scholarly journals Distinct Functions of POT1 at Telomeres

2008 ◽  
Vol 28 (17) ◽  
pp. 5251-5264 ◽  
Author(s):  
Katharine S. Barrientos ◽  
Megan F. Kendellen ◽  
Brian D. Freibaum ◽  
Blaine N. Armbruster ◽  
Katherine T. Etheridge ◽  
...  
Keyword(s):  
Dna Damage ◽  
Protein Interaction ◽  
Dna Binding Protein ◽  
Binding Activity ◽  
Terminal Protein ◽  
Interaction Domain ◽  
Ssdna Binding ◽  
Double Stranded Dna ◽  
Mammalian Protein

ABSTRACT The mammalian protein POT1 binds to telomeric single-stranded DNA (ssDNA), protecting chromosome ends from being detected as sites of DNA damage. POT1 is composed of an N-terminal ssDNA-binding domain and a C-terminal protein interaction domain. With regard to the latter, POT1 heterodimerizes with the protein TPP1 to foster binding to telomeric ssDNA in vitro and binds the telomeric double-stranded-DNA-binding protein TRF2. We sought to determine which of these functions—ssDNA, TPP1, or TRF2 binding—was required to protect chromosome ends from being detected as DNA damage. Using separation-of-function POT1 mutants deficient in one of these three activities, we found that binding to TRF2 is dispensable for protecting telomeres but fosters robust loading of POT1 onto telomeric chromatin. Furthermore, we found that the telomeric ssDNA-binding activity and binding to TPP1 are required in cis for POT1 to protect telomeres. Mechanistically, binding of POT1 to telomeric ssDNA and association with TPP1 inhibit the localization of RPA, which can function as a DNA damage sensor, to telomeres.

scholarly journals POT1 Association with TRF2 Regulates Telomere Length

2009 ◽  
Vol 29 (20) ◽  
pp. 5611-5619 ◽  
Author(s):  
Megan F. Kendellen ◽  
Katharine S. Barrientos ◽  
Christopher M. Counter
Keyword(s):  
Telomere Length ◽  
Ectopic Expression ◽  
Dna Binding Protein ◽  
Binding Activity ◽  
Critical Aspect ◽  
Interaction Domain ◽  
Ssdna Binding ◽  
Telomere Elongation ◽  
Double Stranded Dna ◽  
Telomeric Protein

ABSTRACT Deleting the OB folds encoding the telomeric single-stranded DNA (ssDNA)-binding activity of the human telomeric protein POT1 induces significant telomere elongation, suggesting that at least one critical aspect of the regulation of telomere length is disrupted by this POT1ΔOB mutant protein. POT1 is known to associate with two proteins through the protein interaction domain retained in POT1ΔOB—the telomeric double-stranded DNA-binding protein TRF2 and the telomere-associated protein TPP1. We report that introducing a mutation that reduces association of POT1 with TRF2, but not a mutation that reduces the association with TPP1, abrogates the ability of POT1ΔOB to promote telomere elongation. Mechanistically, expression of POT1ΔOB reduced the association of TRF2 with POT1, RAP1, and TIN2; however, of these proteins, only ectopic expression of POT1 suppressed the telomere elongation induced by POT1ΔOB. Lastly, replacing endogenous POT1 with a full-length POT1 mutant defective in the association with TRF2 induced telomere elongation. Thus, we conclude that the association of POT1 with both ssDNA and TRF2 is critical for telomere length homeostasis.

Inhibition of Adenovirus DNA Synthesis In Vitro by Sera from Patients with Systemic Lupus Erythematosus

1982 ◽  
Vol 2 (12) ◽  
pp. 1492-1500
Author(s):  
Marshall S. Horwitz ◽  
Beth R. Friefeld ◽  
Harold D. Keiser
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Dna Synthesis ◽  
Lupus Erythematosus ◽  
Inhibitory Activity ◽  
Antinuclear Antibodies ◽  
Dna Binding Protein ◽  
Systemic Lupus ◽  
Double Stranded Dna ◽  
The Individual

Sera containing antinuclear antibodies from patients with systemic lupus erythematosus (SLE) and related disorders were tested for their effect on the synthesis of adenovirus (Ad) DNA in an in vitro replication system. After being heated at 60°C for 1 h, some sera from patients with SLE inhibited Ad DNA synthesis by 60 to 100%. Antibodies to double-stranded DNA were present in 15 of the 16 inhibitory sera, and inhibitory activity copurified with anti-double-stranded DNA in the immunoglobulin G fraction. These SLE sera did not inhibit the DNA polymerases α, β, γ and had no antibody to the 72,000-dalton DNA-binding protein necessary for Ad DNA synthesis. The presence of antibodies to single-stranded DNA and a variety of saline-extractable antigens (Sm, Ha, nRNP, and rRNP) did not correlate with SLE serum inhibitory activity. Methods previously developed for studying the individual steps in Ad DNA replication were used to determine the site of inhibition by the SLE sera that contained antibody to double-stranded DNA. Concentrations of the SLE inhibitor that decreased the elongation of Ad DNA by greater than 85% had no effect on either the initiation of Ad DNA synthesis or the polymerization of the first 26 deoxyribonucleotides.

scholarly journals The rat cytomegalovirus homologue of parvoviral rep genes, r127, encodes a nuclear protein with single- and double-stranded DNA-binding activity that is dispensable for virus replication

2004 ◽  
Vol 85 (7) ◽  
pp. 2001-2013 ◽  
Author(s):  
Koen W. R. van Cleef ◽  
Wendy M. A. Scaf ◽  
Karen Maes ◽  
Suzanne J. F. Kaptein ◽  
Erik Beuken ◽  
...  
Keyword(s):  
Dna Binding ◽  
Virus Replication ◽  
Deletion Mutant ◽  
Nuclear Protein ◽  
Human Herpesvirus ◽  
Binding Activity ◽  
Dna Binding Activity ◽  
Double Stranded Dna

An intriguing feature of the rat cytomegalovirus (RCMV) genome is open reading frame (ORF) r127, which shows similarity to the rep genes of parvoviruses as well as the U94 genes of human herpesvirus type 6A (HHV-6A) and 6B (HHV-6B). Counterparts of these genes have not been found in other herpesviruses. Here, it is shown that the r127 gene is transcribed during the early and late phases of virus replication in vitro as an unspliced 1·1 kb transcript containing the complete r127 ORF. Transcripts of r127 were also detected in various organs of RCMV-infected rats at 1 week post-infection (p.i.), but only in the salivary gland at 4 months p.i. Using rabbit polyclonal antibodies raised against the r127-encoded protein (pr127), pr127 was found to be expressed as early as 12 h p.i. within the nuclei of RCMV-infected cells in vitro. Expression of pr127 was also observed within the nuclei of cells in various organs of RCMV-infected rats at 3 weeks p.i. Moreover, pr127 was demonstrated to bind single- as well as double-stranded DNA. Finally, an RCMV r127 deletion mutant (RCMVΔr127) was generated, in which the r127 ORF was disrupted. This deletion mutant, however, was shown to replicate with a similar efficiency as wild-type RCMV (wt RCMV), both in vitro and in vivo. Taken together, it is concluded that the RCMV r127 gene encodes a nuclear protein with single- and double-stranded DNA-binding activity that is dispensable for virus replication, not only in vitro, but also during the acute phase of infection in vivo.

scholarly journals DNA Damage-induced Association of ATM with Its Target Proteins Requires a Protein Interaction Domain in the N Terminus of ATM

2005 ◽  
Vol 280 (15) ◽  
pp. 15158-15164 ◽  
Author(s):  
Norvin Fernandes ◽  
Yingli Sun ◽  
Shujuan Chen ◽  
Proma Paul ◽  
Reuben J. Shaw ◽  
...  
Keyword(s):  
Dna Damage ◽  
Protein Interaction ◽  
Interaction Domain ◽  
Target Proteins ◽  
N Terminus

scholarly journals Evaluating Effects of HypomorphicThoc1Alleles on Embryonic Development inRb1Null Mice

2016 ◽  
Vol 36 (11) ◽  
pp. 1621-1627 ◽  
Author(s):  
Meenalakshmi Chinnam ◽  
Xiaoling Wang ◽  
Xiaojing Zhang ◽  
David W. Goodrich
Keyword(s):  
Embryonic Development ◽  
Protein Interaction ◽  
Tissue Expression ◽  
Regulatory Genes ◽  
Terminal Protein ◽  
Interaction Domain ◽  
Ribonucleoprotein Complex ◽  
Protein Levels ◽  
Amino Terminal ◽  
Expression Of Genes

The Rb1 tumor suppressor protein is a molecular adaptor that physically links transcription factors like E2f with various proteins acting on DNA or RNA to repress gene expression. Loss of Rb1 liberates E2f to activate the expression of genes mediating resulting phenotypes. Most Rb1 binding proteins, including E2f, interact through carboxyl-terminal protein interaction domains, but genetic evidence suggests that an amino-terminal protein interaction domain is also important. One protein that binds Rb1 through the amino-terminal domain is encoded byThoc1, a required component of the THO ribonucleoprotein complex important for RNA processing and transport. The physiological relevance of this interaction is unknown. Here we tested whetherThoc1mediates effects ofRb1loss on mouse embryonic development. We found thatThoc1deficiency delays embryo death, and this delay correlates with reduced apoptosis in the brain. E2f protein levels are reduced inRb1:Thoc1-deficient brain tissue. Expression of apoptotic regulatory genes regulated by E2f, like Apaf1 and Bak1, is also reduced. These observations suggest thatThoc1is required to support increased expression of E2f and apoptotic regulatory genes that trigger apoptosis uponRb1loss. These findings implicate Rb1 in the regulation of the THO ribonucleoprotein complex.

scholarly journals The Single-Strand DNA Binding Activity of Human PC4 Prevents Mutagenesis and Killing by Oxidative DNA Damage

2004 ◽  
Vol 24 (13) ◽  
pp. 6084-6093 ◽  
Author(s):  
Jen-Yeu Wang ◽  
Altaf Hossain Sarker ◽  
Priscilla K. Cooper ◽  
Michael R. Volkert
Keyword(s):  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Binding Activity ◽  
Single Strand ◽  
Physical Interaction ◽  
Ssdna Binding ◽  
Dna Binding Activity ◽  
Resistance Pathway ◽  
Domain Of Unknown Function ◽  
Human Proteins

ABSTRACT Human positive cofactor 4 (PC4) is a transcriptional coactivator with a highly conserved single-strand DNA (ssDNA) binding domain of unknown function. We identified PC4 as a suppressor of the oxidative mutator phenotype of the Escherichia coli fpg mutY mutant and demonstrate that this suppression requires its ssDNA binding activity. Saccharomyces cerevisiae mutants lacking their PC4 ortholog Sub1 are sensitive to hydrogen peroxide and exhibit spontaneous and peroxide-induced hypermutability. PC4 expression suppresses the peroxide sensitivity of the yeast sub1Δ mutant, suggesting that the human protein has a similar function. A role for yeast and human proteins in DNA repair is suggested by the demonstration that Sub1 acts in a peroxide resistance pathway involving Rad2 and by the physical interaction of PC4 with the human Rad2 homolog XPG. We show that XPG recruits PC4 to a bubble-containing DNA substrate with a resulting displacement of XPG and formation of a PC4-DNA complex. We discuss the possible requirement for PC4 in either global or transcription-coupled repair of oxidative DNA damage to mediate the release of XPG bound to its substrate.

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