ABSTRACT
XRCC1 is a molecular scaffold protein that assembles multi-protein complexes involved in DNA single-strand break repair. Here we show that biallelic mutations in the human XRCC1 gene are associated with ocular motor apraxia, axonal neuropathy, and progressive cerebellar ataxia. Cells from a patient with mutations in XRCC1 exhibited not only reduced rates of single-strand break repair but also elevated levels of protein ADP-ribosylation. This latter phenotype is recapitulated in a related syndrome caused by mutations in the XRCC1 partner protein PNKP and implicates hyperactivation of poly(ADP-ribose) polymerase/s as a cause of cerebellar ataxia. Indeed, remarkably, genetic deletion of Parp1 rescued normal cerebellar ADP-ribose levels and reduced the loss of cerebellar neurons and ataxia in Xrcc1-defective mice, identifying a molecular mechanism by which endogenous single-strand breaks trigger neuropathology. Collectively, these data establish the importance of XRCC1 protein complexes for normal neurological function and identify PARP1 as a therapeutic target in DNA strand break repair-defective disease.
Subject(s)
Cerebellar Ataxia/genetics , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Mutation , Poly (ADP-Ribose) Polymerase-1/metabolism , Adenosine Diphosphate Ribose/metabolism , Alleles , Animals , Apraxias/congenital , Apraxias/genetics , Ataxia/genetics , Axons/pathology , Cerebellar Ataxia/pathology , Cerebellum/metabolism , Cerebellum/pathology , Chromatin/metabolism , Cogan Syndrome/genetics , DNA Breaks, Single-Stranded , DNA Repair/genetics , DNA Repair Enzymes/genetics , DNA Repair Enzymes/metabolism , DNA-Binding Proteins/deficiency , Female , Humans , Interneurons/metabolism , Interneurons/pathology , Male , Mice , Pedigree , Phenotype , Phosphotransferases (Alcohol Group Acceptor)/genetics , Phosphotransferases (Alcohol Group Acceptor)/metabolism , Poly (ADP-Ribose) Polymerase-1/deficiency , Poly (ADP-Ribose) Polymerase-1/genetics , X-ray Repair Cross Complementing Protein 1ABSTRACT
Efficient recycling of synaptic vesicles is thought to be critical for sustained information transfer at central terminals. However, the specific contribution that retrieved vesicles make to future transmission events remains unclear. Here we exploit fluorescence and time-stamped electron microscopy to track the functional and positional fate of vesicles endocytosed after readily releasable pool (RRP) stimulation in rat hippocampal synapses. We show that most vesicles are recovered near the active zone but subsequently take up random positions in the cluster, without preferential bias for future use. These vesicles non-selectively queue, advancing towards the release site with further stimulation in an actin-dependent manner. Nonetheless, the small subset of vesicles retrieved recently in the stimulus train persist nearer the active zone and exhibit more privileged use in the next RRP. Our findings reveal heterogeneity in vesicle fate based on nanoscale position and timing rules, providing new insights into the origins of future pool constitution.