Telomerase-associated protein 1, HSP90, and topoisomerase IIalpha associate directly with the BLM helicase in immortalized cells using ALT and modulate its helicase activity using telomeric DNA substrates

The BLM helicase associates with the telomere structural proteins TRF1 and TRF2 in immortalized cells using the alternative lengthening of telomere (ALT) pathways. This work focuses on identifying protein partners of BLM in cells using ALT. Mass spectrometry and immunoprecipitation techniques have i...

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Bibliographic Details
Published in:The Journal of biological chemistry 2009-05, Vol.284 (22), p.14966-14977
Main Authors: Bhattacharyya, Saumitri, Keirsey, Jeremy, Russell, Beatriz, Kavecansky, Juraj, Lillard-Wetherell, Kate, Tahmaseb, Kambiz, Turchi, John J, Groden, Joanna
Format: Article
Language:English
Subjects:
Antigens, Neoplasm - metabolism
Blotting, Western
Carrier Proteins - metabolism
Cell Line, Transformed
Cell Nucleus Structures - metabolism
DNA - biosynthesis
DNA - metabolism
DNA Topoisomerases, Type II - metabolism
DNA-Binding Proteins - metabolism
HSP90 Heat-Shock Proteins - metabolism
Humans
Mass Spectrometry
Protein Transport
RecQ Helicases - chemistry
RecQ Helicases - metabolism
RNA-Binding Proteins
Telomere - metabolism
Telomeric Repeat Binding Protein 2 - chemistry
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Summary:The BLM helicase associates with the telomere structural proteins TRF1 and TRF2 in immortalized cells using the alternative lengthening of telomere (ALT) pathways. This work focuses on identifying protein partners of BLM in cells using ALT. Mass spectrometry and immunoprecipitation techniques have identified three proteins that bind directly to BLM and TRF2 in ALT cells: telomerase-associated protein 1 (TEP1), heat shock protein 90 (HSP90), and topoisomerase IIalpha (TOPOIIalpha). BLM predominantly co-localizes with these proteins in foci actively synthesizing DNA during late S and G(2)/M phases of the cell cycle when ALT is thought to occur. Immunoprecipitation studies also indicate that only HSP90 and TOPOIIalpha are components of a specific complex containing BLM, TRF1, and TRF2 but that this complex does not include TEP1. TEP1, TOPOIIalpha, and HSP90 interact directly with BLM in vitro and modulate its helicase activity on telomere-like DNA substrates but not on non-telomeric substrates. Initial studies suggest that knockdown of BLM in ALT cells reduces average telomere length but does not do so in cells using telomerase.
ISSN:0021-9258
DOI:10.1074/jbc.M900195200