A rapid system for analyzing histone H4 mutations in the yeast Saccharomyces cerevisiae

Genetically engineered, specific histone H4 mutations have been analyzed in yeast which lack normal H4 protein using the time consuming techniqUe of tetrad dissection. The ~urpose of the present research proj ect has been to develop a rapid new system for analyzing histone H4 mutations in yeast. The new system has made it possible to isolate quickly cells which are surviving on mutant H4 only. The system will make it possible to analyze more quickly large numbers of H4 mutations. It is termed a plasmid shuttle system because of the use of plasmid vectors for shuttling histone H4 alleles in and out of the yeast cell. The system relies on the availibility of haploid yeast cells which have been deleted for both copies of their histone H4 gene and which are dependent on plasmid encoded wild type H4. The system's effectiveness has been tested by analyzingH4 mutations which have been previously examined by another more time consuming method. These mutations have been defined as encoding functional or nonfunctional H4. An H4 mutation never examined has also been analyzed using the new system. The mutation consists of 6 amino acid SUbstitutions: arginine' is replaced by lysine at position 3, glycine by aspartic acid at position 6 and 42, glycine by serine at position 7 and 41, and alanine by threonine at position 38. statistical analysis of the data provides evidence that this mutation in the H4 gene does not encode functional H4 protein. A Rapid System For Analyzing Histone H4 Mutations In The Yeast Saccharomyces Cerevisiae By E. Rex L. Harkrader