umuDC -Mediated Cold Sensitivity Is a Manifestation of Functions of the UmuD 2 C Complex Involved in a DNA Damage Checkpoint Control

Abstract

ABSTRACT The umuDC genes are part of the Escherichia coli SOS response, and their expression is induced as a consequence of DNA damage. After induction, they help to promote cell survival via two temporally separate pathways. First, UmuD and UmuC together participate in a cell cycle checkpoint control; second, UmuD′ 2 C enables translesion DNA replication over any remaining unrepaired or irreparable lesions in the DNA. Furthermore, elevated expression of the umuDC gene products leads to a cold-sensitive growth phenotype that correlates with a rapid inhibition of DNA synthesis. Here, using two mutant umuC alleles, one that encodes a UmuC derivative that lacks a detectable DNA polymerase activity ( umuC104 ; D101N) and another that encodes a derivative that is unable to confer cold sensitivity but is proficient for SOS mutagenesis ( umuC125 ; A39V), we show that umuDC -mediated cold sensitivity can be genetically separated from the role of UmuD′ 2 C in SOS mutagenesis. Our genetic and biochemical characterizations of UmuC derivatives bearing nested deletions of C-terminal sequences indicate that umuDC -mediated cold sensitivity is not due solely to the single-stranded DNA binding activity of UmuC. Taken together, our analyses suggest that umuDC -mediated cold sensitivity is conferred by an activity of the UmuD 2 C complex and not by the separate actions of the UmuD and UmuC proteins. Finally, we present evidence for structural differences between UmuD and UmuD′ in solution, consistent with the notion that these differences are important for the temporal regulation of the two separate physiological roles of the umuDC gene products.