Patrick K.K. Yeung

Mimosine, the allelochemical from the leguminous tree Leucaena leucocephala, selectively enhances cell proliferation in dinoflagellates

ABSTRACT Mimosine, the allelochemical from the leguminous tree Leucaena leucocephala, is toxic to most terrestrial animals and plants. We report here that while mimosine inhibits major phytoplankton groups, it enhances cell proliferation in dinoflagellates. On addition to coastal seawater samples, mimosine is able to confer a growth advantage to dinoflagellates. The use of mimosine will promote the isolation and culture of this group of phytoplankton.

Characterization of a Karenia papilionacea -like dinoflagellate from the South China Sea

The morphological characteristics of a new isolate of a dorso-ventrally flattened athecate dinoflagellate obtained from Hong Kong (HK) waters is described. Partial sequences of the large subunit ribosomal RNA gene were obtained and a high degree of similarity was observed with various species within the genus Karenia. Phylogenetic comparison suggested the Karenia sp. HK forms a clade with the newly described species K. papilionacea from New Zealand but not with authentic K. brevis from North America. Karenia sp. HK is the first K. brevis-like dinoflagellate characterized by molecular cladistics from the South China Sea and the results call for re-examination of the status of other K. brevis-like morphotypes described from the region.

Large subunit rDNA sequences from Alexandrium catenella strains isolated during harmful algal blooms in Hong Kong

The partial sequences of the large subunit (24S) ribosomal RNA gene(LsurRNA) of three Alexandrium catenella strains isolatedduring the 1998 and 1989 red tide incidents in Hong Kong were compared,togetherwith the sequences of other Asian A. catenella speciescomplex. The sequences of two A. catenella strainsisolatedin 1998 were identical, despite being isolated at different time (March andApril) and different locations (eastern and southern Hong Kong). In cladisticanalysis, the two A. catenella HK98A & B strains, theA. catenella HK1989 strain, the Chinese strain clade andtheKorean/Japanese A. catenella form a clade distinct fromother A. catenella strains. Interestingly, the twoA. catenella HK98A & B strains only form a clade withtheA. catenella HK1989 strain/ChineseA. catenella (CCMP1493) with low bootstrap values (49%).

Cyclic ADP-ribose links metabolism to multiple fission in the dinoflagellate Crypthecodinium cohnii

Cellular metabolism is required for cell proliferation. However, the way in which metabolic signals are conveyed to cell cycle decisions is unclear. Cyclic ADP-ribose (cADPR), the NAD(+) metabolite, mobilizes calcium from calcium stores in many cells. We found that dinoflagellate cells with higher metabolic rate underwent multiple fission (MF), a division mode in which cells can exceed twice their sizes at G1. A temperature shift-down experiment suggested that MF involves a commitment point at late G1. In fast-growing cells, cADPR level peaked in G(1) and increased with increasing concentrations of glucose in the medium. Addition of glycolytic poison iodoacetate inhibited cell growth, reduced cADPR levels as well as the commitment of cell cycles in fast-growing cells. Commitment of MF cell cycles was induced by a cell permeant cADPR agonist, but blocked by a specific antagonist of cADPR-induced Ca(2+) release. Our results establish cADPR as a link between cellular metabolism and cell cycle control.

Anxiety and Depression with Neurogenesis Defects in Exchange Protein Directly Activated by cAMP 2-Deficient Mice are Ameliorated by a Selective Serotonin Reuptake Inhibitor, Prozac

Intracellular cAMP and serotonin are important modulators of anxiety and depression. Fluoxetine, a selective serotonin reuptake inhibitor (SSRI) also known as Prozac, is widely used against depression, potentially by activating cAMP response element-binding protein (CREB) and increasing brain-derived neurotrophic factor (BDNF) through protein kinase A (PKA). However, the role of Epac1 and Epac2 (Rap guanine nucleotide exchange factors, RAPGEF3 and RAPGEF4, respectively) as potential downstream targets of SSRI/cAMP in mood regulations is not yet clear. Here, we investigated the phenotypes of Epac1 (Epac1−/−) or Epac2 (Epac2−/−) knockout mice by comparing them with their wild-type counterparts. Surprisingly, Epac2−/− mice exhibited a wide range of mood disorders, including anxiety and depression with learning and memory deficits in contextual and cued fear-conditioning tests without affecting Epac1 expression or PKA activity. Interestingly, rs17746510, one of the three single-nucleotide polymorphisms (SNPs) in RAPGEF4 associated with cognitive decline in Chinese Alzheimers disease (AD) patients, was significantly correlated with apathy and mood disturbance, whereas no significant association was observed between RAPGEF3 SNPs and the risk of AD or neuropsychiatric inventory scores. To further determine the detailed role of Epac2 in SSRI/serotonin/cAMP-involved mood disorders, we treated Epac2−/− mice with a SSRI, Prozac. The alteration in open field behavior and impaired hippocampal cell proliferation in Epac2−/− mice were alleviated by Prozac. Taken together, Epac2 gene polymorphism is a putative risk factor for mood disorders in AD patients in part by affecting the hippocampal neurogenesis.

Flow cytometric analysis of nocodazole-induced cell-cycle arrest in the pennate diatom Phaeodactylum tricornutum Bohlin

The microtubule inhibitor, nocodazole (2.5 mg L-1), can arrest the cell cycle of the pennate diatom Phaeodactylum tricornutum Bohlin at G2 + M phase. Flow cytometric analysis of cells treated with nocodazole suggest that the proportion of cells at G2 + M phase can accumulate to over 95%. Even after a 48-h treatment with nocodazole (2.5 mg L-1), the cells can still exit mitosis, suggesting that the cell-cycle arrest is reversible.

Multiplex fluorimetric assays for monitoring algal toxins

Most known algal toxins act on ion channels either directly or indirectly, resulting in a change in intracellular ion concentrations when administered to targeted cells. The present project developed the working conditions for the use of fluorescent dyes in monitoring changes in membrane potential, intracellular calcium, and intracellular sodium levels in mammalian cell lines. Using these conditions, we were able to demonstrate specific changes in fluorescent signals in response to several purified toxins. We were also able to generate algal extracts which, when administered to the developed fluorimetric assays, were able to elicit different pattern of changes in membrane potential, intracellular calcium, and intracellular sodium levels. The differential pattern of responses induced by the different algal toxins in the three fluorimetric assays serve as a proof of concept for the use of multiplex fluorimetric assays in the laboratory monitoring of algal toxins.