The yeast Saccharomyces cerevisiae gene CCHl (ORF YGR217w) shows high homology with animal calcium channel al-subunit genes. Knock-out mutants were constructed of Cchl and of Midi which is known to mediate Ca 2+ influx in response to the a-mating pheromone. Cchl is involved in calcium influx and the late stage of the mating process. The cchl mutant sensitivity against the a-mating pheromone can be reduced by the addition of extra calcium. The product of this gene is likely to interact with the MIDI gene product in Ca influx or its control.

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Internal pH (pHi) was determined inEmiliania huxleyi (Lohmann) using the probe 2',7'-bis-(2-carboxyethyl)-5(and-6)carboxyfluoresceinacetoxymethylester (BCEF-AM) and digital imaging microscopy. The probe BCECF-AM was taken up and hydrolysed to the free acid by the cells. A linear relationship was established between pHi and the 490/450 fluorescence ratio of BCECF-AM over the pH range 6.0 to 8.0 using the ionophore nigericin. Two distinct pH domains were identified within the cell, the cytoplasmic domain (approx. pH 7.0) and the chloroplast domain (approx. pH 8.0). The average pHi was 7.29 (±0.11) for cells in the presence of 2 mM HCO 3 (-) . In the absence of HCO 3 (-) the pHi was decreased by 0.8 pH unit. The importance of these changes in pHi is considered in relation to inorganic-carbon uptake.

inorganic carbon-dependent photosynthetic oxygen evolution was saturated at a photon flux density of 100//mol m"^ s"^ for air-grown cells of a low calcifying strain of Emiliania huxleyi (Lohmann) Kamptner. Measurement of photosynthetic oxygen evolution at constant inorganic carbon concentration but varying pH showed that exogenous bicarbonate was not a major carbon source for photosynthesis. At pH 8-0 the concentration of dissolved inorganic carbon (DIC) required for the half-maximal rate of photosynthetic O^ evolution (K'^.j [DIC]) was 2'86 mM; the rate of non-enzymic dehydration of HCO^" greatly exceeding the rate of CO.^ fixation. Carbon dioxide uptake occurs by diffusive entr\' as shown by the A',,-[DIC] of 12-5 fiM at pH 5-0.Bicarhonate uptake, measured by the silicone-oil-layer centrigual filtering technique, did not show Michaeiis-Menten type kinetics. The electrical membrane potential difference was determined from the distribution of the lipophilic cation tetra[*^H]phenylphosphonium (TPP*) between cells and the media. Cells grown at pH 8-0 exhibited a negative membrane potential (inside of ce!! relative to outside) of about -60 mV.

summary We have used a pulsed u.v. laser to ablate the cell wall of the filamentous fungus Aspergillus niger van Tiegh., and expose the plasma membrane at defined points along the hypha. Using the patch clamp technique, giga ohm seals (up to 20 GΩ) and recordings of plasma membrane ion channels were obtained. We describe, for the first time in fungi, a 43 pS plasma membrane anion efflux channel. Its potential role in cell signalling and pH homeostasis is discussed.