Mechanism and Regulation of DNA-Protein Crosslink Repair by the DNA-Dependent Metalloprotease SPRTN

Abstract

SummaryCovalent DNA-protein crosslinks (DPCs) are toxic DNA lesions that interfere with essential chromatin transactions, such as replication and transcription. Little was known about DPC-specific repair mechanisms until the recent identification of a DPC-processing protease in yeast. The existence of a DPC protease in higher eukaryotes is inferred from data in Xenopus laevis egg extracts, but its identity remains elusive. Here we identify the metalloprotease SPRTN as the DPC protease acting in metazoans. Loss of SPRTN results in failure to repair DPCs and hypersensitivity to DPC-inducing agents. SPRTN accomplishes DPC processing through a unique DNA-induced protease activity, which is controlled by several sophisticated regulatory mechanisms. Cellular, biochemical, and structural studies define a DNA switch triggering its protease activity, a ubiquitin switch controlling SPRTN chromatin accessibility, and regulatory autocatalytic cleavage. Our data also provide a molecular explanation on how SPRTN deficiency causes the premature aging and cancer predisposition disorder Ruijs-Aalfs syndrome.

Formation and repair of DNA-protein crosslink damage

“…Consistently, replication-arrested worm larvae depleted of Dvc-1 exhibited severe sensitivity toward formaldehyde exposure [Stingele et al, 2016]. These results suggest that SPRTN is involved in DPC repair.…”

“…Several studies have identified the metalloprotease SPRTN as a DPC-cleaving protease [Lopez-Mosqueda et al, 2016; Stingele et al, 2016; Vaz et al, 2016]. SPRTN is evolutionarily related to Wss1 in budding yeast, and they share similar domains in the C-terminus and other structural features.…”

“…In mammalian cells, knockdown of SPRTN with siRNA resulted in hypersensitivity toward formaldehyde and the persistence of DPCs on DNA [Lopez-Mosqueda et al, 2016; Stingele et al, 2016; Vaz et al, 2016]. Consistently, replication-arrested worm larvae depleted of Dvc-1 exhibited severe sensitivity toward formaldehyde exposure [Stingele et al, 2016].…”

“…SPRTN has several regulatory mechanisms to prevent rampant, uncontrolled proteolysis of proteins not involved in DPC lesions. The catalytic center of the protein is exposed to the solvent, making these regulatory mechanisms especially crucial to avoid excessive proteolysis on non-target proteins [Stingele et al, 2016]. The first of these mechanisms to be triggered involves a ubiquitin switch to control SPRTN activity.…”

“…SPRTN in its monoubiquitinated state does not bind to chromatin; only when a DPC is detected does SPRTN become deubiquitinated, enabling the protein to bind to damaged chromatin sites. Therefore, the amount of SPRTN bound to chromatin is expected to be directly proportional to the abundance of DPC lesions [Stingele et al, 2016]. …”