A Novel Interaction Between RAD23A/B and Y-family DNA Polymerases.

The Y-family DNA polymerases - Pol ι, Pol η, Pol κ and Rev1 - are most well-known for their roles in the DNA damage tolerance pathway of translesion synthesis (TLS). They function to overcome replication barriers by bypassing DNA damage lesions that cannot be normally replicated, allowing replication forks to continue without stalling. In this work, we demonstrate a novel interaction between each Y-family polymerase and the nucleotide excision repair (NER) proteins, RAD23A and RAD23B. We initially focus on the interaction between RAD23A and Pol ι, and through a series of biochemical, cell-based, and structural assays, find that the RAD23A ubiquitin-binding domains (UBA1 and UBA2) interact with separate sites within the Pol ι catalytic domain. While this interaction involves the ubiquitin-binding cleft of UBA2, Pol ι interacts with a distinct surface on UBA1. We further find that mutating or deleting either UBA domain disrupts the RAD23A-Pol ι interaction, demonstrating that both interactions are necessary for stable binding. We also provide evidence that both RAD23 proteins interact with Pol ι in a similar manner, as well as with each of the Y-family polymerases. These results shed light on the interplay between the different functions of the RAD23 proteins and reveal novel binding partners for the Y-family TLS polymerases.

Protein engineering reveals that gasdermin A preferentially targets mitochondrial membranes over the plasma membrane during pyroptosis.

When activated, gasdermin family members are thought to be pore-forming proteins that cause lytic cell death. Despite this, numerous studies have suggested that the threshold for lytic cell death is dependent on which gasdermin family member is activated. Determination of the propensity of various gasdermin family members to cause pyroptosis has been handicapped by the fact that for many of them, the mechanisms and timing of their activation are uncertain. In this article, we exploit the recently discovered exosite-mediated recognition of gasdermin D (GSDMD) by the inflammatory caspases to develop a system that activates gasdermin family members in an efficient and equivalent manner. We leverage this system to show that upon activation, GSDMD and gasdermin A (GSDMA) exhibit differential subcellular localization, differential plasma membrane permeabilization, and differential lytic cell death. While GSDMD localizes rapidly to both the plasma membrane and organelle membranes, GSDMA preferentially localizes to the mitochondria with delayed and diminished accumulation at the plasma membrane. As a consequence of this differential kinetics of subcellular localization, N-terminal GSDMA results in early mitochondrial dysfunction relative to plasma membrane permeabilization. This study thus challenges the assumption that gasdermin family members effect cell death through identical mechanisms and establishes that their activation in their respective tissues of expression likely results in different immunological outcomes.

Network localization of clinical, cognitive, and neuropsychiatric symptoms in Alzheimer's disease.

There is both clinical and neuroanatomical variability at the single-subject level in Alzheimer's disease, complicating our understanding of brain-behaviour relationships and making it challenging to develop neuroimaging biomarkers to track disease severity, progression, and response to treatment. Prior work has shown that both group-level atrophy in clinical dementia syndromes and complex neurological symptoms in patients with focal brain lesions localize to brain networks. Here, we use a new technique termed 'atrophy network mapping' to test the hypothesis that single-subject atrophy maps in patients with a clinical diagnosis of Alzheimer's disease will also localize to syndrome-specific and symptom-specific brain networks. First, we defined single-subject atrophy maps by comparing cortical thickness in each Alzheimer's disease patient versus a group of age-matched, cognitively normal subjects across two independent datasets (total Alzheimer's disease patients = 330). No more than 42% of Alzheimer's disease patients had atrophy at any given location across these datasets. Next, we determined the network of brain regions functionally connected to each Alzheimer's disease patient's location of atrophy using seed-based functional connectivity in a large (n = 1000) normative connectome. Despite the heterogeneity of atrophied regions at the single-subject level, we found that 100% of patients with a clinical diagnosis of Alzheimer's disease had atrophy functionally connected to the same brain regions in the mesial temporal lobe, precuneus cortex, and angular gyrus. Results were specific versus control subjects and replicated across two independent datasets. Finally, we used atrophy network mapping to define symptom-specific networks for impaired memory and delusions, finding that our results matched symptom networks derived from patients with focal brain lesions. Our study supports atrophy network mapping as a method to localize clinical, cognitive, and neuropsychiatric symptoms to brain networks, providing insight into brain-behaviour relationships in patients with dementia.

When Do Patients Return to Physical Activities and Athletics After Scoliosis Surgery?: A Validated Patient Questionnaire Based Study.

STUDY DESIGN: A retrospective chart review with a survey. OBJECTIVES: This study seeks to determine time of return to normal, physical and athletic activities, and delaying factors after all pedicle screw fixation. SUMMARY OF BACKGROUND DATA: Return to athletic activity after posterior spine fusion (PSF) in adolescent idiopathic scoliosis (AIS) is largely dependent on a surgeon's philosophy. Some allow contact and collision sports by 6 and 12 months, while others avoid contact sports for 1 year and never allow collision sports. We have utilized a patient driven self-directed approach. METHODS: The sports activity questionnaire (SAQ) was developed and activities were categorized into normal (school, gym, and backpack), physical (running, bending, and bicycling) and athletics (AAP criteria: noncontact, contact and collision sports). SAQ was validated through the "test-retest" method on 25 patients and retesting after 3 weeks to minimize recall bias. Questions with kappa >0.7 were included. Patient demographics, x-ray measurements, and perioperative details were recorded. RESULTS: Ninety five patients completed the SAQ. By 3 months; 77% (72/93) returned to school, 60% (54/90) to bending, 52% (48/93) to carrying backpacks, 43% (37/87) to running, and 37% (30/81) to gym. By 6 months, 54% (27/50) returned to noncontact sports, and 63% (21/33) to contact sports. 79% and 53% returned to preoperative level of contact and noncontact sports, respectively. Higher body mass index (BMI) was a risk for delayed return (>3 mo) to school and gym (P < 0.05), while fusion below L2 and younger age for running, bending, and carrying backpacks (P < 0.05). In contrast, there was no patient/curve characteristics associated with a delay to sports. Lowest instrumented vertebra (LIV), Lenke types were not risk factors. There was no correction loss, implant failure, or complications. CONCLUSION: Patients return to athletics much earlier than expected; a quarter returned by 3 months, and over half by 6 months. Age and LIV are determinants for return to "physical activity." LEVEL OF EVIDENCE: 3.