Mutants in the arom gene cluster of Neurospora crassa specific for biosynthetic dehydroquinase.

I N Neurospora crassa a gene cluster encodes a multienzyme aggregate of five enzymes catalyzing steps two through six in the pre-chorismic acid part (Figure 1) of the aromatic synthetic pathway (GILES, CASE, PARTRIDGE, and AHMED 1967a). Two distinct categories of mutants have been detected within this arom cluster: single gene mutants which lack only one of the activities and pleiotropic (“polarity”) mutants which lack two or more activities (Figure 2). A major class of pleiotropic mutants consists of non-complementing types which lack all five activities, map within the proximal (arom-2-DHQ synthetase) structural gene in the cluster, and are (in at least some cases) suppressed by nonsense suppressors (CASE and GILES 1968). These mutants provide the major evidence that the arom cluster is normally transcribed in a polarized fashion as a single m-RNA which is subsequently translated also in a polarized fashion into the multienzyme aggregate. Previous studies failed to yield mutants lacking only dehydroquinase (DHQase), despite the fact that the absence of this activity in many non-complementing mutants indicated that the arom cluster encodes the DHQase. Subsequently, evidence was obtained that Neurospora has two different DHQases (GILES, PARTRIDGE, AHMED, and CASE 1967b). One of these is a catabolic enzyme induced by quinic acid (QA) or dehydroquinic acid (DHQ). The other, which is constitutive and associated with the arom aggregate, is the biosynthetic enzyme. The hypothesis was proposed that single gene mutants in the arom cluster lacking the synthetic activity were not recovered because such mutants would accumulate DHQ, induce the catabolic enzyme, and thus exhibit no aromatic requirement. Proof of this hypothesis required the development of a procedure for isolating mutants in this presumptive structural gene in the arom cluster encoding the synthetic DHQase. The present paper describes such a procedure and discusses the genetical and biochemical properties of mutants which have been obtained in the predicted category.