Mechanochemical consequences of myopathy-linked mutations in Tpm2.2 on striated muscle contractility.

Tropomyosin (Tpm) is an actin-binding protein central to muscle contraction regulation. The Tpm sequence consists of periodic repeats corresponding to seven actin-binding sites, further divided in two functionally distinct halves. To clarify the importance of the first and second halves of the actin-binding periods in regulating the interaction of myosin with actin, we introduced hypercontractile mutations D20H, E181K located in the N-terminal halves of periods 1 and 5 and hypocontractile mutations E41K, N202K located in the C-terminal halves of periods 1 and 5 of the skeletal muscle Tpm isoform Tpm2.2. Wild-type and mutant Tpms displayed similar actin-binding properties, however, as revealed by FRET experiments, the hypercontractile mutations affected the binding geometry and orientation of Tpm2.2 on actin, causing a stimulation of myosin motor performance. Contrary, the hypocontractile mutations led to an inhibition of both, actin activation of the myosin ATPase and motor activity, that was more pronounced than with wild-type Tpm2.2. Single ATP turnover kinetic experiments indicate that the introduced mutations have opposite effects on product release kinetics. While the hypercontractile Tpm2.2 mutants accelerated product release, the hypocontractile mutants decelerated product release from myosin, thus having either an activating or inhibitory influence on myosin motor performance, which agrees with the muscle disease phenotypes caused by these mutations.

Protein interactome of 3',5'-cAMP reveals its role in regulating the actin cytoskeleton.

Identification of protein interactors is ideally suited for the functional characterization of small molecules. 3',5'-cAMP is an evolutionary ancient signaling metabolite largely uncharacterized in plants. To tap into the physiological roles of 3',5'-cAMP, we used a chemo-proteomics approach, thermal proteome profiling (TPP), for the unbiased identification of 3',5'-cAMP protein targets. TPP measures shifts in the protein thermal stability upon ligand binding. Comprehensive proteomics analysis yielded a list of 51 proteins significantly altered in their thermal stability upon incubation with 3',5'-cAMP. The list contained metabolic enzymes, ribosomal subunits, translation initiation factors, and proteins associated with the regulation of plant growth such as CELL DIVISION CYCLE 48. To functionally validate obtained results, we focused on the role of 3',5'-cAMP in regulating the actin cytoskeleton suggested by the presence of actin among the 51 identified proteins. 3',5'-cAMP supplementation affected actin organization by inducing actin-bundling. Consistent with these results, the increase in 3',5'-cAMP levels, obtained either by feeding or by chemical modulation of 3',5'-cAMP metabolism, was sufficient to partially rescue the short hypocotyl phenotype of the actin2 actin7 mutant, severely compromised in actin level. The observed rescue was specific to 3',5'-cAMP, as demonstrated using a positional isomer 2',3'-cAMP, and true for the nanomolar 3',5'-cAMP concentrations reported for plant cells. In vitro characterization of the 3',5'-cAMP-actin pairing argues against a direct interaction between actin and 3',5'-cAMP. Alternative mechanisms by which 3',5'-cAMP would affect actin dynamics, such as by interfering with calcium signaling, are discussed. In summary, our work provides a specific resource, 3',5'-cAMP interactome, as well as functional insight into 3',5'-cAMP-mediated regulation in plants.

Distinct roles of adipose triglyceride lipase and hormone-sensitive lipase in the catabolism of triacylglycerol estolides.

Branched esters of palmitic acid and hydroxy stearic acid are antiinflammatory and antidiabetic lipokines that belong to a family of fatty acid (FA) esters of hydroxy fatty acids (HFAs) called FAHFAs. FAHFAs themselves belong to oligomeric FA esters, known as estolides. Glycerol-bound FAHFAs in triacylglycerols (TAGs), named TAG estolides, serve as metabolite reservoir of FAHFAs mobilized by lipases upon demand. Here, we characterized the involvement of two major metabolic lipases, adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL), in TAG estolide and FAHFA degradation. We synthesized a library of 20 TAG estolide isomers with FAHFAs varying in branching position, chain length, saturation grade, and position on the glycerol backbone and developed an in silico mass spectra library of all predicted catabolic intermediates. We found that ATGL alone or coactivated by comparative gene identification-58 efficiently liberated FAHFAs from TAG estolides with a preference for more compact substrates where the estolide branching point is located near the glycerol ester bond. ATGL was further involved in transesterification and remodeling reactions leading to the formation of TAG estolides with alternative acyl compositions. HSL represented a much more potent estolide bond hydrolase for both TAG estolides and free FAHFAs. FAHFA and TAG estolide accumulation in white adipose tissue of mice lacking HSL argued for a functional role of HSL in estolide catabolism in vivo. Our data show that ATGL and HSL participate in the metabolism of estolides and TAG estolides in distinct manners and are likely to affect the lipokine function of FAHFAs.

Longitudinal Associations of Psychiatric Risk Factors with Non-psychiatric Hospitalization in a Large Cohort of People Living with HIV in New York City.

Hospitalizations among people living with HIV (PLWH) are frequent and costly. This study examined the association between psychiatric, HIV-related, and demographic factors and hospitalization rates among PLWH using data from the Einstein-Rockefeller-City University of New York Center for AIDS Research Clinical Cohort Database. Of the 10,215 PLWH included in the sample, 45% had at least one non-psychiatric hospitalization between 2009 and 2018, with significant risk factors including prior psychiatric outpatient visits, depression, or alcohol-related disorder diagnoses, female sex, older age, CD4 count < 500 cells/uL, and detectable viral load. Additionally, 14% had an HIV-related hospitalization, with significant risk factors including prior psychiatric outpatient visits, alcohol- and substance-related disorder diagnoses, female sex, older age, CD4 count < 500 cells/uL, and detectable viral load. The study emphasizes the need for tailored interventions, including integrated treatment and comprehensive case management, for PLWH with comorbid psychiatric disorders, women, and older adults.

RESUMEN: Las hospitalizaciones son frecuentes y costosas entre las personas que viven con VIH (PVVIH). Este estudio examinó la asociación entre factores psiquiátricos, relacionados con el VIH y demográficos, y las tasas de hospitalización en 10,215 PVVIH. Entre 2009 y 2018, el 45% de los pacientes tuvieron al menos una hospitalización no psiquiátrica. Los factores de riesgo significativos incluyeron más visitas previas a la consulta psiquiátrica ambulatoria, diagnóstico previo de depresión o trastorno relacionado con el alcohol, sexo femenino, edad avanzada, conteo de células CD4 < 500 células/uL, y carga viral detectable. De las 10,215 PVVIH, el 14% tuvo una hospitalización relacionada con el VIH. Los resultados destacan la necesidad urgente de apoyo dirigido a PVVIH con trastornos psiquiátricos comorbilidades, y para mujeres y adultos mayores que viven con VIH.

[Little frog in the mouth: a visual diagnosis?] / Fröschlein im Mund: eine Blickdiagnose?

Ranula, from the Latin "little frog", is a retention cyst filled with saliva in the oral cavity. Simple ranulas most commonly affect the sublingual gland and typically present as a hemispherical bluish cyst on the floor of the mouth, making it a visual diagnosis. A 7-year-old girl presented with a swelling on the underside of the tongue, an uncommon location for a ranula that made diagnostic assignment difficult. The optimal treatment of a ranula is still controversial in the literature. Many authors favor surgery as the treatment of choice. Our case shows that a watch and wait approach with simple mechanical pressure on the cyst can be sufficient.

Correction: Franz et al. Unraveling a Force-Generating Allosteric Pathway of Actomyosin Communication Associated with ADP and Pi Release. Int. J. Mol. Sci. 2021, 22, 104.

The author wishes to make the following correction to this paper [...].

Single-molecule analysis reveals that regulatory light chains fine-tune skeletal myosin II function.

Myosin II is the main force-generating motor during muscle contraction. Myosin II exists as different isoforms that are involved in diverse physiological functions. One outstanding question is whether the myosin heavy chain (MHC) isoforms alone account for these distinct physiological properties. Unique sets of essential and regulatory light chains (RLCs) are known to assemble with specific MHCs, raising the intriguing possibility that light chains contribute to specialized myosin functions. Here, we asked whether different RLCs contribute to this functional diversification. To this end, we generated chimeric motors by reconstituting the MHC fast isoform (MyHC-IId) and slow isoform (MHC-I) with different light-chain variants. As a result of the RLC swapping, actin filament sliding velocity increased by ∼10-fold for the slow myosin and decreased by >3-fold for the fast myosin. Results from ensemble molecule solution kinetics and single-molecule optical trapping measurements provided in-depth insights into altered chemo-mechanical properties of the myosin motors that affect the sliding speed. Notably, we found that the mechanical output of both slow and fast myosins is sensitive to the RLC isoform. We therefore propose that RLCs are crucial for fine-tuning the myosin function.

Profilin2a-phosphorylation as a regulatory mechanism for actin dynamics.

Profilin is a major regulator of actin dynamics in multiple specific processes localized in different cellular compartments. This specificity is not only meditated by its binding to actin but also its interaction with phospholipids such as phosphatidylinositol (4,5)-bisphosphate (PIP2 ) at the membrane and a plethora of proteins containing poly-L-proline (PLP) stretches. These interactions are fine-tuned by posttranslational modifications such as phosphorylation. Several phospho-sites have already been identified for profilin1, the ubiquitously expressed isoform. However, little is known about the phosphorylation of profilin2a. Profilin2a is a neuronal isoform important for synapse function. Here, we identified several putative profilin2a phospho-sites in silico and tested recombinant phospho-mimetics with regard to their actin-, PLP-, and PIP2 -binding properties. Moreover, we assessed their impact on actin dynamics employing a pyrene-actin polymerization assay. Results indicate that distinct phospho-sites modulate specific profilin2a functions. We could identify a molecular switch site at serine residue 71 which completely abrogated actin binding-as well as other sites important for fine-tuning of different functions, for example, tyrosine 29 for PLP binding. Our findings suggest that differential profilin2a phosphorylation is a sensitive mechanism for regulating its neuronal functions. Moreover, the dysregulation of profilin2a phosphorylation may contribute to neurodegeneration.

Positive and negative uses of social media among adolescents hospitalized for suicidal behavior.

INTRODUCTION: There is public concern about potential associations between adolescent social media/smartphone use and risk for suicide. However, no prior studies leverage qualitative methods to explore the experiences of adolescents currently at-risk for suicide. METHODS: This study examined social technology use from the perspectives of adolescents (n = 30; Mage = 16.1 years) currently hospitalized for a recent suicide attempt or severe ideation. We conducted in-depth interviews and coded transcripts using thematic analysis. We had three research questions: What (1) negative and (2) positive experiences do suicidal adolescents report related to their use of social media/smartphones? (3) How do adolescents describe their disconnection from these technologies use during inpatient hospitalization and views on a subsequent return to digital connectivity after discharge? RESULTS AND CONCLUSIONS: Results reveal both positive and negative social technology uses, with most participants reporting mixed (positive and negative) experiences. Negatives/risks included trouble regulating use, stress related to social media metrics, encounters with "triggering" content, hostility and meanness, self-denigrating comparisons, and burdensome friendship expectations. Positives/benefits included social connection, social support, affect-enhancing content, shared interests, and resources for mental health and coping. Overall, the documented risks and benefits of social technology use correspond with established (offline) risk and protective factors for suicidal thoughts and behaviors. Participants generally valued the break from social technologies during hospitalization, and also viewed them as integral to social re-entry and identified related concerns. Future studies should test well-being focused 'digital hygiene' interventions for maximizing potential benefits and minimizing potential harms of social technologies for at-risk adolescents.

Symmetry Effects in Photoinduced Electron Transfer in Chlorin-Quinone Dyads: Adiabatic Suppression in the Marcus Inverted Region.

In donor-acceptor dyads undergoing photoinduced electron transfer (PET), a direction or pathway for electron movement is usually dictated by the redox properties and the separation distance between the donor and acceptor subunits, while the effect of symmetry is less recognized. We have designed and synthesized two isomeric donor-acceptor assemblies in which electronic coupling between donor and acceptor is altered by the orbital symmetry control with the reorganization energy and charge transfer exothermicity being kept unchanged. Analysis of the optical absorption and luminescence spectra, supported by the DFT and TD-DFT calculations, showed that PET in these assemblies corresponds to the Marcus inverted region (MIR) and has larger rate for isomer with weaker electronic coupling. This surprising observation provides the first experimental evidence for theoretically predicted adiabatic suppression of PET in MIR, which unambiguously controlled solely by symmetry.

Unraveling a Force-Generating Allosteric Pathway of Actomyosin Communication Associated with ADP and Pi Release.

The actomyosin system generates mechanical work with the execution of the power stroke, an ATP-driven, two-step rotational swing of the myosin-neck that occurs post ATP hydrolysis during the transition from weakly to strongly actin-bound myosin states concomitant with Pi release and prior to ADP dissociation. The activating role of actin on product release and force generation is well documented; however, the communication paths associated with weak-to-strong transitions are poorly characterized. With the aid of mutant analyses based on kinetic investigations and simulations, we identified the W-helix as an important hub coupling the structural changes of switch elements during ATP hydrolysis to temporally controlled interactions with actin that are passed to the central transducer and converter. Disturbing the W-helix/transducer pathway increased actin-activated ATP turnover and reduced motor performance as a consequence of prolonged duration of the strongly actin-attached states. Actin-triggered Pi release was accelerated, while ADP release considerably decelerated, both limiting maximum ATPase, thus transforming myosin-2 into a high-duty-ratio motor. This kinetic signature of the mutant allowed us to define the fractional occupancies of intermediate states during the ATPase cycle providing evidence that myosin populates a cleft-closure state of strong actin interaction during the weak-to-strong transition with bound hydrolysis products before accomplishing the power stroke.

Structural and Computational Insights into a Blebbistatin-Bound Myosin•ADP Complex with Characteristics of an ADP-Release Conformation along the Two-Step Myosin Power Stoke.

The motor protein myosin drives a wide range of cellular and muscular functions by generating directed movement and force, fueled through adenosine triphosphate (ATP) hydrolysis. Release of the hydrolysis product adenosine diphosphate (ADP) is a fundamental and regulatory process during force production. However, details about the molecular mechanism accompanying ADP release are scarce due to the lack of representative structures. Here we solved a novel blebbistatin-bound myosin conformation with critical structural elements in positions between the myosin pre-power stroke and rigor states. ADP in this structure is repositioned towards the surface by the phosphate-sensing P-loop, and stabilized in a partially unbound conformation via a salt-bridge between Arg131 and Glu187. A 5 Å rotation separates the mechanical converter in this conformation from the rigor position. The crystallized myosin structure thus resembles a conformation towards the end of the two-step power stroke, associated with ADP release. Computationally reconstructing ADP release from myosin by means of molecular dynamics simulations further supported the existence of an equivalent conformation along the power stroke that shows the same major characteristics in the myosin motor domain as the resolved blebbistatin-bound myosin-II·ADP crystal structure, and identified a communication hub centered on Arg232 that mediates chemomechanical energy transduction.

Annual Research Review: Suicide among youth - epidemiology, (potential) etiology, and treatment.

BACKGROUND: Suicide is a leading cause of death and a complex clinical outcome. Here, we summarize the current state of research pertaining to suicidal thoughts and behaviors in youth. We review their definitions/measurement and phenomenology, epidemiology, potential etiological mechanisms, and psychological treatment and prevention efforts. RESULTS: We identify key patterns and gaps in knowledge that should guide future work. Regarding epidemiology, the prevalence of suicidal thoughts and behaviors among youth varies across countries and sociodemographic populations. Despite this, studies are rarely conducted cross-nationally and do not uniformly account for high-risk populations. Regarding etiology, the majority of risk factors have been identified within the realm of environmental and psychological factors (notably negative affect-related processes), and most frequently using self-report measures. Little research has spanned across additional units of analyses including behavior, physiology, molecules, cells, and genes. Finally, there has been growing evidence in support of select psychotherapeutic treatment and prevention strategies, and preliminary evidence for technology-based interventions. CONCLUSIONS: There is much work to be done to better understand suicidal thoughts and behaviors among youth. We strongly encourage future research to: (1) continue improving the conceptualization and operationalization of suicidal thoughts and behaviors; (2) improve etiological understanding by focusing on individual (preferably malleable) mechanisms; (3) improve etiological understanding also by integrating findings across multiple units of analyses and developing short-term prediction models; (4) demonstrate greater developmental sensitivity overall; and (5) account for diverse high-risk populations via sampling and reporting of sample characteristics. These serve as initial steps to improve the scientific approach, knowledge base, and ultimately prevention of suicidal thoughts and behaviors among youth.

vlPFC-vmPFC-Amygdala Interactions Underlie Age-Related Differences in Cognitive Regulation of Emotion.

Emotion regulation is a critical life skill that develops throughout childhood and adolescence. Despite this development in emotional processes, little is known about how the underlying brain systems develop with age. This study examined emotion regulation in 112 individuals (aged 6-23 years) as they viewed aversive and neutral images using a reappraisal task. On "reappraisal" trials, participants were instructed to view the images as distant, a strategy that has been previously shown to reduce negative affect. On "reactivity" trials, participants were instructed to view the images without regulating emotions to assess baseline emotional responding. During reappraisal, age predicted less negative affect, reduced amygdala responses and inverse coupling between the ventromedial prefrontal cortex (vmPFC) and amygdala. Moreover, left ventrolateral prefrontal (vlPFC) recruitment mediated the relationship between increasing age and diminishing amygdala responses. This negative vlPFC-amygdala association was stronger for individuals with inverse coupling between the amygdala and vmPFC. These data provide evidence that vmPFC-amygdala connectivity facilitates vlPFC-related amygdala modulation across development.

Curbing craving: behavioral and brain evidence that children regulate craving when instructed to do so but have higher baseline craving than adults.

Although one third of children and adolescents are overweight or obese, developmental changes in food craving and the ability to regulate craving remain poorly understood. We addressed this knowledge gap by examining behavioral and neural responses to images of appetizing unhealthy foods in individuals ages 6 through 23 years. On close trials (assessing unregulated craving), participants focused on a pictured food's appetitive features. On far trials (assessing effortful regulation), participants focused on a food's visual features and imagined that it was farther away. Across conditions, older age predicted less craving, less striatal recruitment, greater prefrontal activity, and stronger frontostriatal coupling. When effortfully regulating their responses to the images, all participants reported less craving and exhibited greater recruitment of lateral prefrontal cortex and less recruitment of ventromedial prefrontal cortex. Greater body mass predicted less regulation-related prefrontal activity, particularly among children. These results suggest that children experience stronger craving than adults but can also effectively regulate craving. Moreover, the mechanisms underlying regulation may differ for heavy and lean children.

Switch-2 determines Mg2+ADP-release kinetics and fine-tunes the duty ratio of Dictyostelium class-1 myosins.

Though myosins share a structurally conserved motor domain, single amino acid variations of active site elements, including the P-loop, switch-1 and switch-2, which act as nucleotide sensors, can substantially determine the kinetic signature of a myosin, i.e., to either perform fast movement or enable long-range transport and tension generation. Switch-2 essentially contributes to the ATP hydrolysis reaction and determines product release. With few exceptions, class-1 myosin harbor a tyrosine in the switch-2 consensus sequence DIYGFE, at a position where class-2 myosins and a selection of myosins from other classes have a substitution. Here, we addressed the role of the tyrosine in switch-2 of class-1 myosins as potential determinant of the duty ratio. We generated constitutively active motor domain constructs of two class-1 myosins from the social amoeba Dictyostelium discoideum, namely, Myo1E, a high duty ratio myosin and Myo1B, a low duty ratio myosin. In Myo1E we introduced mutation Y388F and in Myo1B mutation F387Y. The detailed functional characterization by steady-state and transient kinetic experiments, combined with in vitro motility and landing assays revealed an almost reciprocal relationship of a number of critical kinetic parameters and equilibrium constants between wild-type and mutants that dictate the lifetime of the strongly actin-attached states of myosin. The Y-to-F mutation increased the duty ratio of Moy1B by almost one order of magnitude, while the introduction of the phenylalanine in switch-2 of Myo1E transformed the myosin into a low duty ratio motor. These data together with structural considerations propose a role of switch-2 in fine-tuning ADP release through a mechanism, where the class-specific tyrosine together with surrounding residues contributes to the coordination of Mg2+ and ADP. Our results highlight the importance of conserved switch-2 residues in class-1 myosins for efficient chemo-mechanical coupling, revealing that switch-2 is important to adjust the duty ratio of the amoeboid class-1 myosins for performing movement, transport or gating functions.

Examining tradeoffs between cognitive effort and relief among adults with self-injurious behavior.

BACKGROUND: People engage in nonsuicidal self-injury (NSSI) to reduce negative affect, but it is not clear why they engage in this harmful type of behavior instead of using healthier strategies. The primary goal of this study was to evaluate whether people choose NSSI to reduce negative affect because they perceive it to be less cognitively costly than other available strategies. METHOD: In experiment one, 43 adults completed a novel, relief-based effort discounting task designed to index preferences about exerting cognitive effort to achieve relief. In experiment two, 149 adults, 52 % with a history of NSSI, completed our effort discounting task. RESULTS: Our main results suggest that people will accept less relief from an aversive experience if doing so requires expending less effort, i.e. they demonstrate effort discounting in the context of decisions about relief. We also found and that effort discounting is stronger among those with a history of NSSI, but this association became nonsignificant when simultaneously accounting for other conditions associated with aberrant effort tradeoffs. LIMITATIONS: The use of a control group without NSSI or other potentially harmful relief-seeking behaviors limits our ability to draw specific conclusions about NSSI. The ecological validity of our task was limited by a modestly effective affect manipulation, and because participants made hypothetical choices. CONCLUSIONS: This study demonstrates that preferences about exerting cognitive effort may be a barrier to using healthier affect regulation strategies. Further, the preference not to exert cognitive effort, though present in NSSI, is likely not unique to NSSI. Instead, effort discounting may be a transdiagnostic mechanism promoting an array of harmful relief-seeking behaviors.

Racial and Ethnic Disparities in COVID-19-Related Stressor Exposure and Adverse Mental Health Outcomes Among Health Care Workers.

OBJECTIVE: Racial and ethnic disparities in exposure to COVID-19-related stressors, pandemic-related distress, and adverse mental health outcomes were assessed among health care workers in the Bronx, New York, during the first wave of the pandemic. METHODS: The authors analyzed survey data from 992 health care workers using adjusted logistic regression models to assess differential prevalence of outcomes by race/ethnicity and their interactions. RESULTS: Compared with their White colleagues, Latinx, Black, Asian, and multiracial/other health care workers reported significantly higher exposure to multiple COVID-19-related stressors: redeployment, fear of being sick, lack of autonomy at work, and inadequate access to personal protective equipment. Endorsing a greater number of COVID-19-related stressors was associated with pandemic-related distress in all groups and with adverse mental health outcomes in some groups; it was not related to hazardous alcohol use in any of the groups. These associations were not significantly different between racial and ethnic groups. Latinx health care workers had significantly higher probabilities of pandemic-related distress and posttraumatic stress than White colleagues. Despite greater exposure to COVID-19-related stressors, Black, Asian, and multiracial/other health care workers had the same, if not lower, prevalence of adverse mental health outcomes. Conversely, White health care workers had a higher adjusted prevalence of moderate to severe anxiety compared with Asian colleagues and greater hazardous alcohol use compared with all other groups. CONCLUSIONS: Health care workers from racial and ethnic minority groups reported increased exposure to COVID-19-related stressors, suggestive of structural racism in the health care workforce. These results underscore the need for increased support for health care workers and interventions aimed at mitigating disparities in vocational exposure to risk and stress.