Species-specific sperm adhesion in sea urchins. A quantitative investigation of bindin- mediated egg agglutination

Bindin, a protein component of the acrosomal vesicle of sea urchin sperm, has been isolated from Arbacia punctulata and strongylocentrous purpuratus. Using this isolated bindin, we have devised a quantitative assay for bindin-mediated egg agglutination and compared the agglutination of bindin eggs from A. puntulata and S. purpuratus. Bindin- mediated agglutination is species –specific in both species, although a measurable degree of heterotypic interaction is observed. Homotypic bindin-egg interactions differ significantly from heterotypic interactions both in the extent of agglutination and the size of the resulting aggregates. We also provide direct evidence that bindin particles agglutinate eggs by adhering to the surfaces of adjacent eggs. Although the A. punctulata bindin preparation displays the same functional properties and consists of one major polypeptide of the same apparent molecular weight as S. purpuratus bindin, its morphology is very different. Unlike the spherical aggregates observed with S. purpuratus bindin, A punctulata bindin exists as lamellar vesicles and binds significant amounts of phospholipids and Triton X-100, suggesting that it may be tightly associated with the acrosomal membrane. Having defined a number of the basic parameters of bindin-mediated agglutination, we examined the effect of a number of saccharides and glycopeptides on bindin-mediated egg agglutination. Carbohydrate-containing components derived from the egg cell surface by proteolysis were found to inhibit bindin-mediated egg agglutination at low concentrations, but this inhibition is not species specific.     

Preventing DNA over-replication: a Cdk perspective

The basal-like breast cancer, a new category of breast cancer associated with poor prognosis and possibly unique chemosensitivity, is a current topic in the breast cancer field. Evidence from multiple sources strongly indicate that impairment of BRCA1 pathways is responsible for this phenotype, implying the importance of BRCA1 not only in familial breast cancers but also in sporadic cancers. BRCA1 acts as a hub protein that coordinates a diverse range of cellular pathways to maintain genomic stability. BRCA1 participates in multiple cellular supercomplexes to execute its tasks and, in most of the complexes, BRCA1 exists as a RING heterodimer with BARD1 to provide ubiquitin E3 ligase activity that is required for its tumor suppressor function. It was revealed recently that the BRCA1 RING finger is capable of catalyzing multiple types of ubiquitination depending upon the interacting E2, the ubiquitin carrier protein. BRCA1 may catalyze distinct ubiquitination on different substrates as the situation demands. On the other hand, in response to DNA double-strand breaks where BRCA1 plays its major role for homologous recombination repair, recent evidence showed that ubiquitination is a critical step to recruit BRCA1 to the damaged site through UIM (ubiquitin interacting motif) containing protein RAP80. Thus, ubiquitin and BRCA1 likely affect each other in many ways to perform cellular functions. Elucidation of this mechanism in relation to cell survival is now much anticipated because it could be a key to predict chemosensitivity of basal-like breast cancer.     

Lectin histochemistry for detecting cadmium-induced changes in the glycosylation pattern of rat placenta

Cadmium (Cd) is an industrial and environmental pollutant that produces toxic effects on gametogenesis, pre- and post-implantation embryos, and the placenta. Because the effects of acute Cd intoxication on the placenta are not well understood, we investigated changes in its glycosylated components in Cd treated dams at days 4, 7, 10 and 15 of gestation using lectin histochemistry. CdCl₂ was administered to pregnant rats; control animals received sterile normal saline. Placentas were processed for DBA, Con A, SBA, PNA, UEA-I, RCA-I and WGA lectin histochemistry to evaluate changes in the carbohydrate pattern of the placenta that might modify cell interactions and contribute to embryonic alterations. Lectin binding was analyzed in the yolk sac; trophoblast giant cells; trophoblast I, II and III; spongiotrophoblast cells and endovascular trophoblast cells in the chorioallantoic placenta. Our lectin binding patterns showed that Cd caused alteration of SBA and DBA labeling of trophoblast-derived cells, which suggested increased expressions of α and β GalNAc. Cd also caused decreased UEA-1 binding affinity, which indicated fewer α-L-Fuc residues in placentas of Cd treated dams. The nonreactivity in trophoblast I of the control placentas incubated with Con-A contrasted with the labeling in placentas of experimental dams, which indicated increased expression of terminal α-D-Man, and α-D-Glc residues. We found that Cd altered the reactivity of placenta to several lectins, which indicated modification of the glycotype presented by the fetal component of the placenta. We report that Cd exerts a deleterious effect on the glycosylation pattern of the placenta.     

Overexpression of the ped/pea-15 gene causes diabetes by impairing glucose-stimulated insulin secretion in addition to insulin action

Overexpression of the ped/pea-15 gene is a common feature of type 2 diabetes. In the present work, we show that transgenic mice ubiquitously overexpressing ped/pea-15 exhibited mildly elevated random-fed blood glucose levels and decreased glucose tolerance. Treatment with a 60% fat diet led ped/pea-15 transgenic mice to develop diabetes. Consistent with insulin resistance in these mice, insulin administration reduced glucose levels by only 35% after 45 min, compared to 70% in control mice. In vivo, insulin-stimulated glucose uptake was decreased by almost 50% in fat and muscle tissues of the ped/pea-15 transgenic mice, accompanied by protein kinase Calpha activation and block of insulin induction of protein kinase Czeta. These changes persisted in isolated adipocytes from the transgenic mice and were rescued by the protein kinase C inhibitor bisindolylmaleimide. In addition to insulin resistance, ped/pea-15 transgenic mice showed a 70% reduction in insulin response to glucose loading. Stable overexpression of ped/pea-15 in the glucose-responsive MIN6 beta-cell line also caused protein kinase Calpha activation and a marked decline in glucose-stimulated insulin secretion. Antisense block of endogenous ped/pea-15 increased glucose sensitivity by 2.5-fold in these cells. Thus, in vivo, overexpression of ped/pea-15 may lead to diabetes by impairing insulin secretion in addition to insulin action.     

Phylogeny and biogeography of ratite birds inferred from DNA sequences of the mitochondrial ribosomal genes

The origin of the flightless ratite birds of the southern continents has been debated for over a century. Whether dispersal or vicariance (continental breakup) best explains their origin depends largely on their phylogenetic relationships. No consensus has been reached on this issue despite many morphological and molecular studies. To address this question further we sequenced a 2.8-kb region of mitochondrial DNA containing the ribosomal genes in representative ratites and a tinamou. Phylogenetic analyses indicate that Struthio (Africa) is basal and Rhea (South America) clusters with living Australasian ratites. This phylogeny agrees with transferrin and DNA hybridization studies but not with sequence analyses of some protein-coding genes. These results also require reevaluation of the phylogenetic position of the extinct moas of New Zealand. We propose a new hypothesis for the origin of ratites that combines elements of dispersal and vicariance.      

Griseofulvin-induced aneuploidy and meiotic delay in female mouse germ cells. II. Cytogenetic analysis of one-cell zygotes.

The effects of griseofulvin (GF) upon the first meiotic division of female mouse germ cells were evaluated by cytogenetic analysis of first-cleavage (1-Cl) zygotes. The present study is an extension of an investigation that began with the cytogenetic analysis of metaphase II (M II) oocytes. Different doses (200, 666, 1332, 2000 mg/kg) were tested by oral administration of GF to superovulated animals either at the time of human chorionic gonadotrophin (HCG) injection or 2 h post HCG. When GF was given at the time of HCG, significant dose-dependent increases of different types of cytogenetically abnormal cells were found. These included zygotes containing ostensibly female-derived M I or M II arrested chromosomes and polyploid zygotes. The total yields of these aberrations were 2.9, 4.3, 26.2, 60.6, and 64.1% for control, 200, 666, 1332, and 2000 mg/kg, respectively. The origin of these zygotes was attributed to the fertilization of oocytes that had been previously arrested at M I. No significant induction of hyperploidy was detected. Developmentally abnormal zygotes were still observed when GF was administered 2 h post HCG, although their frequencies were significantly lower than in the first series of experiments. The yields of developmentally abnormal zygotes were 49, 10.2, and 23.6% at 200, 666, and 2000 mg/kg. Additionally, a dose-dependent increase in the frequency of hyperploid zygotes was detected up to a maximum of 36.5% at 2000 mg/kg. These results confirm the cytogenetic observations from M II oocytes after GF treatment under the same experimental conditions; namely, a dramatic change in the oocyte target susceptibility to GF occurred within a short time period. Also, the present study demonstrated that most of GF-induced aneuploid oocytes were fertilized and reached first-cleavage metaphase.