A multi-subunit enzyme, DNA-PK consists of a catalytic subunit (DNA-PKcs), p460, and a regulatory subunit called Ku. The Ku protein is a heterodimer composed of 70-kDa (Ku70) and 80-kDa (Ku80) subunits and has the capability of binding selectively to specific forms of Lapatinib Ditosylate DNA [25,26]. In this capacity, it functions as the regulator of DNA-PK that is active in transcription, DNA recombination, and DNA repair [27-29]. Its role in DNA repair was not formally proven until the emergence of studies implicating Ku as the defective factor in cells hypersensitive to DNA-damaging agents [30]. DNA-dependent protein kinase activators Ku binds DNA ends in a sequence-independent manner, and in the absence of DNA-PKcs, the extreme DNA terminus is bound in an accessible channel [31].

Ku has strong avidity for DNA with a variety of end structures, such as blunt, over-hanged, hair-pinned, and damaged. Ku can also recognize gaps and nicks in dsDNA, indicating possible roles of DNA-PK in the repair of damage other than DSBs [32], and is particularly suited to do this since DNA-PKcs assembles onto Ku-bound DNA regardless of end structure [33]. Although DNA-PKcs has innate affinity (itself) for DNA ends (in low salt conditions), Ku is required for targeting DNA-PKcs to damaged DNA in physiologic conditions and in living cells [34]. Although any DSB discontinuity can activate DNA-PK, its activation varies considerably depending on the end structure, and studies show kinase activation in trans (achieved by kinase autophosphrylation) or cis (achieved by specific DNA strand orientation and sequence bias) [35].

Activation by these dual processes represents a potentially powerful mechanism by which DNA-PK protects DNA ends to maintain genome integrity. After the DSB repair, Ku likely remains trapped on the DNA [36]. How Ku gets removed from the DNA is not clear, although a protease-mediated degradation of Ku80 has been speculated [36]. Cellular and molecular targets of DNA-dependent protein kinase in non-homologous end joining Many proteins have been listed as excellent in vitro and in vivo DNA-PK targets, but the functional relevance of their phosphorylation by DNA-PK remains mostly unclear. Most of the proteins involved in NHEJ (XRCC4, Ku70, Ku80, Artemis, DNA-PKcs, and XLF) are excellent in vitro and in vivo targets of DNA-PK [2,37-40].

When a DSB occurs, the Ku heterodimer (Ku80/Ku70) first binds to the broken ends by using Ku80 and then recruits the DNA-PKcs, which is activated upon binding to Artemis nuclease, and the repair process is completed by XRCC4-DNA ligase IV [39,41] (Figure ?(Figure1).1). Physical association of DNA-PKcs Drug_discovery and its enzymatic selleck chemicals llc activity are required for Artemis’s endonucleolytic activity [39]. In the absence of DNA damage, Artemis is complexed with DNA-PKcs. It has been shown that DNA-PKcs is targeted to Ku-bound DNA; Artemis is released from DNA-PKcs and is rebound again only when the kinase is activated [34].