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Abstract
Cyclic AMP (cAMP)- and Ca2+-dependent protein kinase activities of the fungus Metarhizium anisopliae were sought in extracts of ungerminated conidia, germinating conidia and mycelium, as well as in purified plasma membranes from mycelium. Ungerminated conidia contained a Ca2+/calmodulin-dependent protein kinase capable of phosphorylating multiple endogenous proteins and involved in triggering germination. cAMP-dependent protein kinase activity was not detected in ungerminated conidia in spite of the presence of cAMP in these conidia and the pre-germination synthesis of two cAMP-binding proteins. Most phosphorylation events in crude mycelial extracts were Ca2+-dependent but H-series inhibitors of cAMP-dependent kinases selectively repressed phosphorylation of a 27 kDA protein. Plasma membranes from mycelium contained a Ca2+-independent but H-8-sensitive protein kinase with multiple endogenous substrates for phosphorylation. 8-Azido[32P]cAMP bound selectively to a 52 kDa membrane protein indicative of a single cAMP-binding protein. Plasma membranes contained a phosphatase which rapidly (< 1 min) and selectively dephosphorylated a polypeptide of 15·5 kDa, thus being suited to cause rapid and reversible changes in membrane function. Membranes also contained an adenylate cyclase apparently involved in transmembrane signalling reactions, since mechanical or chemical treatments which stress the fungus caused rapid increases in intracellular levels of cAMP. Reconstitution experiments with a homogenate from a crisp-1 mutant of Neurospora crassa suggested G-protein regulation of Metarhizium plasmalemma adenylate cyclase.
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