Phospho-RPA32/RPA2 (Ser4/8) Antibody [M17K6] Datasheet

Biological Description

Specificity	Phospho-RPA32 (Ser4/8) Antibody [M17K6] detects endogenous levels of total RPA32 protein only when it is phosphorylated at Ser4/8.
Background	Phospho‑RPA32 (Ser4/8) marks a hyperphosphorylated, damage‑responsive state of the 32‑kDa subunit of replication protein A in which DNA‑PK–dependent modification of two N‑terminal serines reprograms RPA’s functions at stalled replication forks and double‑strand breaks, coordinating checkpoint signaling, recombination, fork restart, and the decision between recovery and mitotic catastrophe. RPA32 resides in the trimeric RPA complex together with RPA70 and RPA14 and contributes an N‑terminal regulatory tail that is phosphorylated by PIKK kinases and CDKs and a central OB‑fold ssDNA‑binding domain; Ser4 and Ser8 lie in this unstructured N‑terminus and are among the earliest sites to become phosphorylated when replication stress generates extended RPA‑coated ssDNA. Under replication stress, ATR activation by RPA–ssDNA complexes triggers a first wave of RPA32 phosphorylation at sites such as Ser33 that supports Chk1 activation and replication arrest, while DNA‑PK subsequently phosphorylates Ser4/Ser8, and cells either lacking DNA‑PK activity or expressing a Ser4/8A mutant show nearly identical phenotypes, including defective replication checkpoint arrest, premature fork restart, failure to suppress late origin firing, ATM‑dependent hyper‑recombination, and increased mitotic catastrophe, indicating that Ser4/8 phosphorylation is a critical effector branch of DNA‑PK signaling at stalled forks. These modifications influence how RPA interfaces with other repair factors: Ser4/8 phosphorylation promotes proper MRE11 and TopBP1 phosphorylation and helps maintain ATR–Chk1 signaling, while also modulating recruitment and turnover of recombination proteins such as RAD51, so that loss of this modification leads to excessive homologous recombination events that compromise genome stability rather than resolving damage faithfully.

Specificity

Phospho-RPA32 (Ser4/8) Antibody [M17K6] detects endogenous levels of total RPA32 protein only when it is phosphorylated at Ser4/8.

Background

Phospho‑RPA32 (Ser4/8) marks a hyperphosphorylated, damage‑responsive state of the 32‑kDa subunit of replication protein A in which DNA‑PK–dependent modification of two N‑terminal serines reprograms RPA’s functions at stalled replication forks and double‑strand breaks, coordinating checkpoint signaling, recombination, fork restart, and the decision between recovery and mitotic catastrophe. RPA32 resides in the trimeric RPA complex together with RPA70 and RPA14 and contributes an N‑terminal regulatory tail that is phosphorylated by PIKK kinases and CDKs and a central OB‑fold ssDNA‑binding domain; Ser4 and Ser8 lie in this unstructured N‑terminus and are among the earliest sites to become phosphorylated when replication stress generates extended RPA‑coated ssDNA. Under replication stress, ATR activation by RPA–ssDNA complexes triggers a first wave of RPA32 phosphorylation at sites such as Ser33 that supports Chk1 activation and replication arrest, while DNA‑PK subsequently phosphorylates Ser4/Ser8, and cells either lacking DNA‑PK activity or expressing a Ser4/8A mutant show nearly identical phenotypes, including defective replication checkpoint arrest, premature fork restart, failure to suppress late origin firing, ATM‑dependent hyper‑recombination, and increased mitotic catastrophe, indicating that Ser4/8 phosphorylation is a critical effector branch of DNA‑PK signaling at stalled forks. These modifications influence how RPA interfaces with other repair factors: Ser4/8 phosphorylation promotes proper MRE11 and TopBP1 phosphorylation and helps maintain ATR–Chk1 signaling, while also modulating recruitment and turnover of recombination proteins such as RAD51, so that loss of this modification leads to excessive homologous recombination events that compromise genome stability rather than resolving damage faithfully.

Usage Information

Application

WB, IP, IF, FCM

Dilution

WB	IP	IF	FCM
1:1000	1:1000	1:5000	1:2000

Reactivity

Human

Source

Rabbit Monoclonal Antibody

MW

29 kDa

Storage Buffer

PBS, pH 7.2+50% Glycerol+0.05% BSA+0.01% NaN3

Storage
(from the date of receipt)

-20°C (avoid freeze-thaw cycles), 2 years

References

https://pubmed.ncbi.nlm.nih.gov/24819595/
https://pubmed.ncbi.nlm.nih.gov/22977173/

Phospho-RPA32/RPA2 (Ser4/8) Antibody [M17K6]

Biological Description

Usage Information

References

Application Data