Emerging Roles of DNA Glycosylases and the Base Excision Repair Pathway.

The base excision repair (BER) pathway historically has been associated with maintaining genome integrity by eliminating nucleobases with small chemical modifications. In the past several years, however, BER was found to play additional roles in genome maintenance and metabolism, including sequence-specific restriction modification and repair of bulky adducts and interstrand crosslinks. Central to this expanded biological utility are specialized DNA glycosylases - enzymes that selectively excise damaged, modified, or mismatched nucleobases. In this review we discuss the newly identified roles of the BER pathway and examine the structural and mechanistic features of the DNA glycosylases that enable these functions.

Maneuvering through Life with Positivity: Estimating the Effects of Foster Youth's Appraisal on Coping Styles.

Youth in foster care tend to experience a disproportional number of adverse life experiences and demonstrate high rates of emotional and behavioral difficulties. According to the transactional model of stress and coping, how youth appraise their experiences influences the type of coping strategies they use in response to adversity, and these relations are key components to understanding later adjustment. However, few studies have examined potential effects of appraisal on coping for youth in foster care. Furthermore, it is not well understood if or how such interaction may vary across age. To address this gap, this study examined potential age moderation of contemporaneous primary, threat-based appraisal effects on coping in a large sample of 490 youth in foster care (48% female, ages 8 to 18) using a series of statistical models which were capable of detecting very general forms of effect moderation. Results indicated that primary appraisal positively predicted direct and prosocial coping, and negatively predicted asocial coping. The linear effects of appraisal on coping did not vary based on age of the youth. The findings suggest that primary appraisals of life events for youth in foster care does have a unique influence on certain coping styles, suggesting perhaps new directions for research on youth exposed to multiple adversities. To promote wider use of the non-parametric time-varying effect model in R, the analysis syntax is also included in the appendix.

Kinetics, Products, and Brown Carbon Formation by Aqueous-Phase Reactions of Glycolaldehyde with Atmospheric Amines and Ammonium Sulfate.

Glycolaldehyde (GAld) is a C2 water-soluble aldehyde produced during the atmospheric oxidation of isoprene and many other species and is commonly found in cloudwater. Previous work has established that glycolaldehyde evaporates more readily from drying aerosol droplets containing ammonium sulfate (AS) than does glyoxal, methylglyoxal, or hydroxyacetone, which implies that it does not oligomerize as quickly as these other species. Here, we report NMR measurements of glycolaldehyde's aqueous-phase reactions with AS, methylamine, and glycine. Reaction rate constants are smaller than those of respective glyoxal and methylglyoxal reactions in the pH range of 3-6. In follow-up cloud chamber experiments, deliquesced glycine and AS seed particles were found to take up glycolaldehyde and methylamine and form brown carbon. At very high relative humidity, these changes were more than 2 orders of magnitude faster than predicted by our bulk liquid NMR kinetics measurements, suggesting that reactions involving surface-active species at crowded air-water interfaces may play an important role. The high-resolution liquid chromatography-electrospray ionization-mass spectrometric analysis of filter extracts of unprocessed AS + GAld seed particles identified sugar-like C6 and C12 GAld oligomers, including proposed product 3-deoxyglucosone, with and without modification by reactions with ammonia to diimine and imidazole forms. Chamber exposure to methylamine gas, cloud processing, and simulated sunlight increased the incorporation of both ammonia and methylamine into oligomers. Many C4-C16 imidazole derivatives were detected in an extract of chamber-exposed aerosol along with a predominance of N-derivatized C6 and C12 glycolaldehyde oligomers, suggesting that GAld is capable of forming brown carbon SOA.

An autoinhibitory role for the GRF zinc finger domain of DNA glycosylase NEIL3.

The NEIL3 DNA glycosylase maintains genome integrity during replication by excising oxidized bases from single-stranded DNA (ssDNA) and unhooking interstrand cross-links (ICLs) at fork structures. In addition to its N-terminal catalytic glycosylase domain, NEIL3 contains two tandem C-terminal GRF-type zinc fingers that are absent in the other NEIL paralogs. ssDNA binding by the GRF-ZF motifs helps recruit NEIL3 to replication forks converged at an ICL, but the nature of DNA binding and the effect of the GRF-ZF domain on catalysis of base excision and ICL unhooking is unknown. Here, we show that the tandem GRF-ZFs of NEIL3 provide affinity and specificity for DNA that is greater than each individual motif alone. The crystal structure of the GRF domain shows that the tandem ZF motifs adopt a flexible head-to-tail configuration well-suited for binding to multiple ssDNA conformations. Functionally, we establish that the NEIL3 GRF domain inhibits glycosylase activity against monoadducts and ICLs. This autoinhibitory activity contrasts GRF-ZF domains of other DNA-processing enzymes, which typically use ssDNA binding to enhance catalytic activity, and suggests that the C-terminal region of NEIL3 is involved in both DNA damage recruitment and enzymatic regulation.

Too Old to Be Breastfed? Examination of Pre-Healthcare Professionals' Beliefs About, and Emotional and Behavioral Responses toward Extended Breastfeeding.

While breastfeeding seems to be widely accepted in the United States, extended breastfeeding (defined as breastfeeding beyond 12 months of age) tends to be stigmatized. Healthcare professionals are assumed to play a significant role in supporting women who desire to practice extended breastfeeding; however, how healthcare professionals react to extended breastfeeding has not been well understood. This research surveyed 116 healthcare students, who majored in nursing and human medicine, and examined their perceived advantages and disadvantages, emotional responses to, and advice that they would provide to future mothers regarding extended breastfeeding. The results indicated that students responded predominantly with negative emotions and neutral responses to extended breastfeeding, with a small number of participants responding with positive emotions. Many participants believed that it would bring benefits to the child and that it would be burdensome to the mother. Participants displayed a variety of behavioral responses when asked about advice that they would provide to future mothers with whom they will interact in a clinical setting. Practical implications are discussed.

Molecular crowding enhances facilitated diffusion of two human DNA glycosylases.

Intracellular space is at a premium due to the high concentrations of biomolecules and is expected to have a fundamental effect on how large macromolecules move in the cell. Here, we report that crowded solutions promote intramolecular DNA translocation by two human DNA repair glycosylases. The crowding effect increases both the efficiency and average distance of DNA chain translocation by hindering escape of the enzymes to bulk solution. The increased contact time with the DNA chain provides for redundant damage patrolling within individual DNA chains at the expense of slowing the overall rate of damaged base removal from a population of molecules. The significant biological implication is that a crowded cellular environment could influence the mechanism of damage recognition as much as any property of the enzyme or DNA.

Free energy map for the co-oligomerization of formaldehyde and ammonia.

Density functional theory calculations, including Poisson-Boltzmann implicit solvent and free energy corrections, are applied to construct a free energy map of formaldehyde and ammonia co-oligomerization in aqueous solution at pH 7. The stepwise route to forming hexamethylenetetramine (HMTA), the one clearly identified major product in a complex mixture, involves a series of addition reactions of formaldehyde and ammonia coupled with dehydration and cyclization reactions at key steps in the pathway. The free energy map also allows us to propose the possible identity of some major peaks observed by mass spectroscopy in the reaction mixture being the result of stable species not along the pathway to HMTA, in particular those formed by intramolecular condensation of hydroxyl groups to form six-membered rings with ether linkages. Our study complements a baseline free energy map previously worked out for the self-oligomerization of formaldehyde in solution at pH 7 using the same computational protocol and published in this journal (J. Phys. Chem. A 2013, 117, 12658).

Specific exercise patterns generate an epigenetic molecular memory window that drives long-term memory formation and identifies ACVR1C as a bidirectional regulator of memory in mice.

Exercise has beneficial effects on cognition throughout the lifespan. Here, we demonstrate that specific exercise patterns transform insufficient, subthreshold training into long-term memory in mice. Our findings reveal a potential molecular memory window such that subthreshold training within this window enables long-term memory formation. We performed RNA-seq on dorsal hippocampus and identify genes whose expression correlate with conditions in which exercise enables long-term memory formation. Among these genes we found Acvr1c, a member of the TGF ß family. We find that exercise, in any amount, alleviates epigenetic repression at the Acvr1c promoter during consolidation. Additionally, we find that ACVR1C can bidirectionally regulate synaptic plasticity and long-term memory in mice. Furthermore, Acvr1c expression is impaired in the aging human and mouse brain, as well as in the 5xFAD mouse model, and over-expression of Acvr1c enables learning and facilitates plasticity in mice. These data suggest that promoting ACVR1C may protect against cognitive impairment.

Preschool-aged children's hair cortisol and parents' behavior, psychopathology, and stress.

Parental factors, including parenting behavior, parent mental health, and parent stress, are associated with child stress. More recently, studies have shown that these parental factors may also be associated with children's hair cortisol concentration (HCC). HCC is a novel biomarker for chronic stress. HCC indexes cumulative cortisol exposure thereby reflecting longer-term stress reactivity. Although HCC is associated with a range of problems in adults such as depression, anxiety, appraisal of stressful events, and diabetes, studies investigating HCC in children have been inconsistent, with particularly little information about parental factors and HCC. As chronic stress may have long-term physiological and emotional effects on children, and parent-based interventions can reduce these effects, it is important to identify parental factors that relate to children's HCC. The aim of this study was to examine associations between preschool-aged children's physiological stress measured via HCC and mother- and father-reported parenting behavior, psychopathology, and stress. Participants included N = 140 children ages 3-5-years-old and their mothers (n = 140) and fathers (n = 98). Mothers and fathers completed questionnaire measures on their parenting behavior, depressive and anxiety symptoms, and perceived stress. Children's HCC was assessed by processing small hair samples. HCC levels were higher in boys compared to girls, and higher in children of color compared to white children. There was a significant association between children's HCC and fathers' authoritarian parenting. Children's HCC was positively associated with physical coercion, a specific facet of fathers' authoritarian parenting, even after accounting for sex of the child, race/ethnicity of the child, stressful life events, fathers' depression, fathers' anxiety, and fathers' perceived stress. In addition, there was a significant interaction between higher levels of both mothers' and fathers' authoritarian parenting and children's HCC. Children's HCC was not significantly related to mothers' and fathers' anxiety and depression or mothers' and fathers' perceived stress. These findings contribute to the large literature that links harsh and physical parenting practices with problematic outcomes in children.

COVID-19 and the Latinx Community: "Promotoras Represent a Community in Pain".

COVID-19 has disproportionately affected the Latinx community, leading to heightened economic instability and increased mortality/morbidity. Frontline community health workers (promotoras) have played an integral role in serving low-income Latinx immigrant communities, disseminating health information to this vulnerable community while also facing heightened risks to their own health and wellbeing. This study explores the impact of the pandemic on Latinx communities and the promotoras that serve them, examining how the stresses and inequities the pandemic wrought might be mitigated. Promotoras (N = 15, all female) were recruited from a local health agency in Santa Ana, CA and completed a semi-structured interview about their experiences during COVID-19. Qualitative analyses demonstrated that the pandemic substantially affected the daily lives both of community members, via economic challenges, limited access to reliable pandemic-related information, and psychological and social stress, and of promotoras, via changes to the nature of their work and psychological and social stress. Promotoras perceived that these harms might be mitigated by providing for economic and material needs in the community, and that promotoras can be fortified to continue serving the community through self-care and psychosocial healing practices. According to promotoras, the Latinx community needs economic and material resources to overcome COVID-19 related stressors. Additionally, promotoras may benefit from programming to preserve mental and physical health in the wake of new stressors. Lending greater support to promotoras within the agencies in which they are nested may enable them to be more successful in fulfilling their mission and sustaining their own health.

Confirming putative variants at ≤ 5% allele frequency using allele enrichment and Sanger sequencing.

Whole exome sequencing (WES) is used to identify mutations in a patient's tumor DNA that are predictive of tumor behavior, including the likelihood of response or resistance to cancer therapy. WES has a mutation limit of detection (LoD) at variant allele frequencies (VAF) of 5%. Putative mutations called at ≤ 5% VAF are frequently due to sequencing errors, therefore reporting these subclonal mutations incurs risk of significant false positives. Here we performed ~ 1000 × WES on fresh-frozen and formalin-fixed paraffin-embedded (FFPE) tissue biopsy samples from a non-small cell lung cancer patient, and identified 226 putative mutations at between 0.5 and 5% VAF. Each variant was then tested using NuProbe NGSure, to confirm the original WES calls. NGSure utilizes Blocker Displacement Amplification to first enrich the allelic fraction of the mutation and then uses Sanger sequencing to determine mutation identity. Results showed that 52% of the 226 (117) putative variants were disconfirmed, among which 2% (5) putative variants were found to be misidentified in WES. In the 66 cancer-related variants, the disconfirmed rate was 82% (54/66). This data demonstrates Blocker Displacement Amplification allelic enrichment coupled with Sanger sequencing can be used to confirm putative mutations ≤ 5% VAF. By implementing this method, next-generation sequencing can reliably report low-level variants at a high sensitivity, without the cost of high sequencing depth.

Unhooking of an interstrand cross-link at DNA fork structures by the DNA glycosylase NEIL3.

Interstrand DNA-DNA cross-links (ICLs) are generated by endogenous processes, drugs, and environmental toxins. Understanding the cellular pathways by which various ICLs are repaired is critical to understanding their biological effects. Recent studies showed that replication-dependent repair of an ICL derived from the reaction of an abasic (AP) site with an adenine residue (dA) on the opposing strand of duplex DNA proceeds via a novel mechanism in which the DNA glycosylase NEIL3 unhooks the ICL. Here we examined the ability of the glycosylase domain of murine NEIL3 (MmuNEIL3-GD) to unhook dA-AP ICLs. The enzyme selectively unhooks the dA-AP ICL located at the duplex/single-strand junction of splayed duplexes that model the strand-separated DNA at the leading edge of a replication fork. We show that the ability to unhook the dA-AP ICL is a specialized function of NEIL3 as this activity is not observed in other BER enzymes. Importantly, NEIL3 only unhooks the dA-AP ICL when the AP residue is located on what would be the leading template strand of a model replication fork. The same specificity for the leading template strand was observed with a 5,6-dihydrothymine monoadduct, demonstrating that this preference is a general feature of the glycosylase and independent of the type of DNA damage. Overall, the results show that the glycosylase domain of NEIL3, lacking the C-terminal NPL4 and GRF zinc finger motifs, is competent to unhook the dA-AP ICL in splayed substrates and independently enforces important substrate preferences on the repair process.

Conducting a randomized clinical trial of an psychological intervention for parents/caregivers of children with cancer shortly after diagnosis.

OBJECTIVE: To report acceptability, feasibility, and outcome data from a randomized clinical trial (RCT) of a brief intervention for caregivers of children newly diagnosed with cancer. METHOD: Eighty-one families were randomly assigned following collection of baseline data to Intervention or Treatment as Usual (TAU). Recruitment and retention rates and progression through the protocol were tracked. Measures of state anxiety and posttraumatic stress symptoms served as outcomes. RESULTS: Difficulties enrolling participants included a high percentage of newly diagnosed families failing to meet inclusion criteria (40%) and an unexpectedly low participation rate (23%). However, movement through the protocol was generally completed in a timely manner and those completing the intervention provided positive feedback. Outcome data showed no significant differences between the arms of the RCT. CONCLUSIONS: There are many challenges inherent in conducting a RCT shortly after cancer diagnosis. Consideration of alternative research designs and optimal timing for interventions are essential next steps.

Feasibility and preliminary outcomes from a pilot study of a brief psychological intervention for families of children newly diagnosed with cancer.

OBJECTIVE: To report initial feasibility and outcome from a pilot study of a new three-session intervention for caregivers of children newly diagnosed with cancer, Surviving Cancer Competently Intervention Program-Newly Diagnosed (SCCIP-ND). METHOD: Nineteen families (38 caregivers) were randomly assigned to SCCIP-ND or treatment as usual subsequent to learning of their child's illness. The study design included pre- and 2-month postintervention assessments, with state anxiety and posttraumatic stress symptoms as outcomes. Feasibility was based on therapist feedback and supervision, program evaluations, and data from study-tracking procedures. RESULTS: SCCIP-ND appears to be an acceptable intervention that can be used successfully with caregivers over the first few months after diagnosis. Recruitment and retention data document feasibility but also highlight challenges. Preliminary outcome data show changes in the desired direction [e.g., reduced anxiety and parental posttraumatic stress symptoms (PTSS)]. CONCLUSIONS: The pilot data are supportive of the value and challenges of developing evidence-based family interventions in pediatric psychology.

Improving physical functional status in patients with fibromyalgia: a brief cognitive behavioral intervention.

OBJECTIVE: Sustained improvement in physical functional status was the primary goal of a brief, 6 session cognitive behavioral therapy (CBT) protocol for fibromyalgia (FM). METHODS: One hundred forty-five patients with FM were randomly assigned to either (1) standard medical care that included pharmacological management of symptoms and suggestions for aerobic fitness, or (2) the same standard medical treatment plus 6 sessions of CBT aimed at improving physical functioning. Outcome measures included the Medical Outcome Study Short Form-36 Physical Component Score and McGill ratings of pain. Outcomes were treated dichotomously using a preestablished criterion for clinically significant success based upon the reliability of change index from baseline to one year posttreatment. RESULTS: Twenty-five percent of the patients receiving CBT were able to achieve clinically meaningful levels of longterm improvement in physical functioning, whereas only 12% of the patients receiving standard care achieved the same level of improvement. There were no lasting differences on pain ratings between groups. CONCLUSION: Lasting improvements in physical functioning have been among the most difficult outcomes to obtain in studies of FM. These data suggest that the inclusion of CBT to a standard medical regimen for FM can favorably influence physical functioning in a subset of patients.