The autoantigen Ku is indistinguishable from NF IV, a protein forming multimeric protein-DNA complexes.

Stuiver, Maarten H.; Coenjaerts, Frank E. J.; Vliet, Peter C. van der

doi:10.1084/jem.172.4.1049

Cited by 50 publications

(8 citation statements)

References 14 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Third, on extraction of nuclei with 2 M NaCl, a large proportion of Ku protein remains insoluble (ref. 16; and data not shown). Finally, immunofluorescence studies show that Ku protein is localized in the nucleus and that its distribution is grossly altered by DNase I, but not by RNase treatment (3,4).…”

Section: Discussionmentioning

confidence: 79%

See 1 more Smart Citation

Analysis of the mechanism of interaction of simian Ku protein with DNA

1991

View full text Add to dashboard Cite

Ku protein is a relatively abundant DNA-binding protein which was first detected as the autoantigen in a patient with scleroderma-polymyositis overlap syndrome (hence the name 'Ku'). It is a heterodimer of two polypeptide chains of molecular weights 85,000 and 72,000, and it characteristically binds, in vitro, to the ends of DNA fragments, and translocates to form regular multimeric complexes, with one protein bound per 30 bp of DNA. We have studied the mechanism of interaction of Ku protein with DNA in vitro, using protein extracted from cultured monkey cells. We find that the precise structure of the DNA ends is not important for binding, as Ku protein can bind to hairpin loops and to mononucleosomes. Bound protein also does not require DNA ends for continued binding, since complexes formed with linear DNAs can be circularized by DNA ligase. Dissociation of the complex also appears to require DNA ends, since ligase closed circular complexes were found to be extremely stable even in the presence of 2 M NaCl. We also found that Ku molecules slide along DNA, with no preferential binding to specific sequences. Thus, Ku protein behaves like a bead threaded on a DNA string, a binding mechanism which allows us to make a new hypothesis concerning the function of this protein in the nucleus.

show abstract

Section: Discussionmentioning

confidence: 79%

“…The protein is a heterodimer composed of two polypeptide chains of molecular weights 85 000 and 72 000. The cDNAs coding for both protein subunits have been cloned and sequenced (13)(14)(15)(16)(17), and the deduced amino acid sequences suggest that the subunits might be held together by an element similar to a leucine zipper (18).…”

Section: Introductionmentioning

confidence: 99%

Analysis of the mechanism of interaction of simian Ku protein with DNA

1991

View full text Add to dashboard Cite

show abstract

“…NRE1 serves as a sequence-specific DNA binding site for Ku/ DNA-PK CS , and kinase activity correlates specifically with Kus binding to this element. Additionally, several heterodimeric transcription factors which bind to specific DNA sequences have been described, including proximal sequence element binding protein, transferrin receptor element binding protein, heat shock element binding factor, and nuclear factor IV, that are closely related to, or identical to Ku (Knuth et al 1990, Stuiver et al 1990, Roberts et al 1994, Kim et al 1995. Thus, specific DNA binding sites for Ku on gene promoters may play a more important role in localizing Ku-associated kinase activity than had previously been thought.…”

Section: Introductionmentioning

confidence: 95%

Association of the Ku autoantigen/DNA-dependent protein kinase holoenzyme and poly(ADP-ribose) polymerase with the DNA binding domain of progesterone receptors

Sartorius¹,

Takimoto²,

Richer³

et al. 2000

Journal of Molecular Endocrinology

View full text Add to dashboard Cite

Ligand-activated progesterone receptors (PR) bind to DNA at specific progesterone response elements by means of a DNA binding domain (DBD PR ) containing two highly conserved zinc fingers. DNA-bound PRs regulate transcription via interaction with other nuclear proteins and transcription factors. We have now identified four HeLa cell nuclear proteins that copurify with a glutathionine-S-transferase-human DBD PR fusion protein.Microsequence and immunoblot analyses identified one of these proteins as the 113 kDa poly(ADPribose) polymerase. The three other proteins were identified as subunits of the DNA-dependent protein kinase (DNA-PK) holoenzyme: its DNA binding regulatory heterodimers consisting of Ku70 and Ku86, and the 460 kDa catalytic subunit, DNA-PK CS . DNA-PK that was 'pulleddown' by DBD PR on the affinity resin was able to (1) autophosphorylate Ku70, Ku86, and DNA-PK CS , (2) transphosphorylate DBD PR , and (3) phosphorylate a DNA-PK-specific p53 peptide substrate. DNA-PK was also able to associate with the DBD of the yeast activator GAL4. However, neither a PR DBD mutant lacking a structured first zinc finger (DBD CYS ) nor the core DBD of the estrogen receptor (DBD ER ) copurified DNA-PK, suggesting the interaction is not non-specific for DBDs. Lastly, we found that DNA-PK copurified with full-length human PR transiently expressed in HeLa cells, suggesting that the human PR/ DNA-PK complex can assemble in vivo. These data show that DNA-PK and DBD PR interact, that DBD PR is a phosphorylation substrate of DNA-PK, and suggest a potential role for DNA-PK in PR-mediated transcription.

show abstract

“…There is abundant evidence that Ku is a DNA binding protein. It has been isolated as a nuclear factor (NFIV) from HeLa cells (6), and localized to the peripheral area of the nucleus and nucleolus often associated with active chromatin (7,8). In addition, the deduced amino acid sequence of the p70 subunit contains periodic leucine repeats with considerable homology to some well characterized transcription factors (9,10).…”

Section: Introductionmentioning

confidence: 99%

Production and characterization of recombinant human Ku antigen

Ono

1994

Nucl Acids Res

View full text Add to dashboard Cite

Ku is an ubiquitous nuclear heterodimeric protein consisting of p70 and p86 subunits that binds double-stranded DNA termini and associates with chromosomes in vivo. It was originally described as an autoantigen in patients with certain autoimmune diseases. The individual subunits of Ku have been difficult to isolate from human cells without denaturation and attempts to produce functional recombinant Ku have been largely unsuccessful. Here, we utilize two recombinant baculoviral vectors that carry p70 or p86 cDNA and express the Ku subunits individually as well as assemble them into the complete Ku heterodimer. In an electrophoretic mobility shift assay, recombinant Ku binds to linear double-stranded DNA but not to supercoiled, nicked circular, nor linear single-stranded DNA. Neither subunit binds DNA by itself indicating that heterodimerization is essential for function. We also describe a simple purification method for the isolation of highly purified recombinant Ku using a hexahistidine tag. The baculovirus expression system provides a stable and efficient source of not only the p70 and p86 subunits but also the functional Ku heterodimer.

show abstract

The autoantigen Ku is indistinguishable from NF IV, a protein forming multimeric protein-DNA complexes.

Cited by 50 publications

References 14 publications

Analysis of the mechanism of interaction of simian Ku protein with DNA

Analysis of the mechanism of interaction of simian Ku protein with DNA

Association of the Ku autoantigen/DNA-dependent protein kinase holoenzyme and poly(ADP-ribose) polymerase with the DNA binding domain of progesterone receptors

Production and characterization of recombinant human Ku antigen

Contact Info

Product

Resources

About