Effects of the diatom-Emiliana huxleyi succession on photosynthesis, calcification and carbon metabolism by size-fractioned phytoplankton

Changes in the species composition, photosynthesis, calcification and size-fractionated carbon metabolism by natural phytoplankton assemblages were monitored in three mesocosms under different nutrient conditions during May 1993. In the 3 enclosures, the decline of the diatom-dominated assemblages was followed by the development of a bloom of the coccolithoporid Emiliania huxleyi. Highest growth of E. huxleyi was observed in the mesocosm with a high N : P ratio, suggesting this species is a good competitor at low phosphate concentrations. The transition from diatom- to E. huxleyi-dominated assemblages brought about a sharp reduction of the phytoplankton standing stock and carbon-specific photosynthetic rate. The relative contribution of the smaller size fraction to total photosynthesis increased as the succession progressed. Calcification rate and E. huxleyi cell-specified calcite production were highest during the early stages of development of the E. huxleyi bloom. Distinct changes in the patterns of ¹⁴C allocation into biomolecules were noticed during the diatom-E. huxleyi succession. The diatom-dominated assemblage showed high relative ¹⁴C incorporation into low molecular weight metabolites (LMWM), whereas proteins and, specially, lipids accounted for the largest proportion of carbon incorporation in the E. huxleyi bloom. The patterns of photoassimilated carbon metabolism proved to be strongly dependent on cellular size, as protein relative synthesis was significantly higher in the smaller than in the larger size fraction, irrespective of the nutrient regime and the successional stage. These results are discussed in relation to the ecological and physiological features of small phytoplankton.     

Patterns of carbon and nitrogen uptake during blooms of Emiliania huxleyi in two Norwegian fjords

Blooms of the coccolithophorid Emiliania huxleyi were monitoredin two land-locked fjords, Fauskangerpollen and Nordåsvannet(Western Norway), in May 1993. The chemical composition of particulatematter, size-fractionated photosynthesis, calcification, nitrogenuptake rates and the patterns of macromolecular synthesis wereexamined during the peak and decline of E.huxleyi blooms. Thetemporal evolution of the bloom in Fauskangerpollen was definedby a gradual decrease in cell abundance and cell-specific calcificationrates, and by increasing numbers of empty coccospheres and theratio detached coccoliths/living cells. A large proportion ofthe nitrogen required for microplankton growth was suppliedby aminonium and dissolved organic compounds such as urea and,as a consequence, the f-ratios were low (0.2). In general, nitrogenuptake patterns were consistent with ambient concentrationsof nitrogenous species. The photosynthetic carbon metabolismof E.huxleyi dominated phytoplankton assemblages was characterizedby high carbon allocation into lipids and relatively low carbonincorporation into protein as compared with diatom-dominatedassemblages. Consequently, the organic C/nitrogen uptake ratiodetermined stoichiometrically was significantly higher (up to10.8) when coccolithophorids were dominant than in diatom-basedor mixed-phyto-plankton assemblages. These carbon incorporationpatterns were reflected in differences in the chemical compositionof particulate matter.     

Carbohydrate and amino acid metabolism during batch culture of a human lymphoblastoid cell line,BTSN6

This work presents data on the carbohydrate and amino acid metabolism of a lymphoblastoid cell line producing an IgG1 antibody. In static culture, it was observed that lactate levels were significantly lowered when the cells were cultured on galactose as a carbon source. The use of carbohydrate substitution may be useful in lowering lactate levels, if it is established that this component is toxic to the cells. In addition, carbohydrate substitution may be used to modify glycosylation patterns and hence pharmacokinetic properties of glycoproteins.The amino acids glutamine and tryptophan were shown to be limiting in batch culture on this medium (DR, a 1:1 mixture of DMEM and RPMI, with 4mM glutamine). Amino acids produced included alanine, proline and glutamate. Serine was consumed to exhaustion, which was followed by a depletion of extracellular glycine. Amino acid metabolism, specific antibody productivity and specific growth rate were shown to be functions of the inoculation density in stirred flask culture. The results have implications for the design of media for both low and high density antibody manufacture by these cell lines.     

Comprehensive Mapping of Regional Expression of the Clock Protein PERIOD2 in Rat Forebrain across the 24-h Day

In mammals, a light-entrainable clock located in the suprachiasmatic nucleus (SCN) regulates circadian rhythms by synchronizing oscillators throughout the brain and body. Notably, the nature of the relation between the SCN clock and subordinate oscillators in the rest of the brain is not well defined. We performed a high temporal resolution analysis of the expression of the circadian clock protein PERIOD2 (PER2) in the rat forebrain to characterize the distribution, amplitude and phase of PER2 rhythms across different regions. Eighty-four LEW/Crl male rats were entrained to a 12-h: 12-h light/dark cycle, and subsequently perfused every 30 min across the 24-h day for a total of 48 time-points. PER2 expression was assessed with immunohistochemistry and analyzed using automated cell counts. We report the presence of PER2 expression in 20 forebrain areas important for a wide range of motivated and appetitive behaviors including the SCN, bed nucleus, and several regions of the amygdala, hippocampus, striatum, and cortex. Eighteen areas displayed significant PER2 rhythms, which peaked at different times of day. Our data demonstrate a previously uncharacterized regional distribution of rhythms of a clock protein expression in the brain that provides a sound basis for future studies of circadian clock function in animal models of disease.     

The AMP-dependent kinase pathway is upregulated in BAP1 mutant uveal melanoma

Metastatic uveal melanoma (UM) responds poorly to targeted therapies and immune checkpoint inhibitors. Loss of BRCA1-associated protein 1 (BAP1) via inactivating mutations in the BAP1 gene is associated with UM progression. Thus, molecular alterations caused by BAP1 dysfunction may be novel therapeutic targets for metastatic UM. Here, we found that phosphorylation of AMP-dependent kinase (AMPK) was elevated in BAP1-altered (or mutant) compared to BAP1-unaltered (or wild-type [WT]) UM tumors. As a readout of AMPK pathway activation, phosphorylation of an AMPK downstream effector, acetyl-CoA-carboxylase (ACC), was also elevated. BAP1 re-expression in BAP1-null UM cell lines decreased phospho-AMPK (pAMPK) and phospho-ACC (pACC) levels. AMPK phosphorylation is mediated by calcium/calmodulin dependent protein kinase kinase 2 (CaMKK2) and potentially liver kinase B1 (LKB1) in BAP1 mutant UM cells. Knockdown of AMPKα1/2 reduced the viability of BAP1 mutant UM cells, indicating a survival function of AMPK in BAP1 mutant UM. Our data suggest that the AMPK pathway is an important mechanism mediating the survival of BAP1 mutant UM. Targeting the AMPK pathway may be a novel therapeutic strategy for metastatic UM.