We describe a method to decipher the complex inter-relationships between metabolite production trends and gene expression events, and show how information gleaned from such studies can be applied to yield improved production strains. Genomic fragment microarrays were constructed for the Aspergillus terreus genome, and transcriptional profiles were generated from strains engineered to produce varying amounts of the medically significant natural product lovastatin. Metabolite detection methods were employed to quantify the polyketide-derived secondary metabolites lovastatin and (+)-geodin in broths from fermentations of the same strains. Association analysis of the resulting transcriptional and metabolic data sets provides mechanistic insight into the genetic and physiological control of lovastatin and (+)-geodin biosynthesis, and identifies novel components involved in the production of (+)-geodin, as well as other secondary metabolites. Furthermore, this analysis identifies specific tools, including promoters for reporter-based selection systems, that we employed to improve lovastatin production by A. terreus.

X-linked hypohidrotic ectodermal dysplasia results in abnormal morphogenesis of teeth, hair and eccrine sweat glands1. The gene (ED1) responsible for the disorder has been identified2,3,4, as well as the analogous X-linked gene (Ta) in the mouse5,6. Autosomal recessive disorders, phenotypically indistinguishable from the X-linked forms, exist in humans7 and at two separate loci (crinkled, cr, and downless, dl) in mice8. Dominant disorders, possibly allelic to the recessive loci, are seen in both species9,10 (ED3, Dlslk). A candidate gene has recently been identified at the dl locus11 that is mutated in both dl and Dlslk mutant alleles. We isolated and characterized its human DL homologue, and identified mutations in three families displaying recessive inheritance and two with dominant inheritance. The disorder does not map to the candidate gene locus in all autosomal recessive families, implying the existence of at least one additional human locus. The putative protein is predicted to have a single transmembrane domain, and shows similarity to two separate domains of the tumour necrosis factor receptor (TNFR) family12,13.

X-linked hypohidrotic ectodermal dysplasia (XLHED), the most common of the ectodermal dysplasias, results in the abnormal development of teeth, hair, and eccrine sweat glands. The gene responsible for this disorder, EDA1, was identified by isolation of a single cDNA that was predicted to encode a 135-amino-acid protein. Mutations in this splice form were detected in <10% of families with XLHED. The subsequent cloning of the murine homologue of the EDA1 gene (Tabby [Ta]) allowed us to identify a second putative isoform of the EDA1 protein (isoform II) in humans. This EDA1 cDNA is predicted to encode a 391-residue protein, of which 256 amino acids are encoded by the new exons. The putative protein is 94% identical to the Ta protein and includes a collagen-like domain with 19 repeats of a Gly-X-Y motif in the presumptive extracellular domain. The genomic structure of the EDA1 gene was established, and the complete sequence of the seven new exons was determined in 18 XLHED-affected males. Putative mutations, including 12 missense, one nonsense, and four deletion mutations, were identified in approximately 95% of the families. The results suggest that EDA1 isoform II plays a critical role in tooth, hair, and sweat gland morphogenesis, whereas the biological significance of isoform I remains unclear. Identification of mutations in nearly all of the XLHED families studied suggests that direct molecular diagnosis of the disorder is feasible. Direct diagnosis will allow carrier detection in families with a single affected male and will assist in distinguishing XLHED from the rarer, clinically indistinguishable, autosomal recessive form of the disorder.