Epidermolysis bullosa simplex is a disease in which keratin gene mutations cause the production of defective intermediate filaments, which leads in turn to epidermal basal cell fragility and blistering. The inheritance in nearly all kindreds is autosomal dominant, most kindreds have missense mutations, and the encoded proteins appear to exert a dominant negative function. One previously reported patient with generalized blistering had a fully dominant mutation of keratin 5; in that kindred a homozygote was affected no more severely than the heterozygotes. By contrast we report-here a keratin 14 mutation that causes blistering limited to the hands and feet in heterozygotes, but homozygotes have more severe, widespread blistering of the skin and mucous membranes. Thus keratin gene mutations may be not only fully recessive or fully dominant but also partially dominant as well.

DNA topoisomerase I (topo I) is known to participate in the process of DNA replication, but is not essential in Saccharomyces cerevisiae. The TRF4 gene is also nonessential and was identified in a screen for mutations that are inviable in combination with a top1 null mutation. Here we report the surprising finding that a top1 trf4-ts double mutant is defective in the mitotic events of chromosome condensation, spindle elongation, and nuclear segregation, but not in DNA replication. Direct examination of rDNA-containing mitotic chromosomes demonstrates that a top1 trf4-ts mutant fails both to establish and to maintain chromosome condensation in the rDNA at mitosis. We show that the Trf4p associates physically with both Smclp and Smc2p, the S. cerevisiae homologs of Xenopus proteins that are required for mitotic chromosome condensation in vitro. The defect in the top1 trf4-ts mutant is sensed by the MAD1-dependent spindle assembly checkpoint but not by the RAD9-dependent DNA damage checkpoint, further supporting the notion that chromosome structure influences spindle assembly. These data indicate that TOP1 (encoding topo I) and TRF4 participate in overlapping or dependent steps in mitotic chromosome condensation and serve to define a previously unrecognized biological function of topo I.

Mutations in genes encoding the keratin intermediate filaments expressed in basal cells have been identified in some families with epidermolysis bullosa simplex as the proximate cause of the fragility. We have systematically scanned genomic sequences of one of these keratins, keratin 14, for mutations in patients from 49 apparently independent kindreds using single-strand conformation polymorphism analysis. The ten mutations identified are clustered at three sites--the ends of the helices and the L12 linker region, mutation sites that have been identified in past, more limited studies. Early onset of blistering in these ten families is correlated with more widespread distribution of lesions.