A Multimodel Study of the Role of Novel PKC Isoforms in the DNA Integrity Checkpoint

The protein kinase C (PKC) family plays important regulatory roles in numerous cellular processes. Saccharomyces cerevisiae contains a single PKC, Pkc1, whereas in mammals, the PKC family comprises nine isoforms. Both Pkc1 and the novel isoform PKC delta are involved in the control of DNA integrity checkpoint activation, demonstrating that this mechanism is conserved from yeast to mammals. To explore the function of PKC delta in a non-tumor cell line, we employed CRISPR-Cas9 technology to obtain PKC delta knocked-out mouse embryonic stem cells (mESCs). This model demonstrated that the absence of PKC delta reduced the activation of the effector kinase CHK1, although it suggested that other isoform(s) might contribute to this function. Therefore, we used yeast to study the ability of each single PKC isoform to activate the DNA integrity checkpoint. Our analysis identified that PKC theta, the closest isoform to PKC delta, was also able to perform this function, although with less efficiency. Then, by generating truncated and mutant versions in key residues, we uncovered differences between the activation mechanisms of PKC delta and PKC theta and identified their essential domains. Our work strongly supports the role of PKC as a key player in the DNA integrity checkpoint pathway and highlights the advantages of combining distinct research models.