Cyclin-Dependent Kinase Phosphorylation of Polynucleotide Kinase Phosphatase Promotes Okazaki Fragment Processing and Genome Stability During DNA Replication
Core Concepts
Phosphorylation of PNKP at threonine 118 by CDKs is essential for its recruitment to single-strand DNA gaps between Okazaki fragments, allowing PNKP to process the ends and promote their ligation, thereby maintaining genome stability during DNA replication.
Abstract
The study investigates the role of polynucleotide kinase phosphatase (PNKP) in DNA replication and genome stability. Key findings:
-
PNKP knockout cells exhibit slower cell growth, accumulation in S phase, and faster replication fork progression, suggesting PNKP is important for proper DNA replication.
-
The linker region of PNKP, especially the phosphorylation of threonine 118 (T118), is critical for cell proliferation and replication fork progression.
-
CDK1/Cyclin A2 and CDK2/Cyclin A2 complexes phosphorylate PNKP at T118, mainly during S phase, allowing PNKP to be recruited to single-strand DNA gaps between Okazaki fragments (OFs).
-
The phosphatase activity of PNKP is required for processing the ends of these single-strand gaps to facilitate OF ligation and prevent post-replicative single-strand DNA gaps.
-
PNKP also plays a role in the PARP1-dependent gap-filling pathway when OFs are not properly ligated.
-
Disruption of PNKP phosphorylation at T118 or its enzymatic activities leads to genome instability, highlighting the importance of PNKP in maintaining replication fidelity.
Translate Source
To Another Language
Generate MindMap
from source content
CDKs-mediated phosphorylation of PNKP is required for end-processing of single-strand DNA gaps on Okazaki Fragments and genome stability
Stats
PNKP knockout cells showed slower cell growth and faster replication fork progression compared to wild-type cells.
Phosphorylation of PNKP at threonine 118 (T118) by CDK1/Cyclin A2 and CDK2/Cyclin A2 complexes is required for its recruitment to single-strand DNA gaps between Okazaki fragments.
The phosphatase activity of PNKP is essential for processing the ends of these single-strand gaps to facilitate Okazaki fragment ligation.
Disruption of PNKP phosphorylation at T118 or its enzymatic activities leads to increased genome instability.
Quotes
"Phosphorylation of PNKP at threonine 118 (T118) in the linker region is required for its recruitment to single-strand DNA nicks and/or gaps between Okazaki fragments (OFs) during DNA replication."
"PNKP enzymatic activity, especially phosphatase activity, is required for processing of the ends of single-strand gaps on OFs."
"Phosphorylation-mediated PNKP recruitment to single-strand DNA gaps on OFs and the end-processing activity of PNKP are critically important for preventing genome instability through appropriate regulation of DNA replication."
Deeper Inquiries
How might the regulation of PNKP phosphorylation and activity be exploited for cancer therapy?
Phosphorylation of PNKP at T118 by CDKs plays a crucial role in the recruitment of PNKP to single-strand DNA gaps, particularly those between Okazaki fragments during DNA replication. This recruitment is essential for the end-processing of these gaps and the prevention of genome instability. Exploiting the regulation of PNKP phosphorylation and activity could be a potential target for cancer therapy. By targeting the phosphorylation of PNKP at T118, it may be possible to disrupt the recruitment of PNKP to DNA gaps, leading to the accumulation of unligated Okazaki fragments and subsequent genome instability. This approach could be particularly effective in cancer cells that rely heavily on rapid DNA replication and are more susceptible to disruptions in the DNA repair process.
What other DNA repair or replication factors might interact with PNKP to coordinate Okazaki fragment processing and genome stability?
Several DNA repair and replication factors are likely to interact with PNKP to coordinate Okazaki fragment processing and maintain genome stability. One key factor is XRCC1, which is a binding scaffold protein of PNKP in the single-strand break repair pathway. Additionally, FEN1, a flap endonuclease involved in the removal of RNA/DNA fragments from Okazaki fragments, likely interacts with PNKP to ensure proper end-processing and ligation of Okazaki fragments. PARP1, which is involved in the repair of single-strand breaks and multiple DNA replication processes, may also interact with PNKP to facilitate the PARP1-dependent gap-filling pathway for Okazaki fragment maturation. Other factors such as RPA2, PCNA, and LIG3 may also play a role in coordinating Okazaki fragment processing and genome stability in conjunction with PNKP.
Could defects in PNKP-mediated Okazaki fragment processing contribute to the neurodevelopmental and neurodegenerative disorders associated with PNKP mutations?
Defects in PNKP-mediated Okazaki fragment processing could indeed contribute to the neurodevelopmental and neurodegenerative disorders associated with PNKP mutations. PNKP is crucial for the repair of DNA damage, including single-strand breaks and double-strand breaks, as well as the processing of Okazaki fragments during DNA replication. Mutations in PNKP that affect its enzymatic activities, such as the phosphatase and kinase functions, can lead to the accumulation of DNA damage and the formation of unligated Okazaki fragments. This accumulation of DNA damage and incomplete DNA replication can result in genomic instability, which is known to be a contributing factor to neurodevelopmental disorders like microcephaly and seizures, ataxia oculomotor apraxia, and Charcot-Marie-Tooth disease. Therefore, defects in PNKP-mediated Okazaki fragment processing could play a significant role in the pathogenesis of these disorders.
