abp1+and abp2+function downstream of SPBC2A9.02 and SPAC27D7.08c to initiate DNA replication. (A) Reduced expression levels of abp1+ and abp2+ in SPBC2A9.02 Δ and SPAC27D7.08c Δ. The mRNA levels were quantified by real time PCR and those of act1+ served as an internal control (n=3). The relative level in WT was designated as arbitrary unit 1. (B) Overexpression of abp1+ and abp2+ partially rescued the growth defect of SPBC2A9.02 Δ and SPAC27D7.08c Δ. pREP1-abp1+ or pREP1-abp2+ were transformed into each deletion separately. pREP1-abp1+ and pJR2-41U-abp2+ were co-transformed into SPBC2A9.02 Δ or SPAC27D7.08c Δ. Transformants were harvested and 5-fold serial dilutions were spotted on plates supplemented with DNA damage reagents. Plates were photographed after 3 days of incubation at 32°C. (C) Overexpression of abp1+ or abp2+ partially relieved the G1-arrest in SPBC2A9.02 Δ and SPAC27D7.08c Δ. Transformants described in Figure 4B were grown to logarithmic phase and harvested for flow cytometry analysis. Reproducible results were obtained in three independent experiments.