Homologous and illegitimate recombination in developing Xenopus oocytes and eggs.
Exogenous DNA is efficiently recombined when injected into the nuclei of Xenopus laevis oocytes. This reaction proceeds by a homologous resection-annealing mechanism which depends on the activity of a 5'-->3' exonuclease. Two possible functions for this recombination activity have been proposed: it may be a remnant of an early process in oogenesis, such as meiotic recombination or amplification of genes coding for rRNA, or it may reflect materials stored for embryogenesis. To test these hypotheses, recombination capabilities were examined with oocytes at various developmental stages. Late-stage oocytes performed only homologous recombination, whereas the smallest oocytes ligated the restriction ends of the injected DNA but supported no homologous recombination. This transition from ligation to recombination activity was also seen in nuclear extracts from these same stages. Exonuclease activity was measured in the nuclear extracts and found to be low in early stages and then to increase in parallel with recombination capacity in later stages. The accumulation of exonuclease and recombination activities during oogenesis suggests that they are stored for embryogenesis and are not present for oocyte-specific functions. Eggs were also tested and found to catalyze homologous recombination, ligation, and illegitimate recombination. Retention of homologous recombination in eggs is consistent with an embryonic function for the resection-annealing mechanism. The observation of all three reactions in eggs suggests that multiple pathways are available for the repair of double-strand breaks during the extremely rapid cleavage stages after fertilization.
Lehman
C W
CW
Department of Biochemistry, University of Utah School of Medicine, Salt Lake City 84132.
Clemens
M
M
Worthylake
D K
DK
Trautman
J K
JK
Carroll
D
D
D000818:Animals; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004275:DNA, Ribosomal / Q000378:metabolism*; D005090:Exodeoxyribonucleases / Q000378:metabolism; D005260:Female; D005306:Fertilization; D066298:In Vitro Techniques; D007700:Kinetics; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism; D009865:Oocytes / Q000166:cytology / Q000502:physiology*; D010062:Ovulation Induction; D010063:Ovum / Q000166:cytology / Q000502:physiology*; D011995:Recombination, Genetic; D015183:Restriction Mapping; D014982:Xenopus laevis
D004275:DNA, Ribosomal; D009687:Nuclear Proteins; D004247:DNA; D005090:Exodeoxyribonucleases