Oxygen fluxes were mapped at the growing apices and along mycelial hyphal segments of the ascomycete Neurospora crassa. High spatial resolution was obtained using micro-oxygen probes (2-3 microm tip diameters) and the self-referencing technique to maximize the sensitivity of oxygen flux measurements. As expected, oxygen influx was inhibited by cyanide, although oxygen influx (and hyphal growth) resumed with the induction of an alternate oxidase activity. Along hyphal segments, variations in oxygen influx were not correlated with location, near or far from septa, and varied over time along the same hyphal segment. Growing hyphae had a region of maximal oxygen influx greater than 10 microm behind the hyphal tip, the oxygen influx was correlated with hyphal growth rate. The region of maximal oxygen influx did not correspond with mitochondrial density, which is maximal (about 30% of hyphal volume) 5-10 microm behind the tip. Therefore, tip-localized mitochondria do not contribute to the respiratory requirements of the growing hypha. The tip-localized mitochondria may function in clearing calcium from the cytoplasm, although a decline in chlortetracycline fluorescence after cyanide inhibition could also be due to ATP-depletion due to inhibition of actively respiring mitochondria. Alternatively, the growing tip may be the site of mitochondrial biogenesis.
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