Modulation of telomerase activity by telomere DNA-binding proteins in Oxytricha.
Telomere proteins protect the chromosomal terminus from nucleolytic degradation and end-to-end fusion, and they may contribute to telomere length control and the regulation of telomerase. The current studies investigate the effect of Oxytricha single-stranded telomere DNA-binding protein subunits alpha and beta on telomerase elongation of telomeric DNA. A native agarose gel system was used to evaluate telomere DNA-binding protein complex composition, and the ability of telomerase to use these complexes as substrates was characterized. Efficient elongation occurred in the presence of the alpha subunit. Moreover, the alpha-DNA cross-linked complex was a substrate for telomerase. At higher alpha concentrations, two alpha subunits bound to the 16-nucleotide single-stranded DNA substrate and rendered it inaccessible to telomerase. The formation of this alpha . DNA . alpha complex may contribute to regulation of telomere length. The alpha . beta . DNA ternary complex was not a substrate for telomerase. Even when telomerase was prebound to telomeric DNA, the addition of alpha and beta inhibited elongation, suggesting that these telomere protein subunits have a greater affinity for the DNA and are able to displace telomerase. In addition, the ternary complex was not a substrate for terminal deoxynucleotidyltransferase. We conclude that the telomere protein inhibits telomerase by rendering the telomeric DNA inaccessible, thereby helping to maintain telomere length.
Froelich-Ammon
S J
SJ
Department of Chemistry and Biochemistry and Howard Hughes Medical Institute, University of Colorado, Boulder, Colorado 80309-0215, USA. thomas.cech@colorado.edu
Dickinson
B A
BA
Bevilacqua
J M
JM
Schultz
S C
SC
Cech
T R
TR
D000818:Animals; D004253:DNA Nucleotidylexotransferase / Q000378:metabolism; D016054:DNA, Protozoan / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004587:Electrophoresis, Agar Gel; D046911:Macromolecular Substances; D008957:Models, Genetic; D016813:Oxytricha / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D015800:Protozoan Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012091:Repetitive Sequences, Nucleic Acid; D013045:Species Specificity; D019098:Telomerase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016615:Telomere / Q000378:metabolism*
D016054:DNA, Protozoan; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D015800:Protozoan Proteins; D004253:DNA Nucleotidylexotransferase; D019098:Telomerase; D004259:DNA-Directed DNA Polymerase