Common mechanisms of PIKK regulation
Introduction
Genome maintenance is critical to prevent disease. Challenges to genome integrity come from environmental mutagens, byproducts of cellular respiration, and errors during nucleic acid metabolism including DNA replication. Cells have DNA damage response (DDR) activities that continually monitor the integrity of the DNA and function to prevent the occurrence of deleterious mutations and rearrangements. The DDR is regulated by the phosphoinositide three-kinase-related protein kinases (PIKKs). The PIKKs primarily responsible for signaling the presence of DNA damage include ATM, ATR and DNA-PKcs. These PIKKs phosphorylate hundreds of proteins that maintain genome integrity through regulation of cell cycle progression, DNA repair, apoptosis, and cellular senescence.
Human cells also contain three additional PIKKs (SMG1, mTOR, and TRRAP) with activities in other biological pathways. SMG1 primarily controls nonsense-mediated mRNA decay, mTOR regulates nutrient-dependent signaling, and TRRAP regulates transcription but lacks kinase activity. Insights into mTOR and SMG1 regulation are pertinent to the DNA damage regulated PIKKs.
Section snippets
PIKK complexes
The PIKK enzymes are large proteins (between 2547 and 4128 amino acids) that share a common domain structure (Fig. 1). The kinase domain is located near the C-terminus and is flanked by two regions of sequence similarity among all the PIKKs called the FAT (FRAP, ATM, TRRAP) and FATC (FAT C-terminus) domains. The FAT domain consists of HEAT (Huntingtin, Elongation factor 3, A subunit of protein phosphatase 2A and TOR1) repeats. The FATC domain is small (32 amino acids) and is required for PIKK
ATR and ssDNA gaps
ATR appears to be the most versatile of the PIKK family of DNA damage-responsive kinases, being activated by a variety of DNA lesions including base adducts, cross-links, DSBs, and compounds that directly promote replication stress such as hydroxyurea and aphidicolin. In addition to responding to environmental mutagens that induce replication stress, ATR is essential for the viability of replicating cells [7], [10]. ATR is activated during every cell cycle to regulate replication origin firing
PIKK activators
The localization of PIKKs to subcellular compartments promotes kinase activation by concentrating each kinase with an activating protein. ATM, ATR, and DNA-PK are activated through protein–protein interactions with Mre11, TopBP1, and Ku70/80, respectively.
Post-translational modifications and PIKK structures
How does the interaction of TopBP1 with ATR or MRN with ATM activate these kinases? Most likely, these interactions cause a conformational change that increases the ability of these kinases to access substrates [52]. In response to DNA damage, ATM transitions from an inactive dimer to an active monomer [29]. This transition may be facilitated by the MRN complex and function to release ATM autoinhibition. ATM autophosphorylation correlates with the transition. Mutations that remove these
Summary and perspective
The PIKKs are unusual protein kinases in that their sequence suggests common evolution from lipid kinases. The common evolution is apparent in the similarity of their regulation. While responding to different input signals, recurrent themes of regulation include dynamic localization dependent on partner proteins and a protein or protein/nucleic acid activator (Fig. 2).
PIKK relocalization concentrates the kinases to where their activator is found. This concentration may be primarily responsible
Note added in proof
TopBP1 was recently discovered to function to recruit the 9-1-1 complex to stalled replication forks placing it both upstream and downstream of Rad9 (S. Yan and W.M. Michael, TopBP1 and DNA polymerase-alpha directly recruit the 9-1-1 complex to stalled DNA replication forks, J Cell Biol. 184 (2009) 793-804).
Conflict of Interest
None.
Funding: C.A.L. is supported by a Department of Defesne Breast Cancer Research Program pre-doctoral fellowship (W81XWH-06-1-0528).
Acknowledgements
The authors would like to thank Daniel Mordes and other members of the Cortez laboratory for their insightful contributions. Studies on ATR in the author's laboratory are supported by R01 CA102729.
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