A negative regulator of telomere-length protein trf1 is associated with interstitial (TTAGGG)n blocks in immortal Chinese hamster ovary cells.
Telomeres of mammalian chromosomes are composed of long tandem repeats (TTAGGG)n which bind in a sequence-specific manner two proteins-TRF1 and TRF2. In human somatic cells both proteins are mostly associated with telomeres and TRF1 overexpression resulting in telomere shortening. However, chromosomes of some mammalian species, e.g., Chinese hamster, have large interstitial blocks of (TTAGGG)n sequence (IBTs) and the blocks are involved in radiation-induced chromosome instability. In normal somatic cells of these species chromosomes are stable, indicating that the IBTs are protected from unequal homologous recombination. In this study we expressed V5-epitope or green fluorescent protein (GFP)-tagged human TRF1 in different lines of mammalian cells and analyzed distribution of the fusion proteins in interphase nucleus. As expected, transient transfection of human (A549) or African green monkey cells with GFP-N-TRF1 or TRF1-C-V5 plasmids resulted in the appearance in interphase nuclei of multiple faint nuclear dots containing GFP or V5 epitope which we believe to represent telomeres. Transfection of immortalized Chinese hamster ovary (CHO) cell line K1 which have extremely short telomeres with GFP-N-TRF1 plasmid leads to the appearance in interphase nuclei of large GFP bodies corresponding in number to the number of IBTs in these cells. Simultaneous visualization of GFP and IBTs in interphase nuclei of transfected CHO-K1 cells showed colocalization of both signals indicating that expressed TRF1 actually associates with IBTs. These results suggest that TRF1 may serve as general sensor of (TTAGGG)n repeats controlling not only telomeres but also interstitial (TTAGGG)n sequences.
Krutilina
R I
RI
Institute of Cytology, Russian Academy of Sciences, Tikchoretskii Av. 4, St. Petersburg, 194064, Russian Federation.
Oei
S
S
Buchlow
G
G
Yau
P M
PM
Zalensky
A O
AO
Zalenskaya
I A
IA
Bradbury
E M
EM
Tomilin
N V
NV
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007399:Interphase; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics*; D016615:Telomere / Q000235:genetics* / Q000378:metabolism*; D035321:Telomeric Repeat Binding Protein 1; D014162:Transfection; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D035321:Telomeric Repeat Binding Protein 1