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.

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.

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.

Survival Outcome in True Carcinoma of Unknown Primary (tCUP) with p16 + Cervical Metastasis.

Introduction Age and lymph node ratio have been attributed as independent predictors for survival and recurrence in carcinoma of unknown primary (CUP). Objective The purpose of this study was to analyze the prognostic value of p16 overexpression for CUP in the absence of true primary (TP). Methods The study involved 43 patients who underwent therapeutic lymph node dissection (LND) from 2000 to 2015 after all the diagnostic work up for CUP. Immunohistochemistry for p16 overexpression was performed. Cox proportional hazard regression analysis was used to analyze the prognostic impact on 5-year overall survival (OS) and recurrence-free survival (RFS). Results The male-to-female ratio was 5.1:1, with a median age of 62 years. The clinicopathological data, except for p16 overexpression, did not differ significantly in terms of 5-year OS and RFS. The Cox regression analysis proposed p16 positivity to be an independent prognosticator of regional recurrence-free survival (RRFS) (hazard ratio [HR] 6.180, p = 0.21). The median time to recurrence and death were 10 and 25 months, respectively. Conclusion Cervical metastasis with p16 overexpression is a significant prognostic factor of improved RFS after surgery in CUP. The prognostic significance of lymph node p16 positivity should be further studied.

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.

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.

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.

[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.

Survival Outcome in True Carcinoma of Unknown Primary (tCUP) with p16 + Cervical Metastasis

Abstract Introduction Age and lymph node ratio have been attributed as independent predictors for survival and recurrence in carcinoma of unknown primary (CUP). Objective The purpose of this study was to analyze the prognostic value of p16 overexpression for CUP in the absence of true primary (TP). Methods The study involved 43 patients who underwent therapeutic lymph node dissection (LND) from 2000 to 2015 after all the diagnostic work up for CUP. Immunohistochemistry for p16 overexpression was performed. Cox proportional hazard regression analysis was used to analyze the prognostic impact on 5-year overall survival (OS) and recurrence-free survival (RFS). Results The male-to-female ratio was 5.1:1, with a median age of 62 years. The clinicopathological data, except for p16 overexpression, did not differ significantly in terms of 5-year OS and RFS. The Cox regression analysis proposed p16 positivity to be an independent prognosticator of regional recurrence-free survival (RRFS) (hazard ratio [HR] 6.180, p = 0.21). The median time to recurrence and death were 10 and 25 months, respectively. Conclusion Cervical metastasis with p16 overexpression is a significant prognostic factor of improved RFS after surgery in CUP. The prognostic significance of lymph node p16 positivity should be further studied.

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 [...].

Digital bibliotherapy as a scalable intervention for suicidal thoughts: A randomized controlled trial.

OBJECTIVE: Suicide is a major public health concern in the United States, but few effective and scalable interventions exist to help those with suicidal thoughts. We hypothesized that reading first-person narratives about working through suicidal thoughts would reduce the desire to die among adults and that this effect would be mediated by increased perceived shared experience and optimism. METHOD: Using a randomized waitlist-controlled trial, we tested the effect of digital narrative-based bibliotherapy among 528 adults visiting a social media platform dedicated to providing mental health support. Participants were randomized to either a treatment condition (n = 266), in which they read one suicide narrative per day for 14 days or to a waitlist control condition (n = 262). The primary outcome was a measure of desire to die assessed daily for the 14-day trial period and at 2-week follow-up. RESULTS: Participants in the treatment condition reported lower desire to die than participants in the control condition during the 14-day trial period (ß = -0.26, p = .001) and at 2-week follow-up (t = -2.82, p = .005). Increased perceived shared experience (indirect effect b = -0.55, p < .001) and optimism (indirect effect b = -0.85, p < .001) mediated the effect of treatment on desire to die. CONCLUSIONS: Digital narrative-based bibliotherapy may be an effective intervention for those at risk for suicide, and may work in part by increasing feelings of perceived shared experience and optimism. Future research is needed to test the generalizability of these results to other platforms, groups, and conditions. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Carcinoma of Unknown Primary (CUP) versus CUP Turned to Primary Carcinoma of the Head and Neck-An Analysis of Diagnostic Methods and the Impact of Primary Tumor on Clinical Outcome.

BACKGROUND: The purpose of this study was to analyze the value of different diagnostic methods in detecting the primary site and the impact of primary tumors on the clinical outcome of carcinoma of unknown primary (CUP). METHODS: In this multicenter, retrospective study, 124 patients with true CUP (n = 94) and CUP turned to primary carcinoma (n = 30) were included. Patients with evidence of primary site during the clinical examination were excluded a priori. The diagnostic procedure was comprised of imaging and invasive methods (fine-needle-aspiration, tonsillectomy and panendoscopy). All patients were treated with curative intent. RESULTS: Despite extensive diagnostic workup, the primary site remained unknown in 75.8%. Invasive diagnostic methods showed higher primary detection rates than imaging modalities (15.1% vs. 7.8%). Tonsillectomy and panendoscopy revealed the primary tumor in 14.9% and 15.2% of patients, whereas the detection rates of CT, MRI and FDG-PET-CT were 10.1%, 4.8% and 6.5%, respectively. The occurrence of primary tumors led to a significantly deteriorating 5-year overall survival (p = 0.002) and emerged as survival prognosticator (HR = 2.764, p = 0.003). CONCLUSION: Clinical examination in combination with tonsillectomy and panendoscopy was superior to imaging alone in detecting the primary tumor. When the CUP of patients turned to a primary tumor, clinical outcome was significantly worse than in CUP patients.

Examining implicit positive affect toward suicide among suicidal and nonsuicidal adults and adolescents.

OBJECTIVE: The purpose of this study was to examine implicit affect toward suicide (i.e., how good/bad suicide is perceived). Some people might be more likely to think about/choose suicide because they perceive it as a good option (to gain relief) relative to available alternatives. METHOD: Implicit affect toward suicide among adults (N = 72) and adolescents (N = 174) with and without suicidal thoughts was examined using first-person (FP) perspective suicide pictures in the affect misattribution procedure (AMP). RESULTS: Suicidal adults' implicit positive affect toward suicide was associated with STB variables, such as explicit valence (r = 0.34) and arousal (r = 0.44) ratings of suicide pictures, and implicit affect differentiated groups above and beyond explicit valence ratings. Contrary to our hypothesis, suicidal participants did not display higher implicit positive affect toward suicide than nonsuicidal participants. However, suicidal participants displayed consistent implicit affect toward different suicide pictures, whereas nonsuicidal participants evaluated some pictures as more pleasant than others (ORs = 1.92-2.27). CONCLUSIONS: Implicit affect toward suicide may relate to STB, but stimuli characteristics (e.g., color) likely influence the accuracy of assessment with the AMP and should be a focus of future research involving this and other implicit measures.

Identification and Speed Estimation of a Moving Object in an Indoor Application Based on Visible Light Sensing of Retroreflective Foils.

Identification and sensing are two of the main tasks a wireless sensor node has to perform in an Internet of Things (IoT) environment. Placing active powered nodes on objects is the most usual approach for the fulfillment of these functions. With the expected massive increase of connected things, there are several issues on the horizon that hamper the further deployment of this approach in an energy efficient, sustainable way, like the usage of environmentally hazardous batteries or accumulators, as well as the required electrical energy for their operation. In this work, we propose a novel approach for performing the tasks of identification and sensing, applying visible light sensing (VLS) based on light emitting diode (LED) illumination and utilizing retroreflective foils mounted on a moving object. This low cost hardware is combined with a self-developed, low complex software algorithm with minimal training effort. Our results show that successful identification and sensing of the speed of a moving object can be achieved with a correct estimation rate of 99.92%. The used foils are commercially available and pose no threat to the environment and there is no need for active sensors on the moving object and no requirement of wireless radio frequency communication. All of this is achievable whilst undisturbed illumination is still provided.

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.

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.

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.