title: Engineering an analog-sensitive CDK12 cell line using CRISPR/Cas.
abstract: The RNA Polymerase II C-terminal domain (CTD) kinase CDK12 has been implicated as a tumor suppressor and regulator of DNA damage response genes. Although much has been learned about CDK12 and its activity, due to the lack of a specific inhibitor and the complications posed by long term RNAi depletion, much is still unknown about the particulars of CDK12 function. Therefore gaining a better understanding of CDK12's roles at the molecular level will be challenging without the development of additional tools. In order to address these issues we have used the CRISPR/Cas gene engineering system to create a mammalian cell line in which the only functional copy of CDK12 is selectively inhibitable by a cell-permeable adenine analog (analog-sensitive CDK12). Inhibition of CDK12 results in a perturbation of the phosphorylation patterns on the CTD and an arrest in cellular proliferation. This cell line should serve as a powerful tool for future studies.
authors.affiliation: Department of Biochemistry, Duke University Medical Center, United States.
authors.forename: Bartlomiej
authors.initials: B
authors.lastname: Bartkowiak
authors.affiliation: Department of Biochemistry, Duke University Medical Center, United States.
authors.forename: Christopher
authors.initials: C
authors.lastname: Yan
authors.affiliation: Department of Biochemistry, Duke University Medical Center, United States. Electronic address: arno@duke.edu.
authors.forename: Arno L
authors.initials: AL
authors.lastname: Greenleaf
mesh_terms: D001483:Base Sequence; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics*; D018844:Cyclin-Dependent Kinases / Q000235:genetics*; D004247:DNA; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D012689:Sequence Homology, Nucleic Acid
chemical_list: D004247:DNA; C447339:CDK12 protein, human; D018844:Cyclin-Dependent Kinases
keywords: Analog-sensitive; CDK12; CRISPR/Cas; CTD phosphorylation; Cell line; Unintended splice site