The modified base J is the target for a novel DNA-binding protein in kinetoplastid protozoans

Abstract

DNA from Kinetoplastida contains the unusual modified base β-D-glucosyl(hydroxymethyl))uracil, called J. Base J is found predominantly in repetitive DNA and correlates with epigenetic silencing of telomeric variant surface glycoprotein genes in Trypanosoma brucei. We have now identified a protein in nuclear extracts of bloodstream stage T. brucei that binds specifically to J-containing duplex DNA. J-specific DNA binding was also observed with extracts from the kinetoplastids Crithidia fasciculata and Leishmania tarentolae. We purified the 90 kDa C. fasciculata J-binding protein 50,000-fold and cloned the corresponding gene from C. fasciculata, T. brucei and L. tarentolae. Recombinant proteins expressed in Escherichia coli demonstrated J-specific DNA binding. The J-binding proteins show 43-63% identity and are unlike any known protein. The discovery of a J-binding protein suggests that J, like methylated cytosine in higher eukaryotes, functions via a protein intermediate.