DNA triplet repeat expansion and mismatch repair.

DNA mismatch repair is a conserved antimutagenic pathway that maintains genomic stability through rectification of DNA replication errors and attenuation of chromosomal rearrangements. Paradoxically, mutagenic action of mismatch repair has been implicated as a cause of triplet repeat expansions that cause neurological diseases such as Huntington disease and myotonic dystrophy. This mutagenic process requires the mismatch recognition factor MutSß and the MutLα (and/or possibly MutLγ) endonuclease, and is thought to be triggered by the transient formation of unusual DNA structures within the expanded triplet repeat element. This review summarizes the current knowledge of DNA mismatch repair involvement in triplet repeat expansion, which encompasses in vitro biochemical findings, cellular studies, and various in vivo transgenic animal model experiments. We present current mechanistic hypotheses regarding mismatch repair protein function in mediating triplet repeat expansions and discuss potential therapeutic approaches targeting the mismatch repair pathway.

Structures of human exonuclease 1 DNA complexes suggest a unified mechanism for nuclease family.

Human exonuclease 1 (hExo1) plays important roles in DNA repair and recombination processes that maintain genomic integrity. It is a member of the 5' structure-specific nuclease family of exonucleases and endonucleases that includes FEN-1, XPG, and GEN1. We present structures of hExo1 in complex with a DNA substrate, followed by mutagenesis studies, and propose a common mechanism by which this nuclease family recognizes and processes diverse DNA structures. hExo1 induces a sharp bend in the DNA at nicks or gaps. Frayed 5' ends of nicked duplexes resemble flap junctions, unifying the mechanisms of endo- and exonucleolytic processing. Conformational control of a mobile region in the catalytic site suggests a mechanism for allosteric regulation by binding to protein partners. The relative arrangement of substrate binding sites in these enzymes provides an elegant solution to a complex geometrical puzzle of substrate recognition and processing.

The secreted protein Cowpox Virus 14 contributes to viral virulence and immune evasion by engaging Fc-gamma-receptors.

The genome of cowpoxvirus (CPXV) could be considered prototypical for orthopoxviridae (OXPV) since it contains many open reading frames (ORFs) absent or lost in other OPXV, including vaccinia virus (VACV). These additional ORFs are non-essential for growth in vitro but are expected to contribute to the broad host range, virulence and immune evasion characteristics of CPXV. For instance, unlike VACV, CPXV encodes proteins that interfere with T cell stimulation, either directly or by preventing antigen presentation or co-stimulation. When studying the priming of naïve T cells, we discovered that CPXV, but not VACV, encodes a secreted factor that interferes with activation and proliferation of naïve CD8+ and CD4+ T cells, respectively, in response to anti-CD3 antibodies, but not to other stimuli. Deletion mapping revealed that the inhibitory protein is encoded by CPXV14, a small secreted glycoprotein belonging to the poxvirus immune evasion (PIE) family and containing a smallpoxvirus encoded chemokine receptor (SECRET) domain that mediates binding to chemokines. We demonstrate that CPXV14 inhibition of antibody-mediated T cell activation depends on the presence of Fc-gamma receptors (FcγRs) on bystander cells. In vitro, CPXV14 inhibits FcγR-activation by antigen/antibody complexes by binding to FcγRs with high affinity and immobilized CPXV14 can trigger signaling through FcγRs, particularly the inhibitory FcγRIIB. In vivo, CPXV14-deleted virus showed reduced viremia and virulence resulting in reduced weight loss and death compared to wildtype virus whereas both antibody and CD8+ T cell responses were increased in the absence of CPXV14. Furthermore, no impact of CPXV14-deletion on virulence was observed in mice lacking the inhibitory FcγRIIB. Taken together our results suggest that CPXV14 contributes to virulence and immune evasion by binding to host FcγRs.

Interaction of proliferating cell nuclear antigen with PMS2 is required for MutLα activation and function in mismatch repair.

Eukaryotic MutLα (mammalian MLH1-PMS2 heterodimer; MLH1-PMS1 in yeast) functions in early steps of mismatch repair as a latent endonuclease that requires a mismatch, MutSα/ß, and DNA-loaded proliferating cell nuclear antigen (PCNA) for activation. We show here that human PCNA and MutLα interact specifically but weakly in solution to form a complex of approximately 1:1 stoichiometry that depends on PCNA interaction with the C-terminal endonuclease domain of the MutLα PMS2 subunit. Amino acid substitution mutations within a PMS2 C-terminal 721QRLIAP motif attenuate or abolish human MutLα interaction with PCNA, as well as PCNA-dependent activation of MutLα endonuclease, PCNA- and DNA-dependent activation of MutLα ATPase, and MutLα function in in vitro mismatch repair. Amino acid substitution mutations within the corresponding yeast PMS1 motif (723QKLIIP) reduce or abolish mismatch repair in vivo. Coupling of a weak allele within this motif (723AKLIIP) with an exo1Δ null mutation, which individually confer only weak mutator phenotypes, inactivates mismatch repair in the yeast cell.

Perceptions of the impact of chorea on health-related quality of life in Huntington disease (HD): A qualitative analysis of individuals across the HD spectrum, family members, and clinicians.

Chorea, a hallmark symptom of Huntington's disease (HD), is characterized by jerky involuntary movements affecting the whole body that can interfere with daily functioning and impact health-related quality of life (HRQOL). To characterize chorea's impact on everyday functioning and HRQOL and identify patterns of perception and experiences of chorea among patients, caregivers, and providers. Data from focus groups of individuals with manifest HD (n = 8 early-stage HD; n = 16 late-stage HD), individuals at-risk or prodromal HD (n = 16), family HD caregivers (n = 17), and HD clinicians (n = 25). Focus group recordings were transcribed verbatim and analysed via constant comparison to identify meaningful and salient themes of living with chorea. Global themes of chorea's impact identified included: watching for chorea, experiences of stigma, and constraints on independence and relationships. Themes distinct to specific respondent groups included: Vigilance (at risk, prodromal); adaptation to chorea (early-stage); loss of autonomy and social life (late-stage); monitoring engagement (family caregivers) and safety (clinical providers). Living with chorea significantly constrains daily functioning, interactions, and HRQOL across the HD disease spectrum. Addressing these impacts via appropriate management of chorea can potentially enhance functioning, HRQOL, and overall satisfaction for persons with HD and their families.

Physician and patient preferences for dosing options in migraine prevention.

BACKGROUND: Adherence to a therapy, though a key factor for successful treatment, is low among patients with chronic conditions such as migraine. Dose frequency plays a major role in adherence. This study evaluated the impact of having flexible dosing options on acceptance of and adherence to a new migraine preventive therapy class among adults with migraine. METHODS: In this observational study, two 20-min online surveys were completed: one by physicians currently treating adult patients with migraine and the other by adults with migraine. Both surveys presented the participants with three scenarios: 1) only monthly, 2) only quarterly, and 3) both dosing options of the new medication are available. Physicians estimated the proportion of their migraine patients who would receive the new medication in each scenario. Patients were asked about their dosing preference when either or both options are available. Respondents were asked to rate the likelihood of their acceptance of and adherence to the therapy. RESULTS: 400 physicians and 417 US adults with migraine completed the surveys. The availability of both dosing options yielded a significant increase in the proportion of patients expected to receive the new medication. The overall proportion of patients favoring monthly dosing (35%) was similar to the proportion favoring quarterly dosing (40%). Among those who preferred monthly dosing (n = 147), a greater proportion indicated they are more likely to fill the prescription (77% vs 56%, P < 0.05) and remain adherent (80% vs 57%, P < 0.05) when only monthly is available versus when only quarterly is available. Similarly, among those who preferred quarterly dosing (n = 166), a greater proportion indicated they are likely to fill (63% vs 55%, P < 0.05) and remain adherent (62% vs 54%, P < 0.05) when only quarterly is available compared with when only monthly is available. CONCLUSIONS: Physicians anticipated that the proportion of patients to receive the new medication would increase when both dosing options are available. Patients stated that they are more likely to fill the prescription and adhere to the new therapy when their preferred dosing regimen is available.

Evidence for a biphasic mode of respiratory syncytial virus transmission in permissive HEp2 cell monolayers.

BACKGROUND: During respiratory syncytial virus (RSV) infection filamentous virus particles are formed on the cell surface. Although the virus infectivity remains cell-associated, low levels of cell-free virus is detected during advanced infection. It is currently unclear if this cell-free virus infectivity is due to a low-efficiency specific cell-release mechanism, or if it arises due to mechanical breakage following virus-induced cell damage at the advanced stage of infection. Understanding the origin of this cell-free virus is a prerequisite for understanding the mechanism of RSV transmission in permissive cells. In this study we describe a detailed examination of RSV transmission in permissive HEp2 cell monolayers. METHODS: HEp2 cell monolayers were infected with RSV using a multiplicity of infection of 0.0002, and the course of infection monitored over 5 days. The progression of the virus infection within the cell monolayers was performed using bright-field microscopy to visualise the cell monolayer and immunofluorescence microscopy to detect virus-infected cells. The cell-associated and cell-free virus infectivity were determined by virus plaque assay, and the virus-induced cell cytotoxicity determined by measuring cell membrane permeability and cellular DNA fragmentation. RESULTS: At 2 days-post infection (dpi), large clusters of virus-infected cells could be detected indicating localised transmission in the cell monolayer, and during this stage we failed to detect either cell-free virus or cell cytotoxicity. At 3 dpi the presence of much larger infected cell clusters correlated with the begining of virus-induced changes in cell permeability. The presence of cell-free virus correlated with continued increase in cell permeability and cytotoxicity at 4 and 5 dpi. At 5 dpi extensive cell damage, syncytial formation, and increased cellular DNA fragmentation was noted. However, even at 5 dpi the cell-free virus constituted less than 1 % of the total virus infectivity. CONCLUSIONS: Our data supports a model of RSV transmission that initially involves the localised cell-to-cell spread of virus particles within the HEp2 cell monolayer. However, low levels of cell free-virus infectivity was observed at the advanced stages of infection, which correlated with a general loss in cell monolayer integrity due to virus-induced cytotoxicity.

Extrahelical (CAG)/(CTG) triplet repeat elements support proliferating cell nuclear antigen loading and MutLα endonuclease activation.

MutLα endonuclease can be activated on covalently continuous DNA that contains a MutSα- or MutSß-recognizable lesion and a helix perturbation that supports proliferating cell nuclear antigen (PCNA) loading by replication factor C, providing a potential mechanism for triggering mismatch repair on nonreplicating DNA. Because mouse models for somatic expansion of disease-associated (CAG)n/(CTG)n triplet repeat sequences have implicated both MutSß and MutLα and have suggested that expansions can occur in the absence of replication, we have asked whether an extrahelical (CAG)n or (CTG)n element is sufficient to trigger MutLα activation. (CAG)n and (CTG)n extrusions in relaxed closed circular DNA do in fact support MutSß-, replication factor C-, and PCNA-dependent activation of MutLα endonuclease, which can incise either DNA strand. Extrahelical elements of two or three repeat units are the preferred substrates for MutLα activation, and extrusions of this size also serve as moderately effective sites for loading the PCNA clamp. Relaxed heteroduplex DNA containing a two or three-repeat unit extrusion also triggers MutSß- and MutLα-endonuclease-dependent mismatch repair in nuclear extracts of human cells. This reaction occurs without obvious strand bias at about 10% the rate of that observed with otherwise identical nicked heteroduplex DNA. These findings provide a mechanism for initiation of triplet repeat processing in nonreplicating DNA that is consistent with several features of the model of Gomes-Pereira et al. [Gomes-Pereira M, Fortune MT, Ingram L, McAbney JP, Monckton DG (2004) Hum Mol Genet 13(16):1815-1825]. They may also have implications for triplet repeat processing at a replication fork.

Spontaneous Coronary Artery Dissection: Case Series from a Tertiary Centre.

Spontaneous coronary artery dissection (SCAD) is a rare cause of non-atherosclerotic acute coronary syndrome (ACS). As it is more commonly seen in young women, the diagnosis can be missed. Current evidence is based on case reports and retrospective studies with no consensus recommendations on immediate management and long-term follow-up. We present a case series of four patients to outline clinical presentation, prognosis and long-term management of this rare clinical entity.

Coupling of human DNA excision repair and the DNA damage checkpoint in a defined in vitro system.

DNA repair and DNA damage checkpoints work in concert to help maintain genomic integrity. In vivo data suggest that these two global responses to DNA damage are coupled. It has been proposed that the canonical 30 nucleotide single-stranded DNA gap generated by nucleotide excision repair is the signal that activates the ATR-mediated DNA damage checkpoint response and that the signal is enhanced by gap enlargement by EXO1 (exonuclease 1) 5' to 3' exonuclease activity. Here we have used purified core nucleotide excision repair factors (RPA, XPA, XPC, TFIIH, XPG, and XPF-ERCC1), core DNA damage checkpoint proteins (ATR-ATRIP, TopBP1, RPA), and DNA damaged by a UV-mimetic agent to analyze the basic steps of DNA damage checkpoint response in a biochemically defined system. We find that checkpoint signaling as measured by phosphorylation of target proteins by the ATR kinase requires enlargement of the excision gap generated by the excision repair system by the 5' to 3' exonuclease activity of EXO1. We conclude that, in addition to damaged DNA, RPA, XPA, XPC, TFIIH, XPG, XPF-ERCC1, ATR-ATRIP, TopBP1, and EXO1 constitute the minimum essential set of factors for ATR-mediated DNA damage checkpoint response.

Diverse crowds using diverse methods improves the scientific dialectic.

In science, diversity is vital to the development of new knowledge. We agree with Duarte et al. that we need more political diversity in social psychology, but contend that we need more religious diversity and methodological diversity as well. If some diversity is good, more is better (especially in science).

Will the real fundamental difference underlying ideology please stand up?

Negativity bias explains many ideological differences, yet does not explain research such as conservatives' greater life satisfaction. Conservatives live in safer communities, perhaps to escape negative emotions, yet display numerous other community preferences unrelated to negativity. This tendency toward cognitive consistency can explain both these phenomena and many of the phenomena described in the target article.

CD8+ T cell targeting of tumor antigens presented by HLA-E.

The nonpolymorphic major histocompatibility complex E (MHC-E) molecule is up-regulated on many cancer cells, thus contributing to immune evasion by engaging inhibitory NKG2A/CD94 receptors on NK cells and tumor-infiltrating T cells. To investigate whether MHC-E expression by cancer cells can be targeted for MHC-E-restricted T cell control, we immunized rhesus macaques (RM) with rhesus cytomegalovirus (RhCMV) vectors genetically programmed to elicit MHC-E-restricted CD8+ T cells and to express established tumor-associated antigens (TAAs) including prostatic acidic phosphatase (PAP), Wilms tumor-1 protein, or Mesothelin. T cell responses to all three tumor antigens were comparable to viral antigen-specific responses with respect to frequency, duration, phenotype, epitope density, and MHC restriction. Thus, CMV-vectored cancer vaccines can bypass central tolerance by eliciting T cells to noncanonical epitopes. We further demonstrate that PAP-specific, MHC-E-restricted CD8+ T cells from RhCMV/PAP-immunized RM respond to PAP-expressing HLA-E+ prostate cancer cells, suggesting that the HLA-E/NKG2A immune checkpoint can be exploited for CD8+ T cell-based immunotherapies.

PCNA function in the activation and strand direction of MutLα endonuclease in mismatch repair.

MutLα (MLH1-PMS2) is a latent endonuclease that is activated in a mismatch-, MutSα-, proliferating cell nuclear antigen (PCNA)-, replication factor C (RFC)-, and ATP-dependent manner, with nuclease action directed to the heteroduplex strand that contains a preexisting break. RFC depletion experiments and use of linear DNAs indicate that RFC function in endonuclease activation is limited to PCNA loading. Whereas nicked circular heteroduplex DNA is a good substrate for PCNA loading and for endonuclease activation on the incised strand, covalently closed, relaxed circular DNA is a poor substrate for both reactions. However, covalently closed supercoiled or bubble-containing relaxed heteroduplexes, which do support PCNA loading, also support MutLα activation, but in this case cleavage strand bias is largely abolished. Based on these findings we suggest that PCNA has two roles in MutLα function: The clamp is required for endonuclease activation, an effect that apparently involves interaction of the two proteins, and by virtue of its loading orientation, PCNA determines the strand direction of MutLα incision. These results also provide a potential mechanism for activation of mismatch repair on nonreplicating DNA, an effect that may have implications for the somatic phase of triplet repeat expansion.

What is freedom--and does wealth cause it?

The target article's climato-economic theory will benefit by allowing for bidirectional effects and the heterogeneity of types of freedom, in order to more fully capture the coevolution of societal wealth and freedom. We also suggest alternative methods of testing climato-economic theory, such as longitudinal analyses of these countries' histories and micro-level experiments of each of the theory's hypotheses.

Late gene expression-deficient cytomegalovirus vectors elicit conventional T cells that do not protect against SIV.

Rhesus cytomegalovirus-based (RhCMV-based) vaccine vectors induce immune responses that protect ~60% of rhesus macaques (RMs) from SIVmac239 challenge. This efficacy depends on induction of effector memory-based (EM-biased) CD8+ T cells recognizing SIV peptides presented by major histocompatibility complex-E (MHC-E) instead of MHC-Ia. The phenotype, durability, and efficacy of RhCMV/SIV-elicited cellular immune responses were maintained when vector spread was severely reduced by deleting the antihost intrinsic immunity factor phosphoprotein 71 (pp71). Here, we examined the impact of an even more stringent attenuation strategy on vector-induced immune protection against SIV. Fusion of the FK506-binding protein (FKBP) degradation domain to Rh108, the orthologue of the essential human CMV (HCMV) late gene transcription factor UL79, generated RhCMV/SIV vectors that conditionally replicate only when the FK506 analog Shield-1 is present. Despite lacking in vivo dissemination and reduced innate and B cell responses to vaccination, Rh108-deficient 68-1 RhCMV/SIV vectors elicited high-frequency, durable, EM-biased, SIV-specific T cell responses in RhCMV-seropositive RMs at doses of ≥ 1 × 106 PFU. Strikingly, elicited CD8+ T cells exclusively targeted MHC-Ia-restricted epitopes and failed to protect against SIVmac239 challenge. Thus, Rh108-dependent late gene expression is required for both induction of MHC-E-restricted T cells and protection against SIV.

Detection of Suicide Risk Using Vocal Characteristics: Systematic Review.

BACKGROUND: In an age when telehealth services are increasingly being used for forward triage, there is a need for accurate suicide risk detection. Vocal characteristics analyzed using artificial intelligence are now proving capable of detecting suicide risk with accuracies superior to traditional survey-based approaches, suggesting an efficient and economical approach to ensuring ongoing patient safety. OBJECTIVE: This systematic review aimed to identify which vocal characteristics perform best at differentiating between patients with an elevated risk of suicide in comparison with other cohorts and identify the methodological specifications of the systems used to derive each feature and the accuracies of classification that result. METHODS: A search of MEDLINE via Ovid, Scopus, Computers and Applied Science Complete, CADTH, Web of Science, ProQuest Dissertations and Theses A&I, Australian Policy Online, and Mednar was conducted between 1995 and 2020 and updated in 2021. The inclusion criteria were human participants with no language, age, or setting restrictions applied; randomized controlled studies, observational cohort studies, and theses; studies that used some measure of vocal quality; and individuals assessed as being at high risk of suicide compared with other individuals at lower risk using a validated measure of suicide risk. Risk of bias was assessed using the Risk of Bias in Non-randomized Studies tool. A random-effects model meta-analysis was used wherever mean measures of vocal quality were reported. RESULTS: The search yielded 1074 unique citations, of which 30 (2.79%) were screened via full text. A total of 21 studies involving 1734 participants met all inclusion criteria. Most studies (15/21, 71%) sourced participants via either the Vanderbilt II database of recordings (8/21, 38%) or the Silverman and Silverman perceptual study recording database (7/21, 33%). Candidate vocal characteristics that performed best at differentiating between high risk of suicide and comparison cohorts included timing patterns of speech (median accuracy 95%), power spectral density sub-bands (median accuracy 90.3%), and mel-frequency cepstral coefficients (median accuracy 80%). A random-effects meta-analysis was used to compare 22 characteristics nested within 14% (3/21) of the studies, which demonstrated significant standardized mean differences for frequencies within the first and second formants (standardized mean difference ranged between -1.07 and -2.56) and jitter values (standardized mean difference=1.47). In 43% (9/21) of the studies, risk of bias was assessed as moderate, whereas in the remaining studies (12/21, 57%), the risk of bias was assessed as high. CONCLUSIONS: Although several key methodological issues prevailed among the studies reviewed, there is promise in the use of vocal characteristics to detect elevations in suicide risk, particularly in novel settings such as telehealth or conversational agents. TRIAL REGISTRATION: PROSPERO International Prospective Register of Systematic Reviews CRD420200167413; https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42020167413.

Using Voice Biomarkers to Classify Suicide Risk in Adult Telehealth Callers: Retrospective Observational Study.

BACKGROUND: Artificial intelligence has the potential to innovate current practices used to detect the imminent risk of suicide and to address shortcomings in traditional assessment methods. OBJECTIVE: In this paper, we sought to automatically classify short segments (40 milliseconds) of speech according to low versus imminent risk of suicide in a large number (n=281) of telephone calls made to 2 telehealth counselling services in Australia. METHODS: A total of 281 help line telephone call recordings sourced from On The Line, Australia (n=266, 94.7%) and 000 Emergency services, Canberra (n=15, 5.3%) were included in this study. Imminent risk of suicide was coded for when callers affirmed intent, plan, and the availability of means; level of risk was assessed by the responding counsellor and reassessed by a team of clinical researchers using the Columbia Suicide Severity Rating Scale (=5/6). Low risk of suicide was coded for in an absence of intent, plan, and means and via Columbia suicide Severity Scale Ratings (=1/2). Preprocessing involved normalization and pre-emphasis of voice signals, while voice biometrics were extracted using the statistical language r. Candidate predictors were identified using Lasso regression. Each voice biomarker was assessed as a predictor of suicide risk using a generalized additive mixed effects model with splines to account for nonlinearity. Finally, a component-wise gradient boosting model was used to classify each call recording based on precoded suicide risk ratings. RESULTS: A total of 77 imminent-risk calls were compared with 204 low-risk calls. Moreover, 36 voice biomarkers were extracted from each speech frame. Caller sex was a significant moderating factor (ß=-.84, 95% CI -0.85, -0.84; t=6.59, P<.001). Candidate biomarkers were reduced to 11 primary markers, with distinct models developed for men and women. Using leave-one-out cross-validation, ensuring that the speech frames of no single caller featured in both training and test data sets simultaneously, an area under the precision or recall curve of 0.985 was achieved (95% CI 0.97, 1.0). The gamboost classification model correctly classified 469,332/470,032 (99.85%) speech frames. CONCLUSIONS: This study demonstrates an objective, efficient, and economical assessment of imminent suicide risk in an ecologically valid setting with potential applications to real-time assessment and response. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry ACTRN12622000486729; https://www.anzctr.org.au/ACTRN12622000486729.aspx.

Using Vocal Characteristics To Classify Psychological Distress in Adult Helpline Callers: Retrospective Observational Study.

BACKGROUND: Elevated psychological distress has demonstrated impacts on individuals' health. Reliable and efficient ways to detect distress are key to early intervention. Artificial intelligence has the potential to detect states of emotional distress in an accurate, efficient, and timely manner. OBJECTIVE: The aim of this study was to automatically classify short segments of speech obtained from callers to national suicide prevention helpline services according to high versus low psychological distress and using a range of vocal characteristics in combination with machine learning approaches. METHODS: A total of 120 telephone call recordings were initially converted to 16-bit pulse code modulation format. Short variable-length segments of each call were rated on psychological distress using the distress thermometer by the responding counselor and a second team of psychologists (n=6) blinded to the initial ratings. Following this, 24 vocal characteristics were initially extracted from 40-ms speech frames nested within segments within calls. After highly correlated variables were eliminated, 19 remained. Of 19 vocal characteristics, 7 were identified and validated as predictors of psychological distress using a penalized generalized additive mixed effects regression model, accounting for nonlinearity, autocorrelation, and moderation by sex. Speech frames were then grouped using k-means clustering based on the selected vocal characteristics. Finally, component-wise gradient boosting incorporating these clusters was used to classify each speech frame according to high versus low psychological distress. Classification accuracy was confirmed via leave-one-caller-out cross-validation, ensuring that speech segments from individual callers were not used in both the training and test data. RESULTS: The sample comprised 87 female and 33 male callers. From an initial pool of 19 characteristics, 7 vocal characteristics were identified. After grouping speech frames into 2 separate clusters (correlation with sex of caller, Cramer's V =0.02), the component-wise gradient boosting algorithm successfully classified psychological distress to a high level of accuracy, with an area under the receiver operating characteristic curve of 97.39% (95% CI 96.20-98.45) and an area under the precision-recall curve of 97.52 (95% CI 95.71-99.12). Thus, 39,282 of 41,883 (93.39%) speech frames nested within 728 of 754 segments (96.6%) were classified as exhibiting low psychological distress, and 71455 of 75503 (94.64%) speech frames nested within 382 of 423 (90.3%) segments were classified as exhibiting high psychological distress. As the probability of high psychological distress increases, male callers spoke louder, with greater vowel articulation but with greater roughness (subharmonic depth). In contrast, female callers exhibited decreased vocal clarity (entropy), greater proportion of signal noise, higher frequencies, increased breathiness (spectral slope), and increased roughness of speech with increasing psychological distress. Individual caller random effects contributed 68% to risk reduction in the classification algorithm, followed by cluster configuration (23.4%), spectral slope (4.4%), and the 50th percentile frequency (4.2%). CONCLUSIONS: The high level of accuracy achieved suggests possibilities for real-time detection of psychological distress in helpline settings and has potential uses in pre-emptive triage and evaluations of counseling outcomes. TRIAL REGISTRATION: ANZCTR ACTRN12622000486729; https://www.anzctr.org.au/ACTRN12622000486729.aspx.

MutLalpha and proliferating cell nuclear antigen share binding sites on MutSbeta.

MutSbeta (MSH2-MSH3) mediates repair of insertion-deletion heterologies but also triggers triplet repeat expansions that cause neurological diseases. Like other DNA metabolic activities, MutSbeta interacts with proliferating cell nuclear antigen (PCNA) via a conserved motif (QXX(L/I)XXFF). We demonstrate that MutSbeta-PCNA complex formation occurs with an affinity of approximately 0.1 microM and a preferred stoichiometry of 1:1. However, up to 20% of complexes are multivalent under conditions where MutSbeta is in molar excess over PCNA. Conformational studies indicate that the two proteins associate in an end-to-end fashion in solution. Surprisingly, mutation of the PCNA-binding motif of MutSbeta not only abolishes PCNA binding, but unlike MutSalpha, also dramatically attenuates MutSbeta-MutLalpha interaction, MutLalpha endonuclease activation, and bidirectional mismatch repair. As predicted by these findings, PCNA competes with MutLalpha for binding to MutSbeta, an effect that is blocked by the cell cycle regulator p21(CIP1). We propose that MutSbeta-MutLalpha interaction is mediated in part by residues ((L/I)SRFF) embedded within the MSH3 PCNA-binding motif. To our knowledge this is the first case where residues important for PCNA binding also mediate interaction with a second protein. These findings also indicate that MutSbeta- and MutSalpha-initiated repair events differ in fundamental ways.