Fig 3.
Cid13 is a Cid1- and poly(A) polymerase-related protein that suppresses the HU sensitivity of rad3 mutants. (A) Alignment of amino acid sequences of the putative catalytic cores of Cid1, Cid11, Cid13, and S. pombe poly(A) polymerase. Amino acid residue numbers are indicated to the left of each sequence, conserved residues are highlighted on a black background, and conservative substitutions are shaded. The Pol β superfamily motif is underlined, and the two conserved aspartate residues changed to alanine in the Cid1 DADA mutant are indicated by asterisks. (B and C) Survival of rad3ts or rad3kd cells transformed with pUR-cid13 (cid13oe) or pUR (vector), as indicated, after plating onto agar containing various doses of HU (B) or after UV irradiation on agar plates (C). Survival was scored by counting colonies after 4 days of incubation at 32°C. (D) Survival of the same strains after short-term exposure to 5 mM HU in liquid culture at 32.5°C. Aliquots of cells taken at the times indicated were scored for viability by plating onto agar lacking HU and incubation at 32°C for 3 days. (E) Cells of the rad3ts strain transformed with pUR-cid13 (cid13oe) or pUR (vector), as indicated, were exposed to 5 mM HU at 32°C in duplicate liquid cultures for 3 h and were then assayed for Cds1 kinase activity in vitro using myelin basic protein (MBP) as substrate. (F) The strains indicated were inoculated onto plates containing various doses of HU, and survival was scored by counting colonies after 4–5 days of incubation at 32°C.