Histone Demethylation Mediated by the Nuclear Amine Oxidase Homolog LSD1

Abstract

LSD1, a homolog of nuclear amine oxidases, functions as a histone demethylase and transcriptional co-repressor. LSD1 specifically demethylates histone H3 lysine 4, which is linked to active transcription. Lysine demethylation occurs via an oxidation reaction that generates formaldehyde. Importantly, RNAi inhibition of LSD1 causes an increase in H3 lysine 4 methylation and concomitant de-repression of target genes, suggesting that LSD1 represses transcription via histone demethylation. The results thus identify a histone demethylase conserved from S. pombe to human and reveal dynamic regulation of histone methylation by both histone methylases and demethylases.

We claim: 
     
         1 . A method of monitoring eukaryotic histone demethylase activity, comprising:
 contacting a eukaryotic histone demethylase protein with a histone peptide, wherein the histone peptide is lysine- or arginine-methylated;   determining the methylation status of the histone peptide.   
     
     
         2 . The method of  claim 1  wherein the eukaryotic histone demethylase protein is LSD1 or AOF1. 
     
     
         3 . A method of screening for modulators of eukaryotic histone demethylase activity, comprising:
 contacting a eukaryotic histone demethylase protein and a histone peptide in the presence and in the absence of a test substance, wherein the histone peptide is lysine-methylated;   determining the methylation status of the histone peptide;   identifying a test substance as an enhancer of eukaryotic histone demethylase activity if less methylated lysine is found in the presence than in the absence of the test substance, and identifying a test substance as an inhibitor of eukaryotic histone demethylase protein activity if more methylated lysine is found in the presence than in the absence of the test substance.   
     
     
         4 . A method for identifying an agent that modulates the interaction between a histone demethylase protein and a CoREST or BHC80 protein, comprising contacting a histone demethylase reagent and a CoREST or BHC80 reagent in the presence of a test agent; and (ii) determining the level of interaction between the histone demethylase reagent and the CoREST or BHC80 reagent, wherein a different level of interaction between the histone demethylase reagent and the CoREST or BHC80 reagent in the presence of the test agent relative to the absence of the test agent indicates that the test agent is an agent that modulates the interaction between a histone demethylase protein and a CoREST or BHC80 protein. 
     
     
         5 . A method for identifying an agent that modulates the biological activity of a histone demethylase, comprising: (i) contacting a histone demethylase reagent with a CoREST or BHC80 reagent in the presence of a test agent; and (ii) determining the biological activity of the histone demethylase reagent, wherein a different activity of the histone demethylase reagent in the presence of the test agent relative to the absence of the test agent indicates that the test agent is an agent that modulates the biological activity of a histone demethylase. 
     
     
         6 . A composition or molecular complex comprising a histone demethylase reagent and a CoREST reagent and/or a BHC80 reagent. 
     
     
         7 . A method for repressing the transcription of a methylated histone activated gene in a cell, comprising contacting the cell with, or administering into the cell, an agent that increases the protein or activity level of a histone demethylase in the cell. 
     
     
         8 . The method of  claim 7 , wherein the agent is a histone demethylase protein or functional homolog thereof; a nucleic acid encoding a histone demethylase protein or functional homolog thereof; an agent that increases the level of protein or activity of CoREST; a CoREST protein or functional homolog thereof; an agent that stimulates the interaction between a histone demethylase and CoREST; or an agent that decreases the level of protein or activity of BHC80. 
     
     
         9 . The method of  claim 8 , wherein the agent is a nucleic acid encoding a CoREST protein or functional homolog thereof; a BHC80 siRNA; or an agent that inhibits the interaction between a histone demethylase and BHC80. 
     
     
         10 . The method of  claim 8 , further comprising contacting the cell with, or administering into the cell, a second agent that increases the level or protein or activity of a histone deacetylase (HDAC). 
     
     
         11 . The method of  claim 10 , wherein the second agent is an HDAC protein or a functional homolog thereof or a nucleic acid encoding an HDAC protein or functional homolog thereof. 
     
     
         12 . A method for increasing the transcription of a methylated histone activated gene in a cell, comprising contacting the cell with an agent that decreases the protein or activity level of a histone demethylase in the cell. 
     
     
         13 . The method of  claim 12 , wherein the agent is a histone demethylase siRNA; an agent that decreases the protein or activity level of CoREST in the cell; an agent that inhibits the interaction between a histone demethylase and CoREST; or an agent that increases the level of protein or activity of BHC80. 
     
     
         14 . The method of  claim 13 , wherein the agent is a CoREST siRNA; a BHC80 protein or functional homolog thereof; a nucleic acid encoding a BHC80 protein or functional homolog thereof; or an agent that stimulates the interaction between a histone demethylase and BHC80. 
     
     
         15 . The method of  claim 14 , further comprising contacting the cell with, or administering into the cell, a second agent that decreases the level or activity of an HDAC. 
     
     
         16 . A method for treating or preventing a disease associated with the abnormal expression of a methylated histone activated gene in a subject, comprising administering to the subject a therapeutically effective amount of an agent that modulates the level of protein or activity of a histone demethylase. 
     
     
         17 . The method of  claim 16 , wherein the disease is a hyperproliferative disease and the agent is an agent that increases the level of protein or activity of a histone demethylase. 
     
     
         18 . The method of  claim 17 , wherein the disease is cancer. 
     
     
         19 . The method of  claim 16 , further comprising administering to the subject a therapeutically effective amount of a second agent that increases the level of protein or activity of a histone deacetylase. 
     
     
         20 . The method of  claim 16 , wherein the agent is administered to the subject by intratumoral injection, perfusion of a target tissue through its vasculature or by direct injection to a target tissue.