The roles of Ca2+ and plasma membrane ion channels in hyphal tip growth of Neurospora crassa

Abstract

Growing hyphae of the ascomycete fungus Neurospora crassa contained a tip-high gradient of cytoplasmic Ca2+, which was absent in non-growing hyphae and was insensitive to Gd3+ in the medium. Patch clamp recordings in the cell-attached mode, from the plasma membrane of these hyphae, showed two types of channel activities; spontaneous and stretch activated. The spontaneous channels were identified as inward K+ channels based on inhibition by tetraethylammonium. The stretch activated channels had increased amplitudes in response to elevated Ca2+ in the pipette solution, and thus are permeable to Ca2+ and mediate inward Ca2+ movement. Gd3+, which is an inhibitor of some stretch activated channels, incompletely inhibited stretch activated channel activity. Both tetraethylammonium and Gd3+ only transiently reduced the rates of tip growth without changing tip morphology, thus indicating that the channels are not absolutely essential for tip growth. Furthermore, in contrast to the hyphae of another tip growing organism, Saprolegnia ferax, tip-high gradients of neither spontaneous nor stretch activated channels were found. Voltage clamping of the apical plasma membrane potential in the range from -300 to +150 mV did not affect the rates of hyphal elongation. Collectively, these data suggest that ion transport across the plasma membrane at the growing tip in Neurospora is not obligatory for the maintenance of tip growth, but that a gradient of Ca2+, possibly generated from internal stores in an unknown way, is required.