Biochemical characterization of human shieldin complex

Abstract

Shieldin is a newly identified DNA repair effector involved in the repair of DNA doublestrand breaks (DSBs) in G1 phase of the cell cycle. Shieldin is a four-component complex consisting of proteins SHLD1, SHLD2, SHLD3 and HORMA domain protein REV7.Shieldin inhibits homologous recombination (HR) through its direct ssDNA binding activity and directs the repair pathway to non-homologous end joining (NHEJ). Despite the clear understanding of Shieldin function, the basis of its recruitment and assembly at DSBs is not well understood. RTRoom temperature PEG Polyethylene glycolNaOH Sodium hydroxide DDR DNA damage response DNA-PK complex. KU70/80 are abundant in nuclear cytoplasm and show high affinity for DNA ends. In the next step, DNA-PK complex recruits endonuclease Artemis. DNA-PKcs activate Artemis by undergoing autophosphorylation (Goodarzi et al., 2006). This makes the DNA ends compatible for ligation reaction. The ligation machinery composed of XRCC4, ligase IV and KLF is then recruited for re-ligation. This type of repair requires minimal to no resection and may potentially lead to small insertion and deletions. Due to this error-prone nature, NHEJ is required for immune receptor diversification as both V(D)J recombination and class switch recombination (CSR) are mediated by it. Along with core factors as mentioned above the pathway activation is carried out by 53BP1 and its