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Biomedical subjects

J Harwood

Publications and source records attributed to J Harwood.

At least 19 recordsLinked to original sources
PubMed2001

Human origin recognition complex binds to the region of the latent origin of DNA replication of Epstein-Barr virus.

Epstein-Barr virus (EBV) replicates in its latent phase once per cell cycle in proliferating B cells. The latent origin of DNA replication, oriP, supports replication and stable maintenance of the EBV genome. OriP comprises two essential elements: the dyad symmetry (DS) and the family of repeats (FR), both containing clusters of binding sites for the transactivator EBNA1. The DS element appears to be the functional replicator. It is not yet understood how oriP-dependent replication is integrated into the cell cycle and how EBNA1 acts at the molecular level. Using chromatin immunoprecipitation experiments, we show that the human origin recognition complex (hsORC) binds at or near the DS element. The association of hsORC with oriP depends on the DS element. Deletion of this element not only abolishes hsORC binding but also reduces replication initiation at oriP to background level. Co-immunoprecipitation experiments indicate that EBNA1 is associated with hsORC in vivo. These results indicate that oriP might use the same cellular initiation factors that regulate chromosomal replication, and that EBNA1 may be involved in recruiting hsORC to oriP.

Animals↗
PubMed2001

Comparing grandchildren's and grandparents' stake in their relationship.

This paper examines the intergenerational stake hypothesis in the grandparent-grandchild relationship. The hypothesis predicts that older generations will perceive more closeness in a relationship than younger generations. Grandparent-grandchild dyads are surveyed concerning various aspects of their relationship. Using measures of closeness and an established typology of grandparent-grandchild relationship style (Cherlin & Furstenberg, 1985), the study finds that grandparents perceive their relationship with their grandchildren to be considerably closer than their grandchildren perceive the relationship. However, their grandchildren perceive the relationship to be more active than their grandparents do. Explanations of these findings consistent with the intergenerational stake hypothesis are advanced. It is found that the two parties often disagree as to the nature of their relationship on the Cherlin and Furstenberg typology. Details of these disagreements are examined in detail, and implications for the grandparent-grandchild relationship are discussed.

Adolescent↗
PubMed2001

Data quality probes--a synergistic method for quality monitoring of electronic medical record data accuracy and healthcare provision.

Increasing reliance is being placed on electronic medical records to support clinical care and achieve improved quality standards. In order for clinical information systems (CIS) to deliver, the data within needs to be complete, consistent and accurate. This data of course only forms part of the process in delivering quality health care during the clinician-patient encounter. This paper outlines a method of assessing the quality of the processes involved in healthcare provision and data quality within a CIS. It proposes a principle of Data Quality Probes (DQP) that can be used to assess the performance of the whole encounter system. The main feature of this is the generation of a query which clinical knowledge predicts should not retrieve any cases in a system performing flawlessly. This approach is applied practically within the paradigm of a UK family practice testing the DQP that only patients who have had a hysterectomy should be prescribed unopposed oestrogen hormone replacement therapy.

Estrogen Replacement Therapy↗
PubMed2000

The rediscovery of Mendelism in agricultural context: Erich von Tschermak as plant-breeder.

Why was Mendelism rediscovered? One way in which historians have addressed this issue is to look at wider trends in research during the 1890s of which the rediscoverers were part. Quite a lot is known about one such research tradition, namely the attempts to resolve the question of evolutionary mechanism through the use of varietal crosses. But another relevant research tradition is still largely unknown: the work of commercial breeders, several of whom were using hybridisation by the 1890s. In this paper I begin by looking at Tschermak's initial career, the sequence of events by which he came upon Mendel's work, and why he was excited by what he read. Then I place Tschermak's early work in the context of commercial plant-breeding in German-speaking Europe toward the end of the 19th century. Finally I look again at the question of Tschermak's somewhat ambivalent relationship to Mendelism after 1900. I argue that his initial misunderstanding of the concept of segregation was due to the fact that he approached Mendel's work with the perspective of a breeder rather than that of a geneticist.

Agriculture↗
PubMed2000

Structural determinants of PIP(2) regulation of inward rectifier K(ATP) channels.

Phosphatidylinositol 4,5-bisphosphate (PIP(2)) activates K(ATP) and other inward rectifier (Kir) channels. To determine residues important for PIP(2) regulation, we have systematically mutated each positive charge in the COOH terminus of Kir6.2 to alanine. The effects of these mutations on channel function were examined using (86)Rb efflux assays on intact cells and inside-out patch-clamp methods. Both methods identify essentially the same basic residues in two narrow regions (176-222 and 301-314) in the COOH terminus that are important for the maintenance of channel function and interaction with PIP(2). Only one residue (R201A) simultaneously affected ATP and PIP(2) sensitivity, which is consistent with the notion that these ligands, while functionally competitive, are unlikely to bind to identical sites. Strikingly, none of 13 basic residues in the terminal portion (residues 315-390) of the COOH terminus affected channel function when neutralized. The data help to define the structural requirements for PIP(2) sensitivity of K(ATP) channels. Moreover, the regions and residues defined in this study parallel those uncovered in recent studies of PIP(2) sensitivity in other inward rectifier channels, indicating a common structural basis for PIP(2) regulation.

Adenosine Triphosphate↗
PubMed1998

Oxygen induces fatty acid (n-6)-desaturation independently of temperature in Acanthamoeba castellanii.

Induction of a microsomal oleate delta12 (n-6) desaturase which is mainly responsible for an increase in membrane lipid unsaturation at low temperature has been observed in the free-living amoeba Acanthamoeba castellanii. In this study we show that the enzyme can also be regulated by oxygen independently of temperature in batch cultures grown to O2-limitation. Raising the oxygen concentration from below the lower limit of detection (< 0.1 microM) to approximately air-saturation (230 microM), whilst maintaining the growth temperature constant (30 degrees C), increased lipid unsaturation and elevated n-6-desaturase activity 2.3-fold. Addition of the protein synthesis inhibitor, anisomycin, showed that increased desaturase activity was due to new protein synthesis rather than activation of pre-existing enzyme. These observations are important for future studies of the mechanism of temperature adaptation in poikilotherms.

Acanthamoeba↗
PubMed1998

Factors affecting levels of genetic diversity in natural populations.

Genetic variability is the clay of evolution, providing the base material on which adaptation and speciation depend. It is often assumed that most interspecific differences in variability are due primarily to population size effects, with bottlenecked populations carrying less variability than those of stable size. However, we show that population bottlenecks are unlikely to be the only factor, even in classic case studies such as the northern elephant seal and the cheetah, where genetic polymorphism is virtually absent. Instead, we suggest that the low levels of variability observed in endangered populations are more likely to result from a combination of publication biases, which tend to inflate the level of variability which is considered 'normal', and inbreeding effects, which may hasten loss of variability due to drift. To account for species with large population sizes but low variability we advance three hypotheses. First, it is known that certain metapopulation structures can result in effective population sizes far below the census size. Second, there is increasing evidence that heterozygous sites mutate more frequently than equivalent homozygous sites, plausibly because mismatch repair between homologous chromosomes during meiosis provides extra opportunities to mutate. Such a mechanism would undermine the simple relationship between heterozygosity and effective population size. Third, the fact that related species that differ greatly in variability implies that large amounts of variability can be gained or lost rapidly. We argue that such cases are best explained by rapid loss through a genome-wide selective sweep, and suggest a mechanism by which this could come about, based on forced changes to a control gene inducing coevolution in the genes it controls. Our model, based on meiotic drive in mammals, but easily extended to other systems, would tend to facilitate population isolation by generating molecular incompatabilities. Circumstances can even be envisioned in which the process could provide intrinsic impetus to speciation.

Animals↗
PubMed1998

Expectations for communication with positive and negative subtypes of older adults.

The study uses a recently-developed scale for eliciting perceptions, expectations, and evaluations of intergenerational communication. As predicted, it is found that younger adults expect to experience more anxiety, receive more complaining, and receive lower levels of attunement from an older adult who is portrayed as "despondent" than one who is portrayed as a "perfect grandparent." In addition, younger adults with more negative attitudes toward older adults expect to experience more negative effect, anxiety, and communication apprehension, to feel more compassion for the older adult, and to receive lower levels of attunement and more complaining from the older adult than those with more positive attitudes. Surprisingly, younger adults with higher levels of young age identification expect to experience lower levels of apprehension, more attunement from the older adult, and to feel more compassion for the older adult than those with lower levels of age identity. These findings are discussed in terms of theoretical models of intergenerational communication, in particular the communication predicament model. In addition, younger people's feelings of having "helped" an older person are discussed in the context of intergroup theory.

Adult↗
PubMed1997

Cdc6p-dependent loading of Mcm proteins onto pre-replicative chromatin in budding yeast.

The Cdc6 protein is essential for the assembly of pre-replicative complexes (pre-RCs) at origins of DNA replication in the budding yeast Saccharomyces cerevisiae. This reaction is blocked in vivo by the cyclin-dependent kinase Cdc28p, together with its regulatory subunits, the B type cyclins that are present throughout S, G2, and M phases. Because the destruction of B type cyclins and the consequent inactivation of the kinase are essential for exit from mitosis, pre-RC formation can only occur after passage through mitosis. Therefore, pre-RC formation has been proposed to be essential for coupling S phase and mitosis and for limiting DNA replication to once per cell cycle. The Mcm2-7 family of proteins has been implicated in limiting replication to once per cell cycle from experiments with Xenopus egg extracts. Here we show that the Mcm proteins of budding yeast are abundant and are quantitatively found in a chromatin-enriched fraction specifically during the G1 phase of the cell cycle. This chromatin binding depends on the de novo synthesis of Cdc6p, providing evidence that a conserved biochemical pathway plays a critical role in coordinating DNA replication with mitosis in both yeast and higher eukaryotes. Cdc6p and the origin recognition complex can be selectively removed from this chromatin-enriched fraction without removing the Mcm proteins. From these results, we propose that Cdc6p (and the origin recognition complex) nucleates the binding of Mcm proteins to chromatin, but once bound, the Mcm proteins appear to interact tightly with some other component of chromatin.

Cell Cycle↗
PubMed1996

Contribution of coplanar and non-coplanar polychlorinated biphenyls to the toxic equivalence of grey seal (Halichoerus grypus) milk.

We measured the concentration of coplanar polychlorinated biphenyl (PCB) congeners #77, #126 and #169 in the milk of 21 individual grey seals (Halichoerus grypus) from the same Scottish colony. Using the toxic equivalency factors (TEFs) of Ahlborg et al. (1994) we found TEQs ranged between 8.4 and 45.7 pg/g, with a mean of 22.3 +/- 9.3 pg/g on a lipid weight basis (n = 32). For some samples we also had concentrations of mono-ortho and di-orthochlorinated PCBs. Inclusion of these congeners approximately doubled the TEQ. There was a good correlation between the concentration of non-ortho PCBs and that of other congeners in the same sample.

Animals↗
PubMed1996

Worldwide patterns of mitochondrial DNA differentiation in the harbor seal (Phoca vitulina).

The harbor seal (Phoca vitulina) has one of the broadest geographic distributions of any pinniped, stretching from the east Baltic, west across the Atlantic and Pacific Oceans to southern Japan. Although individuals may travel several hundred kilometers on annual feeding migrations, harbor seals are generally believed to be philopatric, returning to the same areas each year to breed. Consequently, seals from different areas are likely to be genetically differentiated, with levels of genetic divergence increasing with distance. Differentiation may also be caused by long-standing topographic barriers such as the polar sea ice. We analyzed samples of 227 harbor seals from 24 localities and defined 34 genotypes based on 435 bp of control region sequence. Phylogenetic analysis and analysis of molecular variance showed that populations in the Atlantic and Pacific Oceans and east and west coast populations of these oceans are significantly differentiated. Within these four regions, populations that are geographically farthest apart generally are the most differentiated and often do not share genotypes or differ in genotype frequency. The average corrected sequence divergence between populations in the Atlantic and Pacific Oceans is 3.28% +/- 0.38% and those among populations within each of these oceans are 0.75% +/- 0.69% and 1.19% +/- 0.65%, respectively. Our results suggest that harbor seals are regionally philopatric, on the scale of several hundred kilometers. However, genetic discontinuities may exist, even between neighboring populations such as those on the Scottish and east English coasts or the east and west Baltic. The mitochondrial data are consistent with an ancient isolation of populations in both oceans, due to the development of polar sea ice. In the Atlantic and Pacific, populations appear to have been colonized from west to east with the European populations showing the most recent common ancestry. We suggest the recent ancestry of European seal populations may reflect recolonization from Ice Age refugia after the last glaciation.

Animals↗
PubMed1996

Identification of a domain in Rck, a product of the Salmonella typhimurium virulence plasmid, required for both serum resistance and cell invasion.

Rck is encoded on the Salmonella typhimurium virulence plasmid and is a member of a family of related 17- to 19-kDa outer membrane proteins of Enterobacteriaceae, including Ail (Yersinia enterocolitica) and PagC (S. typhimurium). Structural models for these proteins predict eight membrane-spanning domains alternating with hydrophilic inner and outer loops. When expressed in Escherichia coli, Rck and Ail, but not PagC, confer high-level resistance to the bactericidal activity of complement as well as the ability to adhere to and invade mammalian cell lines. To identify functional domains of Rck, we made and screened random mutations in Rck for decreased bioactivity. We found that a single amino acid substitution (glycine to aspartic acid) in the putative third outer loop greatly reduced Rck-mediated serum resistance and eukaryotic cell invasion. We then constructed two chimeric proteins between Rck and PagC. Substitution of the C-terminal half of Rck with the corresponding PagC fragment containing both the third and the fourth outer loops abolishes the Rck-mediated serum resistance and invasion phenotypes. Substitution of Rck with a smaller C-terminal portion of PagC containing the fourth outer loop did not affect the invasive phenotype or serum resistance. These data reveal that the third putative outer membrane loop region is important for the virulence-associated properties of the Rck protein and suggest a similarity between the mechanism of serum resistance and epithelial cell invasion involving the same domain of Rck.

Amino Acid Sequence↗