Most studies on the structure of DNA in telomeres have been dedicated to the double-stranded region or the guanosine-rich strand and consequently little is known about the factors that may bind to the telomere cytosine-rich (C-rich) strand. This led us to investigate whether proteins exist that can recognise C-rich sequences. We have isolated several nuclear factors from human cell extracts that specifically bind the C-rich strand of vertebrate telomeres [namely a d(CCCTAA)(n) repeat] with high affinity and bind double-stranded telomeric DNA with a 100x reduced affinity. A biochemical assay allowed us to characterise four proteins of apparent molecular weights 66-64, 45 and 35 kDa, respectively. To identify these polypeptides we screened a lambda gt11-based cDNA expression library, obtained from human HeLa cells using a radiolabelled telomeric oligonucleotide as a probe. Two clones were purified and sequenced: the first corresponded to the hnRNP K protein and the second to the ASF/SF2 splicing factor. Confirmation of the screening results was obtained with recombinant proteins, both of which bind to the human telomeric C-rich strand in vitro.
Identification of two human nuclear proteins that recognise the cytosine-rich strand of human telomeres in vitro
Lacroix, L.; Lienard, H.; Labourier, E.; Djavaheri-Mergny, M.; Lacoste, J.; Leffers, H.; Tazi, J.; Helene, C.; Mergny, J. L.
Nucleic Acids Research
2000-04-01 / vol 28 / pages 1564-1575
saccharomyces-cerevisiae; in-vitro; DNA-binding protein; c-myc; circular-dichroism; g-quartet formation; ribonucleoprotein-k; sf2/asf splicing factor; trypanosoma-brucei; yeast telomeres