Research Project
The Neurospora crassa catabolic enzyme, arginase,
exists in
multiple forms. Multiple forms of arginase are found in many vertebrates,
but this is the only reported example in a microbial organism. The two
major forms are structurally similar with subunit sizes of 36 and 41 kDa,
respectively. The larger form is produced by mycelia growing in arginine-supplemented
medium. Both forms are localized in the cytosol. The structural gene for
arginase, aga, has been cloned and sequenced; it contains a 358-codon open
reading frame with three in-frame ATGs at the amino terminus. Mutagenesis
of these ATGs revealed that the first ATG initiates the 41-kDa protein
and the third ATG initiates the 36-kDa protein. Mutation of the second
ATG has no effect on translation. Northern analysis demonstrated that a
1.4-kilobase (kb) transcript is synthesized in minimal medium and both
a 1.4- and 1.7-kb transcript are produced in arginine-supplemented medium.
Primer extension identified the 5' ends of each transcript and demonstrated
that the first and third ATG of the open reading frame are the initial
AUGs of the 1.7- and 1.4-kb mRNA, respectively. The results suggest that
a basal promoter produces the 1.4-kb transcript and an arginine "activated"
promoter is responsible for the 1.7-kb transcript. Tandem promoters are
rare in eukaryotic organisms, and they often regulate developmental or
tissue-specific gene expression. The possibility that arginase has a role
in differentiation in N. crassa is being investigated.