Involvement of autophagy and apoptosis and lipid accumulation in sclerotial morphogenesis of Morchella importuna

Abstract

Sclerotial formation is a key phase of the morel life cycle and lipids have been recorded as the main cytoplasmic reserves in sclerotia of Morchella fungi without any experimental verification. In this study, the ultrastructural features of the undifferentiated mycelia stage (MS) and three main sclerotial differentiation states (sclerotial initial [SI], sclerotial development [SD] and sclerotial maturation [SM]) were compared by transmission electron microscopy. The nature of the energy-rich substance in hypha and sclerotium of Morchella importuna was qualitatively investigated by confocal laser scanning microscopy and quantitatively studied by extraction of lipids. Sclerotia were observed to form from the repeated branching and enlargement of either terminal hyphae or subordinate hyphal branches, indicating a complex type of sclerotial development. Autophagy and apoptosis were involved in the sclerotial metamorphosis of the cultivated strain of M. importuna. During the SI phase, the characteristic features of autophagy (vacuolation, coalescence of small vacuoles, existence of autophagosomes and engulfment of autophagosomes by vacuoles) were observed. At the SD phase, apoptotic characteristics (condensation of the cytoplasm and nucleus, shrinkage of plasma membrane, extensive plasma membrane blebbing and existence of phagosomes) could be seen in some developing sclerotial cells. In the final stage of sclerotial morphogensis, the sclerotial cells showed a necrotic mode of cell death. In addition, confocal laser imaging studies of live cells indicated that the energy-rich substance in morel hyphae and sclerotia was lipid. The lipid content in sclerotia was significantly more than that in hyphal cells. To the best of our knowledge, this is the first detailed ultrastructural description highlighting the involvement of autophagy and apoptosis in sclerotial metamorphosis of Morchella species and lipid accumulation during morel sclerotial development was also first experimentally verified. This work will promote a better understanding of the mechanism of morel sclerotial metamorphosis.

Keywords: Confocal laser scanning microscopy; Metamorphosis; Morel; Programmed cell death; Transmission electron microscopy.