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The human toll of disasters extends beyond death, injury and loss. Post-traumatic stress (PTS) can be common among directly exposed individuals, and children are particularly vulnerable. Even children far removed from harm's way report PTS, and media-based exposure may partially account for this phenomenon. In this study, we examine this issue using data from nearly 400 9- to 11-year-old children collected before and after Hurricane Irma, evaluating whether pre-existing neural patterns moderate associations between hurricane experiences and later PTS. The 'dose' of both self-reported objective exposure and media exposure predicted PTS, the latter even among children far from the hurricane. Furthermore, neural responses in brain regions associated with anxiety and stress conferred particular vulnerability. For example, heightened amygdala reactivity to fearful stimuli moderated the association between self-reported media exposure and PTS. Collectively, these findings show that for some youth with measurable vulnerability, consuming extensive disaster-related media may offer an alternative pathway to disaster exposure that transcends geography and objective risk.

COVID-19 presents significant social, economic, and medical challenges. Because COVID-19 has already begun to precipitate huge increases in mental health problems, clinical psychological science must assert a leadership role in guiding a national response to this secondary crisis. In this article, COVID-19 is conceptualized as a unique, compounding, multidimensional stressor that will create a vast need for intervention and necessitate new paradigms for mental health service delivery and training. Urgent challenge areas across developmental periods are discussed, followed by a review of psychological symptoms that likely will increase in prevalence and require innovative solutions in both science and practice. Implications for new research directions, clinical approaches, and policy issues are discussed to highlight the opportunities for clinical psychological science to emerge as an updated, contemporary field capable of addressing the burden of mental illness and distress in the wake of COVID-19 and beyond. (PsycInfo Database Record (c) 2021 APA, all rights reserved).

Assembly line polyketide synthases (PKSs) are large multimodular enzymes responsible for the biosynthesis of diverse antibiotics in bacteria. Structural and mechanistic analysis of these megasynthases can benefit from the discovery of reagents that recognize individual domains or linkers in a site-specific manner. Monoclonal antibodies not only have proven themselves as premier tools in analogous applications but also have the added benefit of constraining the conformational flexibility of their targets in unpredictable but often useful ways. Here we have exploited a library based on the naïve human antibody repertoire to discover a Fab (3A6) that recognizes the terminal thioesterase (TE) domain of the 6-deoxyerythronolide B synthase with high specificity. Biochemical assays were used to verify that 3A6 binding does not inhibit enzyme turnover. The co-crystal structure of the TE-3A6 complex was determined at 2.45 Å resolution, resulting in atomic characterization of this protein-protein recognition mechanism. Fab binding had minimal effects on the structural integrity of the TE. In turn, these insights were used to interrogate via small-angle X-ray scattering the solution-phase conformation of 3A6 complexed to a catalytically competent PKS module and bimodule. Altogether, we have developed a high-affinity monoclonal antibody tool that recognizes the TE domain of the 6-deoxyerythronolide B synthase while maintaining its native function.

Catalytic modules of assembly-line polyketide synthases (PKSs) have previously been observed in two very different conformations-an "extended" architecture and an "arch-shaped" architecture-although the catalytic relevance of neither has been directly established. By the use of a fully human naïve antigen-binding fragment (Fab) library, a high-affinity antibody was identified that bound to the extended conformation of a PKS module, as verified by X-ray crystallography and tandem size-exclusion chromatography-small-angle X-ray scattering (SEC-SAXS). Kinetic analysis proved that this antibody-stabilized module conformation was fully competent for catalysis of intermodular polyketide chain translocation as well as intramodular polyketide chain elongation and functional group modification of a growing polyketide chain. Thus, the extended conformation of a PKS module is fully competent for all of its essential catalytic functions.

The lentiviral protein Viral Infectivity Factor (Vif) counteracts the antiviral effects of host APOBEC3 (A3) proteins and contributes to persistent HIV infection. Vif targets A3 restriction factors for ubiquitination and proteasomal degradation by recruiting them to a multi-protein ubiquitin E3 ligase complex. Here, we describe a degradation-independent mechanism of Vif-mediated antagonism that was revealed through detailed structure-function studies of antibody antigen-binding fragments (Fabs) to the Vif complex. Two Fabs were found to inhibit Vif-mediated A3 neutralization through distinct mechanisms: shielding A3 from ubiquitin transfer and blocking Vif E3 assembly. Combined biochemical, cell biological and structural studies reveal that disruption of Vif E3 assembly inhibited A3 ubiquitination but was not sufficient to restore its packaging into viral particles and antiviral activity. These observations establish that Vif can neutralize A3 family members in a degradation-independent manner. Additionally, this work highlights the potential of Fabs as functional probes, and illuminates how Vif uses a multi-pronged approach involving both degradation dependent and independent mechanisms to suppress A3 innate immunity.

The lentiviral protein Viral Infectivity Factor (Vif) counteracts the antiviral effects of host APOBEC3 (A3) proteins and contributes to persistent HIV infection. Vif targets A3 restriction factors for ubiquitination and proteasomal degradation by recruiting them to a multi-protein ubiquitin E3 ligase complex. Here, we describe a degradation-independent mechanism of Vif-mediated antagonism that was revealed through detailed structure-function studies of antibody antigen-binding fragments (Fabs) to the Vif complex. Two Fabs were found to inhibit Vif-mediated A3 neutralization through distinct mechanisms: shielding A3 from ubiquitin transfer and blocking Vif E3 assembly. Combined biochemical, cell biological and structural studies reveal that disruption of Vif E3 assembly inhibited A3 ubiquitination but was not sufficient to restore its packaging into viral particles and antiviral activity. These observations establish that Vif can neutralize A3 family members in a degradation-independent manner. Additionally, this work highlights the potential of Fabs as functional probes, and illuminates how Vif uses a multi-pronged approach involving both degradation dependent and independent mechanisms to suppress A3 innate immunity.

The immunoproteasome (iP) has been proposed to perform specialized roles in MHC class I antigen presentation, cytokine modulation, and T cell differentiation and has emerged as a promising therapeutic target for autoimmune disorders and cancer. However, divergence in function between the iP and the constitutive proteasome (cP) has been unclear. A global peptide library-based screening strategy revealed that the proteasomes have overlapping but distinct substrate specificities. Differing iP specificity alters the quantity of production of certain MHC I epitopes but does not appear to be preferentially suited for antigen presentation. Furthermore, iP specificity was found to have likely arisen through genetic drift from the ancestral cP. Specificity differences were exploited to develop isoform-selective substrates. Cellular profiling using these substrates revealed that divergence in regulation of the iP balances its relative contribution to proteasome capacity in immune cells, resulting in selective recovery from inhibition. These findings have implications for iP-targeted therapeutic development.

The immunoproteasome (iP) has been proposed to perform specialized roles in MHC class I antigen presentation, cytokine modulation, and T cell differentiation and has emerged as a promising therapeutic target for autoimmune disorders and cancer. However, divergence in function between the iP and the constitutive proteasome (cP) has been unclear. A global peptide library-based screening strategy revealed that the proteasomes have overlapping but distinct substrate specificities. Differing iP specificity alters the quantity of production of certain MHC I epitopes but does not appear to be preferentially suited for antigen presentation. Furthermore, iP specificity was found to have likely arisen through genetic drift from the ancestral cP. Specificity differences were exploited to develop isoform-selective substrates. Cellular profiling using these substrates revealed that divergence in regulation of the iP balances its relative contribution to proteasome capacity in immune cells, resulting in selective recovery from inhibition. These findings have implications for iP-targeted therapeutic development.

Although serine proteases are important mediators of Mycobacterium tuberculosis (Mtb) virulence, there are currently no tools to selectively block or visualize members of this family of enzymes. Selective reporter substrates or activity-based probes (ABPs) could provide a means to monitor infection and response to therapy using imaging methods. Here, we use a combination of substrate selectivity profiling and focused screening to identify optimized reporter substrates and ABPs for the Mtb "Hydrolase important for pathogenesis 1" (Hip1) serine protease. Hip1 is a cell-envelope-associated enzyme with minimal homology to host proteases, making it an ideal target for probe development. We identified substituted 7-amino-4-chloro-3-(2-bromoethoxy)isocoumarins as irreversible inhibitor scaffolds. Furthermore, we used specificity data to generate selective reporter substrates and to further optimize a selective chloroisocoumarin inhibitor. These new reagents are potentially useful in delineating the roles of Hip1 during pathogenesis or as diagnostic imaging tools for specifically monitoring Mtb infections.

Although serine proteases are important mediators of Mycobacterium tuberculosis (Mtb) virulence, there are currently no tools to selectively block or visualize members of this family of enzymes. Selective reporter substrates or activity-based probes (ABPs) could provide a means to monitor infection and response to therapy using imaging methods. Here, we use a combination of substrate selectivity profiling and focused screening to identify optimized reporter substrates and ABPs for the Mtb "Hydrolase important for pathogenesis 1" (Hip1) serine protease. Hip1 is a cell-envelope-associated enzyme with minimal homology to host proteases, making it an ideal target for probe development. We identified substituted 7-amino-4-chloro-3-(2-bromoethoxy)isocoumarins as irreversible inhibitor scaffolds. Furthermore, we used specificity data to generate selective reporter substrates and to further optimize a selective chloroisocoumarin inhibitor. These new reagents are potentially useful in delineating the roles of Hip1 during pathogenesis or as diagnostic imaging tools for specifically monitoring Mtb infections.