Mechanisms of overlap formation in nonhomologous DNA end joining

1994 ◽  
Vol 14 (2) ◽  
pp. 888-895
Author(s):  
P Pfeiffer ◽  
S Thode ◽  
J Hancke ◽  
W Vielmetter
Keyword(s):  
Single Strand ◽  
Illegitimate Recombination ◽  
Gap Filling ◽  
End Joining ◽  
Base Pairs ◽  
Junction Formation ◽  
Egg Extracts ◽  
Xenopus Egg ◽  
Xenopus Egg Extracts ◽  
Dna Ends

Rejoining of nonhomologous DNA termini plays a central role in processes of illegitimate recombination. In Xenopus egg extracts, DNA ends with noncomplementary 4-nucleotide antiparallel single-strand protrusions are assumed to be joined by formation of short mismatched overlap intermediates. The extents of these overlaps may be set by single fortuitously matching base pairs and determine the patterns of subsequent gap filling and nick ligation. Under conditions of alternative overlap settings, rules for the most probable joining pathway and the effects of mismatches on junction formation were analyzed. We show that in certain cases, fill-in and ligation converting overlap intermediates into covalently closed junctions may proceed in the presence of unrepaired mismatches, whereas in other cases, completion of junction formation is preceded by removal of mismatches. Results are discussed in relation with "alignment" proteins postulated to structurally support overlap heteroduplexes during junction formation.

scholarly journals Mechanisms of overlap formation in nonhomologous DNA end joining.

1994 ◽  
Vol 14 (2) ◽  
pp. 888-895 ◽  
Author(s):  
P Pfeiffer ◽  
S Thode ◽  
J Hancke ◽  
W Vielmetter
Keyword(s):  
Single Strand ◽  
Illegitimate Recombination ◽  
Gap Filling ◽  
End Joining ◽  
Base Pairs ◽  
Junction Formation ◽  
Egg Extracts ◽  
Xenopus Egg ◽  
Xenopus Egg Extracts ◽  
Dna Ends

Rejoining of nonhomologous DNA termini plays a central role in processes of illegitimate recombination. In Xenopus egg extracts, DNA ends with noncomplementary 4-nucleotide antiparallel single-strand protrusions are assumed to be joined by formation of short mismatched overlap intermediates. The extents of these overlaps may be set by single fortuitously matching base pairs and determine the patterns of subsequent gap filling and nick ligation. Under conditions of alternative overlap settings, rules for the most probable joining pathway and the effects of mismatches on junction formation were analyzed. We show that in certain cases, fill-in and ligation converting overlap intermediates into covalently closed junctions may proceed in the presence of unrepaired mismatches, whereas in other cases, completion of junction formation is preceded by removal of mismatches. Results are discussed in relation with "alignment" proteins postulated to structurally support overlap heteroduplexes during junction formation.

scholarly journals N-terminal region of RecQ4 inhibits non-homologous end joining and chromatin association of the Ku heterodimer in Xenopus egg extracts

Gene ◽  
2021 ◽  
pp. 145647
Author(s):  
Takashi Tsuyama ◽  
Kumiko Fujita ◽  
Ryosuke Sasaki ◽  
Shiori Hamanaka ◽  
Yuki Sotoyama ◽  
...  
Keyword(s):  
Terminal Region ◽  
End Joining ◽  
Egg Extracts ◽  
Xenopus Egg ◽  
Non Homologous End Joining ◽  
Xenopus Egg Extracts

scholarly journals The bulk of unpolymerized actin in Xenopus egg extracts is ATP-bound.

1995 ◽  
Vol 6 (2) ◽  
pp. 227-236 ◽  
Author(s):  
J Rosenblatt ◽  
P Peluso ◽  
T J Mitchison
Keyword(s):  
Actin Polymerization ◽  
Critical Concentration ◽  
Large Fraction ◽  
Nucleotide Exchange ◽  
Chromatography Analysis ◽  
Thymosin Beta 4 ◽  
Egg Extracts ◽  
Xenopus Egg ◽  
Xenopus Egg Extracts

Non-muscle cells contain 15-500 microM actin, a large fraction of which is unpolymerized. Thus, the concentration of unpolymerized actin is well above the critical concentration for polymerization in vitro (0.2 microM). This fraction of actin could be prevented from polymerization by being ADP bound (therefore less favored to polymerize) or by being ATP bound and sequestered by a protein such as thymosin beta 4, or both. We isolated the unpolymerized actin from Xenopus egg extracts using immobilized DNase 1 and assayed the bound nucleotide. High-pressure liquid chromatography analysis showed that the bulk of soluble actin is ATP bound. Analysis of actin-bound nucleotide exchange rates suggested the existence of two pools of unpolymerized actin, one of which exchanges nucleotide relatively rapidly and another that apparently does not exchange. Native gel electrophoresis of Xenopus egg extracts demonstrated that most of the soluble actin exists in complexes with other proteins, one of which might be thymosin beta 4. These results are consistent with actin polymerization being controlled by the sequestration and release of ATP-bound actin, and argue against nucleotide exchange playing a major role in regulating actin polymerization.

scholarly journals The Prereplication Complex Recruits XEco2 to Chromatin to Promote Cohesin Acetylation in Xenopus Egg Extracts

2012 ◽  
Vol 22 (11) ◽  
pp. 977-988 ◽  
Author(s):  
Torahiko L. Higashi ◽  
Megumi Ikeda ◽  
Hiroshi Tanaka ◽  
Takuro Nakagawa ◽  
Masashige Bando ◽  
...  
Keyword(s):  
Egg Extracts ◽  
Xenopus Egg ◽  
Xenopus Egg Extracts

scholarly journals Multiple mechanisms determine ER network morphology during the cell cycle in Xenopus egg extracts

2013 ◽  
Vol 203 (5) ◽  
pp. 801-814 ◽  
Author(s):  
Songyu Wang ◽  
Fabian B. Romano ◽  
Christine M. Field ◽  
Tim J. Mitchison ◽  
Tom A. Rapoport
Keyword(s):  
Cell Cycle ◽  
Adaptor Protein ◽  
Sensitive Factor ◽  
Protein Receptors ◽  
Egg Extracts ◽  
Organizing Center ◽  
Xenopus Egg ◽  
Guanosine Triphosphatase ◽  
Xenopus Egg Extracts ◽  
Network Morphology

In metazoans the endoplasmic reticulum (ER) changes during the cell cycle, with the nuclear envelope (NE) disassembling and reassembling during mitosis and the peripheral ER undergoing extensive remodeling. Here we address how ER morphology is generated during the cell cycle using crude and fractionated Xenopus laevis egg extracts. We show that in interphase the ER is concentrated at the microtubule (MT)-organizing center by dynein and is spread by outward extension of ER tubules through their association with plus ends of growing MTs. Fusion of membranes into an ER network is dependent on the guanosine triphosphatase atlastin (ATL). NE assembly requires fusion by both ATL and ER-soluble N-ethyl-maleimide–sensitive factor adaptor protein receptors. In mitotic extracts, the ER converts into a network of sheets connected by ER tubules and loses most of its interactions with MTs. Together, these results indicate that fusion of ER membranes by ATL and interaction of ER with growing MT ends and dynein cooperate to generate distinct ER morphologies during the cell cycle.

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