platelets seen in CMGM display extensive areas of smooth membranes of a spongy structure consisting of dense tubules surrounded by the labyrinth of the surface-connected system. Their physiological significance in these atypical platelets remains unsolved.     

Photo-initiated ion formation from octaethyl-porphyrin and its zinc chelate as a model for electron transfer in reaction centers.

Ion formation from the reaction of triplet (T) and ground state (P) octaethyl-porphyrin (OEP) and zinc octaethyl porphyrin (ZnOEP) and the corresponding cross-reactions have been measured in dry acetonitrile. A uniquely sensitive and fast conductance apparatus and a pulsed dye laser allowed the measurements to be made at the necessarily very low concentrations of T. The hemogeneous reaction of T (ZnOEP) and P (ZnOEP) occurs with rat constant k(1) = 2.0 x 10(8) M(-1)s(-1) and an ion yield of 67%. The similar homogeneous reaction of OEP has k(2) = 1.3 x 10(8)M(-1)s(-1) but an ion yield of only 3%. The cross-reaction of T (OEP) with P (ZnOEP) has k(3) = 1.5 x 10(8) M(-1)s(-1) and an ion yield of 27%, while the inverse cross-reaction of T (ZnOEP) with P (OEP) has k(4) = 3 x 10(8) M(-1)s(-1) and an ion yield of 20%. Thus, the rate constants are only slightly affected but the yields are sensitive to the porphyrin. The possible formation of the heterogeneous ions ZnOEP+ + OEP-, thermodynamically favored by 0.3 V over the homogeneous ions, has little influence on the observed yields. The data are explained by electron transfer and Coulomb field-electon spin-controlled escape of the initial ion-pair.     

Bovine mammary gland UDP-GalNAc:GlcNAcbeta-R beta1-->4-N- acetylgalactosaminyltransferase is glycoprotein hormone nonspecific and shows interaction with alpha-lactalbumin

We have identified a novel N -acetylgalactosaminyltransferase activity in lactating bovine mammary gland membranes. Acceptor specificity studies and analysis of products obtained in vitro by 400 MHz1H-NMR spectroscopy revealed that the enzyme catalyses the transfer of N - acetylgalactosamine (GalNAc) from UDP-GalNAc to acceptor substrates carrying a terminal, beta-linked N -acetylglucosamine (GlcNAc) residue and establishes a beta1-->4-linkage forming a GalNAcbeta1-->4GlcNAc ( N, N '-diacetyllactosediamine, lacdiNAc) unit. Therefore, the enzyme can be identified as a UDP-GalNAc:GlcNAcbeta-R beta1-->4-N- acetylgalactosaminyltransferase (beta4-GalNAcT). This enzyme resembles invertebrate beta4-GalNAcT as well as mammalian beta4- galactosyltransferase (beta4-GalT) in acceptor specificity. It can, however, be clearly distinguished from the pituitary hormone-specific beta4-GalNAcT by its incapability of acting with an elevated activity on a glycoprotein substrate carrying a hormone-specific peptide motif. Furthermore, the GalNAcT activity appeared not to be due to a promiscuous action of a beta4-GalT as could be demonstrated by comparing the beta4-GalNAcT and beta4-GalT activities of the mammary gland, bovine colostrum, and purified beta4-GalT, by competition studies with UDP-GalNAc and UDP-Gal, and by use of an anti-beta4-GalT polyclonal inhibiting antibody. Interestingly, under conditions where mammalian beta4-GalT forms with alpha-lactalbumin (alpha-LA) the lactose synthase complex, the mammary gland beta4-GalNAcT was similarly induced by alpha-LA to act on Glc with an increased efficiency yielding the lactose analog GalNAcbeta1-->4Glc. This enzyme thus forms the second example of a mammalian glycosyltransferase the specificity of which can be modified by this milk protein. It is proposed that the mammary gland beta4-GalNAcT functions in the synthesis of lacdiNAc- based, complex-type glycans frequently occurring on bovine milk glycoproteins. The action of this enzyme is to be considered when aiming at the production of properly glycosylated protein biopharmaceuticals in the milk of transgenic dairy animals.