Figure 3.

Overproduction of Abf2p results in an increase in Holliday junctions in ρ⁺ mtDNA. (A) Schematic of Abf2p with the HMG-box domains represented by the gray boxes. Mutations were made in conserved basic residues, as indicated, from both HMG-box domains resulting in mutant protein Abf2p1⁻2⁻. (B) Overproduction of Abf2p and Abf2p1⁻2⁻ in 14WW ρ⁺ cells after the transition from YNBG to YNBGal medium for the indicated time points. Abf2p and Abf21⁻2⁻ were overexpressed from transformants containing the CEN plasmid GAL1–10 promoter constructs pGal-Abf2 and pGal-Abf21⁻2⁻, respectively. Protein levels were detected by Western blotting with polyclonal antiserum raised against Abf2p (18). (C) 2D gel electrophoresis of ρ⁺ mtDNA replication and recombination intermediates from 14WW cells containing either pGal-Abf2 or pGal-Abf21⁻2⁻ after the transition from YNBG to YNBGal medium. The blots were hybridized with a probe specific for a 5-kb EcoRI fragment from the COX1 gene.