DNA sense-and-respond protein modules for mammalian cells

We generated synthetic protein components that can detect specific DNA sequences and subsequently trigger a desired intracellular response. These modular sensors exploit the programmability of zinc-finger DNA recognition to drive the intein-mediated splicing of an artificial trans-activator that signals to a genetic circuit containing a given reporter or response gene. We used the sensors to mediate sequence recognition−induced apoptosis as well as to detect and report a viral infection. This work establishes a synthetic biology framework for endowing mammalian cells with sentinel capabilities, which provides a programmable means to cull infected cells. It may also be used to identify positively transduced or transfected cells, isolate recipients of intentional genomic edits and increase the repertoire of inducible parts in synthetic biology.United States. Defense Advanced Research Projects Agency (DARPA-BAA-11-23)Defense Threat Reduction Agency (DTRA) (HDTRA1-14-1-0006)United States. Air Force Office of Scientific Research (FA9550-14-1-0060

A Predictive Model of Intein Insertion Site for Use in the Engineering of Molecular Switches

Inteins are intervening protein domains with self-splicing ability that can be used as molecular switches to control activity of their host protein. Successfully engineering an intein into a host protein requires identifying an insertion site that permits intein insertion and splicing while allowing for proper folding of the mature protein post-splicing. By analyzing sequence and structure based properties of native intein insertion sites we have identified four features that showed significant correlation with the location of the intein insertion sites, and therefore may be useful in predicting insertion sites in other proteins that provide native-like intein function. Three of these properties, the distance to the active site and dimer interface site, the SVM score of the splice site cassette, and the sequence conservation of the site showed statistically significant correlation and strong predictive power, with area under the curve (AUC) values of 0.79, 0.76, and 0.73 respectively, while the distance to secondary structure/loop junction showed significance but with less predictive power (AUC of 0.54). In a case study of 20 insertion sites in the XynB xylanase, two features of native insertion sites showed correlation with the splice sites and demonstrated predictive value in selecting non-native splice sites. Structural modeling of intein insertions at two sites highlighted the role that the insertion site location could play on the ability of the intein to modulate activity of the host protein. These findings can be used to enrich the selection of insertion sites capable of supporting intein splicing and hosting an intein switch

Body appreciation around the world: Measurement invariance of the Body Appreciation Scale-2 (BAS-2) across 65 nations, 40 languages, gender identities, and age

The Body Appreciation Scale-2 (BAS-2) is a widely used measure of a core facet of the positive body image construct. However, extant research concerning measurement invariance of the BAS-2 across a large number of nations remains limited. Here, we utilised the Body Image in Nature (BINS) dataset - with data collected between 2020 and 2022 - to assess measurement invariance of the BAS-2 across 65 nations, 40 languages, gender identities, and age groups. Multi-group confirmatory factor analysis indicated that full scalar invariance was upheld across all nations, languages, gender identities, and age groups, suggesting that the unidimensional BAS-2 model has widespread applicability. There were large differences across nations and languages in latent body appreciation, while differences across gender identities and age groups were negligible-to-small. Additionally, greater body appreciation was significantly associated with higher life satisfaction, being single (versus being married or in a committed relationship), and greater rurality (versus urbanicity). Across a subset of nations where nation-level data were available, greater body appreciation was also significantly associated with greater cultural distance from the United States and greater relative income inequality. These findings suggest that the BAS-2 likely captures a near-universal conceptualisation of the body appreciation construct, which should facilitate further cross-cultural research

A new integrative model for the co-occurrence of non-suicidal self-injury behaviours and eating disorder symptoms

OBJECTIVE: The high co-occurrence of non-suicidal self-injury (NSSI) behaviours and eating disorder (ED) symptoms suggests these conditions share common aetiological processes. We assessed a new integrative model of shared factors for NSSI and ED symptoms, where affect dysregulation, impulsivity, self-esteem, and body dissatisfaction mediated the relationship between insecure attachment and maladaptive schemas and NSSI and ED symptoms. A further aim of the study was to assess whether the model behaved similarly across a clinical eating disorder (ED) and a community sample. METHOD: 123 females with a lifetime ED diagnosis and 531 female individuals from the community completed an online survey, which included measures assessing the variables of interest. A cross-sectional single time point analysis was used. RESULTS: Invariance testing indicated that the model was structurally non-invariant (different across groups). The proposed integrative model was a good fit for the ED group, but for the community sample only a revised model reached an acceptable fit. Both attachment and maladaptive schemas, included early in the model, were implicated in the pathways leading to ED and NSSI symptoms in the ED and community groups. In the community group, impulsivity, a mediator, was a shared predictor for NSSI and bulimic symptoms. No other mediating variables were shared by NSSI and ED symptoms in the two groups. Overall, the proposed model explained slightly more variance for the ED group relative to the community group in drive for thinness (R2 = .57 vs .51) and NSSI (R2 = .29 vs .24) but less variance in bulimic symptoms (R2 = .33 vs .39). CONCLUSION: We conclude that the current model provides only limited support for explaining the comorbidity between NSSI and ED symptoms. It is vital to consider both common (e.g., attachment and maladaptive schemas) and specific factors (e.g., impulsivity) to better understand the pathways that lead to the co-occurrence of NSSI and ED symptoms. A new integrative model assessed whether emotion dysregulation, impulsivity, self-esteem, and body dissatisfaction were mediators in the relationship between insecure attachment and maladaptive beliefs about the world and the self and subsequent eating disorder and self-harm symptoms. A further aim was to assess whether the proposed model differed between a clinical eating disorder and a community sample. All participants were female and included 123 patients with a lifetime eating disorder and 531 individuals from the community. Participating individuals completed an online survey at one timepoint, which included measures assessing the variables of interest. The findings of the current study indicated that the proposed model was a good match for the clinical eating disorder sample, but for the community sample only a revised model yielded acceptable statistical fit. Both insecure attachment and maladaptive beliefs about the world and the self, included early in the model, were indirectly related to eating disorder and self-harm symptoms for both the eating disorder and the community groups. Impulsivity, a mediator, was the only shared predictor for self-harm, and bulimic symptoms in the community group. We conclude that the current model provides only limited support for explaining the comorbidity between self-harming behaviours and disordered eating symptoms

Spatiotemporal control of cell signalling using a light-switchable protein interaction

Genetically-encodable optical reporters, such as Green Fluorescent Protein, have revolutionized the observation and measurement of cellular states. However, the inverse challenge of using light to precisely control cellular behavior has only recently begun to be addressed; semi-synthetic chromophore-tethered receptors1 and naturally-occurring channel rhodopsins have been used to directly perturb neuronal networks2,3. The difficulty of engineering light sensitive proteins remains a significant impediment to the optical control to most cell-biological processes. Here we demonstrate the use of a new genetically-encoded light-control system based on an optimized reversible protein-protein interaction from the phytochrome signaling network of Arabidopsis thaliana. Because protein-protein interactions are one of the most general currencies of cellular information, this system can in principal be generically used to control diverse functions. Here we show that this system can be used to precisely and reversibly translocate target proteins to the membrane with micrometer spatial resolution and second time resolution. We show that light-gated translocation of the upstream activators of rho-family GTPases, which control the actin cytoskeleton, can be used to precisely reshape and direct the cell morphology of mammalian cells. The light-gated protein-protein interaction that has been optimized in this work should be useful for the design of diverse light-programmable reagents, potentially enabling a new generation of perturbative, quantitative experiments in cell biology