Dissecting the regulatory circuitry of a eukaryotic genome

Abstract

A method of identifying cellular regulatory circuits which employ at least one component of a subcomplex of regulatory proteins within the RNA II polymerase holoenzyme which behaves as a signal processor for gene-specific regulators (at least one component of a eukaryotic transcription initiation apparatus) and of determining the set of components of the apparatus which are responsible for regulation of each gene and the set of genes which are coordinately controlled by each transcription factor.

What is claimed is:  
     
         1 . A pair of isogenic eukaryotic cells comprising a test cell which contains a mutant component of the general transcription machinery and a control cell which is the wild-type isogenic counterpart of the test cell.  
     
     
         2 . The pair of isogenic eukaryotic cells of  claim 1  wherein the mutant component of the general transcription machinery is a mutant general transcription factor.  
     
     
         3 . The pair of isogenic eukaryotic cells of  claim 1  wherein the mutant component of the general transcription machinery is a temperature sensitive mutant.  
     
     
         4 . The pair of isogenic eukaryotic cells of  claim 1  wherein the mutant component of the general transcription machinery is a point mutant.  
     
     
         5 . The pair of isogenic eukaryotic cells of  claim 1  wherein the mutant component of the general transcription machinery is a deletion mutant.  
     
     
         6 . The pair of isogenic eukaryotic cells of  claim 1  wherein the mutant component of the general transcription machinery is a component of RNA polymerase II holoenzyme.  
     
     
         7 . The pair of isogenic eukaryotic cells of  claim 1  wherein the mutant component of the general transcription machinery is a necessary to reconstitute promoter-dependent transcription in vitro with core RNA polymerase II.  
     
     
         8 . A method of studying the effects of drugs on cells comprising the steps of: 
 (a) contacting a cell with a drug; and    (b) determining the effect of the drug on the cell by assessing expression of one or more of the genes which are determined to be members of the regulatory pathway of the general transcription factor according to  claim 1 .    
     
     
         9 . A method of identifying a cellular regulatory circuit which employs a component of a subcomplex of regulatory proteins within the RNA polymerase II holoenzyme, referred to as the transcription initiation apparatus, comprising: 
 (a) comparing genome expression signature during cellular responses to environmental or other stimuli with the genome expression signature produced by a defect in the transcription initiation apparatus; and    (b) determining differences between the two genome expression signatures and relating the differences to the defect in the transcription initiation apparatus, thereby identifying a component of the transcription initiation apparatus which is responsible for regulation of genes in the cells.    
     
     
         10 . The method of  claim 9  wherein the cellular regulatory circuit is a yeast cell regulatory circuit.