Phylogenetic analyses suggest a hybrid origin of the figs (Moraceae: Ficus) that are endemic to the Ogasawara (Bonin) Islands, Japan

The Ogasawara Islands are oceanic islands and harbor a unique endemic flora. There are three fig species (Ficus boninsimae, F. nishimurae and F. iidaiana) endemic to the Ogasawara Islands, and these species have been considered to be closely related to Ficus erecta, and to have diverged within the islands. However, this hypothesis remains uncertain. To investigate this issue, we assessed the phylogenetic relationships of the Ogasawara figs and their close relatives occurring in Japan, Taiwan and South China based on six plastid genome regions, nuclear ITS region and two nuclear genes. The plastid genome-based tree indicated a close relationship between the Ogasawara figs and F. erecta, whereas some of the nuclear gene-based trees suggested this relationship was not so close. In addition, the phylogenetic analyses of the pollinating wasps associated with these fig species based on the nuclear 28S rRNA and mitochondrial cytB genes suggested that the fig-pollinating wasps of F. erecta are not sister to those of the Ogasawara figs These results suggest the occurrence of an early hybridization event(s) in the lineage leading to the Ogasawara figs.     

Laminin α1 is essential for mouse cerebellar development

Laminin α1 (Lama1), which is a subunit of laminin-1 (laminin-111), a heterotrimeric ECM protein, is essential for embryonic development and promotes neurite outgrowth in culture. Because the deletion of Lama1 causes lethality at early embryonic stages in mice, the in vivo role of Lama1 in neural development and functions has not yet been possible to determine. In this study, we generated conditional Lama1 knockout (Lama1(CKO)) mice in the epiblast lineage using Sox2-Cre mice. These Lama1(CKO) mice survived, but displayed behavioral disorders and impaired formation of the cerebellum. Deficiency of Lama1 in the pial basement membrane of the meninges resulted in defects in the conformation of the meninges. During cerebellar development, Lama1 deficiency also caused a decrease in the proliferation and migration of granule cell precursors, disorganization of Bergmann glial fibers and endfeet, and a transient reduction in the activity of Akt. A marked reduction in numbers of dendritic processes in Purkinje cells was observed in Lama1(CKO) mice. Together, these results indicate that Lama1 is required for cerebellar development and functions.     

TDP2 promotes repair of topoisomerase I-mediated DNA damage in the absence of TDP1

The abortive activity of topoisomerases can result in clastogenic and/or lethal DNA damage in which the topoisomerase is covalently linked to the 3'- or 5'-terminus of a DNA strand break. This type of DNA damage is implicated in chromosome translocations and neurological disease and underlies the clinical efficacy of an important class of anticancer topoisomerase 'poisons'. Tyrosyl DNA phosphodiesterase-1 protects cells from abortive topoisomerase I (Top1) activity by hydrolyzing the 3'-phosphotyrosyl bond that links Top1 to a DNA strand break and is currently the only known human enzyme that displays this activity in cells. Recently, we identified a second tyrosyl DNA phosphodiesterase (TDP2; aka TTRAP/EAPII) that possesses weak 3'-tyrosyl DNA phosphodiesterase (3'-TDP) activity, in vitro. Herein, we have examined whether TDP2 contributes to the repair of Top1-mediated DNA breaks by deleting Tdp1 and Tdp2 separately and together in murine and avian cells. We show that while deletion of Tdp1 in wild-type DT40 cells and mouse embryonic fibroblasts decreases DNA strand break repair rates and cellular survival in response to Top1-induced DNA damage, deletion of Tdp2 does not. However, deletion of both Tdp1 and Tdp2 reduces rates of DNA strand break repair and cell survival below that observed in Tdp1(-)(/)(-) cells, suggesting that Tdp2 contributes to cellular 3'-TDP activity in the absence of Tdp1. Consistent with this idea, over-expression of human TDP2 in Tdp1(-)(/)(-)/Tdp2(-)(/)(-)(/)(-) DT40 cells increases DNA strand break repair rates and cell survival above that observed in Tdp1(-)(/)(-) DT40 cells, suggesting that Tdp2 over-expression can partially complement the defect imposed by loss of Tdp1. Finally, mice lacking both Tdp1 and Tdp2 exhibit greater sensitivity to Top1 poisons than do mice lacking Tdp1 alone, further suggesting that Tdp2 contributes to the repair of Top1-mediated DNA damage in the absence of Tdp1. In contrast, we failed to detect a contribution for Tdp1 to repair Top2-mediated damage. Together, our data suggest that Tdp1 and Tdp2 fulfil overlapping roles following Top1-induced DNA damage, but not following Top2-induced DNA damage, in vivo.     

Nuclear-Cycle Dependence of the Development of the Preprophase Band in Tobacco BY-2 Cells

When tobacco BY-2 cells that had been treated with aphidicolinfor 24 h were cultured in the absence of aphidicolin, DNA synthesiswas initiated within 1 h. DNA synthesis was completed within4 h and then the preprophase band of microtubules (PPB) developed.However, when cells that had been treated with aphidicolin werecultured in the absence of aphidicolin for 1 h and then againin its presence, DNA synthesis, which was initiated during thehour in the absence of aphidicolin, was not completed withina further 10 h in the presence of aphidicolin. Moreover, thePPB did not develop even after the PPB had appeared and disappearedin cells that were cultured contemporaneously in the continuedabsence of aphidicolin. The development of the PPB seems to be causally associated withthe nuclear cycle of cell division in tobacco BY-2 cells. Thisprocess seems to require the completion of the replication ofall, or almost all, of the nuclear DNA. (Received July 25, 1992; Accepted November 24, 1992)     

A novel method of identifying genetic mutations using an electrochemical DNA array

We describe the development of a new type of DNA array chip that utilizes electrochemical reactions and a novel method of simultaneously identifying multiple genetic mutations on an array chip. The electrochemical array (ECA) uses a threading intercalator specific to double-stranded nucleotides, ferrocenylnaphthalene diimide (FND), as the indicator. ECA does not require target labeling, and the equipment is simple, durable and less expensive. The simultaneous multiple mutation detection (SMMD) system using an ECA chip and FND utilizes an enzyme to simultaneously distinguish several genetic mutations such as single nucleotide polymorphism (SNP), insertion, deletion, translocation and short tandem repeat. We examined this SMMD system using an ECA chip, by detecting seven different mutations on the lipoprotein lipase (LPL) gene for 50 patients in a blind test. It turned out that all the results obtained were concordant with the sequencing results, demonstrating that this system is a powerful tool for clinical applications.     

Synaptic scaffolding molecule (S-SCAM) membrane-associated guanylate kinase with inverted organization (MAGI)-2 is associated with cell adhesion molecules at inhibitory synapses in rat hippocampal neurons

Synaptic scaffolding molecule (S-SCAM) is a synaptic protein, which harbors five or six PSD-95/Discs large/ZO-1 (PDZ), a guanylate kinase and two WW domains. It interacts with NMDA receptor subunits, neuroligin and beta-catenin, and is involved in the accumulation of neuroligin at excitatory synapses. In this study, we have demonstrated S-SCAM is localized at inhibitory synapses in rat primary cultured hippocampal neurons. We have identified beta-dystroglycan (beta-DG) as a binding partner for S-SCAM at inhibitory synapses. WW domains of S-SCAM bind to three sequences of beta-DG. We have also revealed that S-SCAM can interact with neuroligin 2, which is known to be exclusively localized at inhibitory synapses. The WW domains and the second PDZ domain of S-SCAM are involved in the interaction with neuroligin 2. Beta-DG, neuroligin 2 and S-SCAM form a tripartite complex in vitro. Neuroligin 2 is detected in the immunoprecipitates by anti-beta-DG antibody from rat brain. S-SCAM, beta-DG and neuroligin 2 are partially co-localized in rat hippocampal neurons. These data suggest that S-SCAM is associated with beta-DG and neuroligin 2 at inhibitory synapses, and functions as a linker between the dystrophin glycoprotein complex and the neurexin-neuroligin complex.     

Participation of glutamate-354 of the CP43 polypeptide in the ligation of manganese and the binding of substrate water in photosystem II

In the current X-ray crystallographic structural models of photosystem II, Glu354 of the CP43 polypeptide is the only amino acid ligand of the oxygen-evolving Mn(4)Ca cluster that is not provided by the D1 polypeptide. To further explore the influence of this structurally unique residue on the properties of the Mn(4)Ca cluster, the CP43-E354Q mutant of the cyanobacterium Synechocystis sp. PCC 6803 was characterized with a variety of biophysical and spectroscopic methods, including polarography, EPR, X-ray absorption, FTIR, and mass spectrometry. The kinetics of oxygen release in the mutant were essentially unchanged from those in wild type. In addition, the oxygen flash yields exhibited normal period four oscillations having normal S state parameters, although the yields were lower, correlating with the mutant's lower steady-state rate (approximately 20% compared to wild type). Experiments conducted with H(2)(18)O showed that the fast and slow phases of substrate water exchange in CP43-E354Q thylakoid membranes were accelerated 8.5- and 1.8-fold, respectively, in the S(3) state compared to wild type. Purified oxygen-evolving CP43-E354Q PSII core complexes exhibited a slightly altered S(1) state Mn-EXAFS spectrum, a slightly altered S(2) state multiline EPR signal, a substantially altered S(2)-minus-S(1) FTIR difference spectrum, and an unusually long lifetime for the S(2) state (>10 h) in a substantial fraction of reaction centers. In contrast, the S(2) state Mn-EXAFS spectrum was nearly indistinguishable from that of wild type. The S(2)-minus-S(1) FTIR difference spectrum showed alterations throughout the amide and carboxylate stretching regions. Global labeling with (15)N and specific labeling with l-[1-(13)C]alanine revealed that the mutation perturbs both amide II and carboxylate stretching modes and shifts the symmetric carboxylate stretching modes of the α-COO(-) group of D1-Ala344 (the C-terminus of the D1 polypeptide) to higher frequencies by 3-4 cm(-1) in both the S(1) and S(2) states. The EPR and FTIR data implied that 76-82% of CP43-E354Q PSII centers can achieve the S(2) state and that most of these can achieve the S(3) state, but no evidence for advancement beyond the S(3) state was observed in the FTIR data, at least not in a majority of PSII centers. Although the X-ray absorption and EPR data showed that the CP43-E354Q mutation only subtly perturbs the structure and spin state of the Mn(4)Ca cluster in the S(2) state, the FTIR and H(2)(18)O exchange data show that the mutation strongly influences other properties of the Mn(4)Ca cluster, altering the response of numerous carboxylate and amide groups to the increased positive charge that develops on the cluster during the S(1) to S(2) transition and weakening the binding of both substrate water molecules (or water-derived ligands), especially the one that exchanges rapidly in the S(3) state. The FTIR data provide evidence that CP43-Glu354 coordinates to the Mn(4)Ca cluster in the S(1) state as a bridging ligand between two metal ions but provide no compelling evidence that this residue changes its coordination mode during the S(1) to S(2) transition. The H(2)(18)O exchange data provide evidence that CP43-Glu354 interacts with the Mn ion that ligates the substrate water molecule (or water-derived ligand) that is in rapid exchange in the S(3) state.     

Papillary serous carcinoma of the peritoneum with paraaortic lymph node metastasis despite minimal intraperitoneal involvement: a case report

BACKGROUND; Papillary serous carcinoma of the peritoneum (PSCP) is a tumor that produces widespread intraperitoneal lesions. Unlike serous ovarian adenocarcinoma, many aspects of its mode of progression and biologic characteristics are unclear. CASE: A 46-year-old woman with PSCP had no detectable ascites and minimal intraperitoneal involvement at the time of the diagnosis, but a paraaortic lymph node metastasis was present. Preoperative endometrial cytology was positive (suspicion of adenocarcinoma). The histologic diagnosis was poorly differentiated serous adenocarcinoma. Cytology of the peritoneal washings demonstrated positive findings, similar to those of endometrial cytology. After cytoreductive surgery, including lymph node dissection and platinum-based chemotherapy, the patient achieved long-term survival. CONCLUSION: PSCP can present with an early paraaortic lymph node metastasis. Endometrial cytology can be valuable in the diagnosis.