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The levels of the endogenous amino acid pools in conidia, germinating conidia, and mycelia of wild-type Neurospora crassa were measured. Three different chromatographic procedures employing the amino acid analyzer were used to identify and quantitatively measure 28 different ninhydrin-positive compounds. All of the common amino acids were detected in conidial extracts except proline, methionine, and cystine. The levels of these three amino acid pools were also very low in mycelia. During the first hour of germination in minimal medium, the levels of most of the free amino acid pools decreased. The pool of glutamic acid, the predominant free amino acid in conidia, decreased 70% during the first hour. Very little glutamic acid or any other amino acid was excreted into the medium. During the first 20 min of germination, the decrease in the glutamic acid pool was nearly equivalent to the increase in the aspartic acid pool. The aspartic acid and lambda-aminobutyric acid pools were the only amino acid pools that increased to maximum levels within the first 20 min of germination and then decreased. It is proposed that an important metabolic event that occurs during the early stages of conidial germination is the production of reduced pyridine nucleotides. The degradation of the large glutamic acid pool existing in the conidia (2.5% of the conidial dry weight) could produce these reduced coenzymes.

The levels of glucosamine and galactosamine were determined in conidia, germinating conidia, and vegetative mycelia of Neurospora crassa. In the vegetative mycelia about 90% of the amino sugars were shown to be components of the cell wall. The remaining 10% of the amino sugars were tentatively identified as the nucleotide sugars uridine diphospho-2-acetamido-2-deoxy-D-glucose and uridine diphospho-2-acetamido-2-deoxy-D-galactose. Conidia and vegetative mycelia contained about the same levels of glucosamine. During the first 9 h after the initiation of germination, the total glucosamine content had increased 3.1-fold, whereas the residual dry weight of the culture had increased 7.7-fold. This led to a drop in the glucosamine concentration from 100 Amol/g of residual dry weight to 42 ,mol/g. During this time, all of the conidia had germinated and

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Neurospora crassa glutamate decarboxylase (GAD) is produced during conidiation and stored in dormant conidia. Polyclonal antibody was generated to GAD that had been purified to homogeneity. The anti-GAD antibody was specific for N. crassa GAD and inhibited GAD activity. The level of GAD protein decreased during conidial germination, indicating that GAD was degraded during this phase of development. The anti-GAD antibody was used to isolate a cDNA clone of GAD from a lambda ZAP cDNA expression library. Escherichia coli containing a plasmid with the cDNA insert produced GAD activity. The cDNA clone contained a 2.6 kbp insert and hybridized to a 2.6 kb mRNA species from conidiating cultures of N. crassa. GAD mRNA was not present in vegetative hyphae. In conidiating cultures, GAD mRNA was first detected when conidia began to appear. The level of GAD mRNA increased as conidiation progressed. This is the first example of the cloning of an enzyme that is regulated at the level of mRNA during the asexual developmental cycle of N. crassa.