Sp110, a polypeptide component of the nuclear body

Abstract

Cloning and characterization of a full length cDNA encoding Sp110 (speckled 110), a novel 110 kDa polypeptide, is disclosed. It is disclosed that Sp110 is a component of the nuclear body, is expressed in leukocytes, and is also expressed in other types of cells, including endothelial cells, smooth muscle cells, liver cells and heart cells, after contact with certain cytokines. The disclosure also includes the following: Sp140 recruits Sp110 to the nuclear body, Sp110 functions as an activator of gene transcription, and Sp110 serves as a nuclear hormone receptor co-activator. Sp110 DNAs, polypeptides, antibodies are disclosed. Also disclosed are Sp110-related screening methods and clinical diagnostic methods.

1 . An isolated DNA comprising a nucleotide sequence whose complement hybridizes under stringent hybridization conditions to a DNA molecule whose nucleotide sequence consists of nucleotides 405 to 797 of SEQ ID NO:1.  
     
     
         2 . The DNA of  claim 1 , wherein the isolated DNA further comprises a nucleotide sequence encoding a domain selected from the group consisting of: 
 a domain having at least 80% sequence identity with amino acids 6-109 of SEQ ID NO:2 (Sp100-like domain);    a domain having at least 80% sequence identity with amino acids 454-532 of SEQ ID NO:2 (SAND domain);    a domain having at least 80% sequence identity with amino acids 537-577 of SEQ ID NO:2 (plant homeobox domain); and    a domain having at least 80% sequence identity with amino acids 606-674 of SEQ ID NO:2 (bromodomain).    
     
     
         3 . The DNA of  claim 1 , wherein the isolated DNA further comprises a nucleotide sequence encoding a domain selected from the group consisting of: 
 amino acids 6-109 of SEQ ID NO:2 (Sp100-like domain) or amino acids 6-109 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein;    amino acids 454-532 of SEQ ID NO:2 (SAND domain) or amino acids 454-532 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein;    amino acids 537-577 of SEQ ID NO:2 (plant homeobox domain) or amino acids 537-577 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein; and    amino acids 606-674 of SEQ ID NO:2 (bromodomain) or amino acids 606-674 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein.    
     
     
         4 . The DNA of  claim 1 , wherein the isolated DNA further comprises a nucleotide sequence encoding: 
 amino acids 6-109 of SEQ ID NO:2 (Sp100-like domain) or amino acids 6-109 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein;    amino acids 454-532 of SEQ ID NO:2 (SAND domain) or amino acids 454-532 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein;    amino acids 537-577 of SEQ ID NO:2 (plant homeobox domain) or amino acids 537-577 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein; and    amino acids 606-674 of SEQ ID NO:2 (bromodomain) or amino acids 606-674 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein.    
     
     
         5 . An isolated DNA comprising a nucleotide sequence that encodes a polypeptide whose amino acid sequence the sequence set forth as SEQ ID NO:2 or the sequence set forth as SEQ ID NO:2, with one or more conservative amino acid substitutions therein.  
     
     
         6 . The DNA of  claim 1 , wherein the isolated DNA comprises a nucleotide sequence encoding a polypeptide comprising: 
 amino acids 6-109 of SEQ ID NO:2 (Sp100-like domain) or amino acids 6-109 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein;    amino acids 537-577 of SEQ ID NO:2 (plant homeobox domain) or amino acids 537-577 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein; and    amino acids 606-674 of SEQ ID NO:2 (bromodomain) or amino acids 606-674 of SEQ ID NO:2 with one or more conservative amino acid substitutions therein;    wherein the polypeptide does not contain a SAND domain.    
     
     
         7 . An isolated DNA comprising a nucleotide sequence that encodes the amino acid sequence set forth as SEQ ID NO:5, or the sequence set forth as SEQ ID NO:5 with one or more conservative amino acid substitutions therein.  
     
     
         8 . A vector comprising the DNA of  claim 1 .  
     
     
         9 . The vector of  claim 8 , wherein the DNA is operably linked to an expression control sequence.  
     
     
         10 . A cell comprising the vector of  claim 8 .  
     
     
         11 - 14 . (canceled)  
     
     
         15 . An antibody that binds specifically to an Sp110 polypeptide.  
     
     
         16 . The antibody of  claim 15 , further comprising a detectable label.  
     
     
         17 . A screening method for identifying a compound that inhibits Sp110 dimerization, the method comprising: 
 providing an Sp110 polypeptide sample solution;    adding to the sample solution a candidate compound; and    detecting a decrease in Sp110 dimerization in the presence of the candidate compound, as compared to Sp110 dimerization in the absence of the candidate compound, as an indication that the candidate compound inhibits Sp110 dimerization.    
     
     
         18 . A screening method for identifying a compound that enhances Sp110 dimerization, the method comprising: 
 providing an Sp110 polypeptide sample solution;    adding to the sample solution a candidate compound; and    detecting an increase in Sp110 dimerization in the presence of the candidate compound, as compared to Sp110 dimerization in the absence of the candidate compound, as an indication that the candidate compound enhances Sp110 dimerization.    
     
     
         19 . A screening method for identifying a polypeptide that dimerizes with Sp110 to form an inactive heterodimer, the method comprising: 
 providing an Sp110 polypeptide sample solution;    adding to the sample solution a candidate polypeptide, thereby forming a test mixture;    providing a gene expression system comprising a reporter gene operably linked to an Sp110-responsive expression control sequence;    contacting the test mixture with the gene expression system;    detecting a decrease in reporter gene expression level in the presence of the test mixture, as compared to gene expression level in the presence of the Sp110 polypeptide sample solution, as an indication that the candidate compound dimerizes with Sp110 to form an inactive heterodimer.    
     
     
         20 . A screening method for identifying a polypeptide that dimerizes with Sp110 to form a constitutively active or hyperactive heterodimer, the method comprising: 
 providing an Sp110 polypeptide sample solution;    adding to the sample solution a candidate polypeptide, thereby forming a test mixture;    providing a gene expression system comprising a reporter gene operably linked to an Sp110-responsive expression control sequence;    contacting the test mixture with the gene expression system;    detecting constitutive expression or an increase in reporter gene expression level in the presence of the test mixture, as compared to gene expression level in the presence of the Sp110 polypeptide sample solution, as an indication that the candidate compound dimerizes with Sp110 to form a constitutively active or hyperactive heterodimer.    
     
     
         21 . A screening method for identifying a compound or polypeptide that inhibits Sp110 binding to a nuclear hormone receptor, the method comprising: 
 providing an Sp110 polypeptide sample solution;    adding to the sample solution a candidate compound;    adding to the sample solution a nuclear hormone receptor; and    detecting a decrease in Sp110 binding to the nuclear hormone receptor in the presence of the candidate compound, as compared to Sp110 binding to the nuclear hormone receptor in the absence of the candidate compound.    
     
     
         22 . A screening method for identifying a compound or polypeptide that enhances Sp110 binding to a nuclear hormone receptor, the method comprising: 
 providing an Sp110 polypeptide sample solution;    adding to the sample solution a candidate compound;    adding to the sample solution a nuclear hormone receptor; and    detecting an increase in Sp110 binding to the nuclear hormone receptor in the presence of the candidate compound, as compared to Sp110 binding to the nuclear hormone receptor in the absence of the candidate compound.    
     
     
         23 . A screening method for identifying a compound or polypeptide that inhibits the binding of an Sp110 dimer to an Sp110-binding nucleotide sequence, the method comprising: 
 providing an Sp110 polypeptide sample solution;    adding to the sample solution a candidate compound or polypeptide;    adding to the sample solution an Sp110-binding nucleotide sequence; and    detecting a decrease in Sp110 binding to the Sp110-binding nucleotide sequence in the presence of the candidate compound, as compared to Sp110 binding to the Sp110-binding nucleotide sequence in the absence of the candidate compound.    
     
     
         24 . A screening method for identifying a compound or polypeptide that enhances the binding of an Sp110 dimer to an Sp110-binding nucleotide sequence, the method comprising: 
 providing an Sp110 polypeptide sample solution;    adding to the sample solution a candidate compound or polypeptide;    adding to the sample solution an Sp110-binding nucleotide sequence; and    detecting an increase in Sp110 binding to the Sp110-binding nucleotide sequence in the presence of the candidate compound, as compared to Sp110 binding to the Sp110-binding nucleotide sequence in the absence of the candidate compound.    
     
     
         25 . A method for diagnosing primary biliary cirrhosis (PBC) in a human patient, the method comprising: 
 providing a substantially pure Sp110 polypeptide;    providing a serum sample from the patient;    contacting Sp110 polypeptide with the serum sample; and    detecting specific binding of an antibody in the serum with the Sp110 polypeptide as an indication of PBC.    
     
     
         26 . A method for assessing the prognosis of a patient diagnosed as having primary biliary cirrhosis (PBC) in a human patient, the method comprising: 
 providing a substantially pure Sp110 polypeptide;    providing a serum sample from the patient;    contacting Sp110 polypeptide with the serum sample; and    detecting specific binding of an antibody in the serum with the Sp110 polypeptide as an indication of the patient's prognosis.