Dectin-1 functions as a pattern recognition receptor for sensing fungal infection. It has been well-established that Dectin-1 induces innate immune responses through caspase recruitment ...domain-containing protein 9 (CARD9)-mediated NF-κB activation. In this study, we find that CARD9 is dispensable for NF-κB activation induced by Dectin-1 ligands, such as curdlan or Candida albicans yeast. In contrast, we find that CARD9 regulates H-Ras activation by linking Ras-GRF1 to H-Ras, which mediates Dectin-1-induced extracellular signal-regulated protein kinase (ERK) activation and proinflammatory responses when stimulated by their ligands. Mechanistically, Dectin-1 engagement initiates spleen tyrosine kinase (Syk)-dependent Ras-GRF1 phosphorylation, and the phosphorylated Ras-GRF1 recruits and activates H-Ras through forming a complex with CARD9, which leads to activation of ERK downstream. Finally, we show that inhibiting ERK activation significantly accelerates the death of C. albicans-infected mice, and this inhibitory effect is dependent on CARD9. Together, our studies reveal a molecular mechanism by which Dectin-1 induces H-Ras activation that leads to ERK activation for host innate immune responses against fungal infection.
Protein termini are determinants of protein stability. Proteins bearing degradation signals, or degrons, at their amino‐ or carboxyl‐termini are eliminated by the N‐ or C‐degron pathways, ...respectively. We aimed to elucidate the function of C‐degron pathways and to unveil how normal proteomes are exempt from C‐degron pathway‐mediated destruction. Our data reveal that C‐degron pathways remove mislocalized cellular proteins and cleavage products of deubiquitinating enzymes. Furthermore, the C‐degron and N‐degron pathways cooperate in protein removal. Proteome analysis revealed a shortfall in normal proteins targeted by C‐degron pathways, but not of defective proteins, suggesting proteolysis‐based immunity as a constraint for protein evolution/selection. Our work highlights the importance of protein termini for protein quality surveillance, and the relationship between the functional proteome and protein degradation pathways.
SYNOPSIS
Proteins with degradation signals (degron) at their amino‐ or carboxyl‐termini are eliminated by the N‐ or C‐degron pathways, respectively. Proteome‐wide analyses suggest functions of human C‐degrons signals in protein quality surveillance, as well as interplay with N‐degron‐dependent mechanisms.
The Global Protein Stability (GPS) random peptide platform enables context‐independent characterization of degron features.
The activity of C‐degrons is tuned by their surrounding sequences.
Gly/C‐degron shortage is limited to functional eukaryotic proteomes accessible to C‐degron pathways.
C‐degron pathways clear products of proteases and mislocalized cellular proteins.
Mitochondrial localization of MIC19 is dually safeguarded by the N‐ and C‐degron pathways.
Coronaviral diGly‐ending proteins evade C‐degron pathway‐mediated degradation.
Proteome‐wide analyses suggest functions of C‐terminal degradation signals in protein quality surveillance as well as interplay with N‐degron‐dependent mechanisms.
The aim of this study was to examine prospective predictors of suicide events, defined as suicide attempts or emergency interventions to reduce suicide risk, in 119 adolescents admitted to an ...in-patient psychiatric unit for suicidal behaviors and followed naturalistically for 6 months. Method Structured diagnostic interviews and self-report instruments were administered to adolescent participants and their parent(s) to assess demographic variables, history of suicidal behavior, psychiatric disorders, family environment and personality/temperament.
Baseline variables that significantly predicted time to a suicide event during follow-up were Black race, high suicidal ideation in the past month, post-traumatic stress disorder (PTSD), childhood sexual abuse (CSA), borderline personality disorder (BPD), low scores on positive affectivity, and high scores on aggression. In a multivariate Cox regression analysis, only Black race, CSA, positive affect intensity and high aggression scores remained significant.
Our findings suggest the following for adolescent populations: (1) in a very high-risk population, risk factors for future attempts may be more difficult to ascertain and some established risk factors (e.g. past suicide attempt) may not distinguish as well; and (2) cross-cutting constructs (e.g. affective and behavioral dysregulation) that underlie multiple psychiatric disorders may be stronger predictors of recurrent suicide events than psychiatric diagnoses. Our finding with respect to positive affect intensity is novel and may have practical implications for the assessment and treatment of adolescent suicide attempters.
BACKGROUND
Daratumumab (DARA) is a human IgG1κ monoclonal antibody directed against CD38, approved for the treatment of multiple myeloma. As CD38 is expressed on RBCs, DARA can interfere with ...pretransfusion testing. DARA interference can be negated by denaturation of CD38 on RBCs with dithiothreitol (DTT) reagents. Because of this interference in pretransfusion testing, our hospital implemented a notification and testing/transfusion algorithm (NATTA) for pretransfusion testing and RBC product provision for DARA patients. This standardized approach combines DTT‐based testing with selective genotyping and the provision of phenotypically similar RBCs for patients with clinically significant antibodies.
STUDY DESIGN AND METHODS
We evaluated pretransfusion test results and transfusion requirements for 91 DARA patients in an academic medical center over 1 year to determine the incremental cost of pretransfusion testing and RBC selection. The actual costs for the NATTA approach were compared to a theoretical approach using universal genotyping with a provision of phenotypically similar RBC transfusions.
RESULTS
The annual cost of testing related to DARA after NATTA implementation was $535.76 per patient. The simulated annual cost for the alternative genotyping with provision of phenotypically similar RBC transfusions approach was $934.83 per patient.
CONCLUSION
In our entire cohort of DARA patients, a DTT‐based testing algorithm with selective genotyping and provision of phenotypically similar RBCs only for patients with clinically significant antibodies was less expensive than a simulated model of universal genotyping and provision of phenotypically similar RBCs.
BACKGROUND
Current regulations do not require blood collection facilities to ask donors about cigarette smoking, and the prevalence of nicotine and its metabolites in blood products is not well ...established. Although smokers have higher hemoglobin (Hb) levels, smoking may adversely affect the quality of donated red blood cells through higher carboxyhemoglobin (COHb) content and premature hemolysis.
STUDY DESIGN AND METHODS
Red blood cell (RBC) unit segments from 100 unique donors were tested for nicotine and its metabolite cotinine by mass spectrometry and for COHb spectrophotometrically. Outcomes were evaluated retrospectively in adult non‐bleeding patients receiving single RBC units.
RESULTS
Thirteen of 100 RBC segments (13%) were positive for cotinine at levels consistent with current smoking (> 10 ng/mL). The cotinine positive RBCs showed significantly greater COHb content compared to cotinine negative units (median 3.0% vs. 0.8%, p = 0.007). For patients transfused cotinine‐positive units, there was no significant change in their vital signs following transfusion and no transfusion reactions were observed. However, patients transfused cotinine‐positive units showed significantly reduced hematocrit and hemoglobin increments (median +1.2% and +0.4 g/dL) following transfusion compared to patients receiving cotinine negative units (median +3.6% and +1.4 g/dL) (p = 0.014).
CONCLUSION
Thirteen percent of RBC units tested positive for cotinine at levels consistent with active smoking, accordant with the estimated national smoking rate of 15.5%. Cotinine‐positive RBC units had greater COHb content and showed reduced hematocrit and hemoglobin increments following transfusion. These preliminary results should be validated in a larger cohort.
See article on page 2485–2488, in this issue
A cell's ability to recognize and adapt to the physical environment is central to its survival and function, but how mechanical cues are perceived and transduced into intracellular signals remains ...unclear. In mesenchymal stem cells (MSCs), high-magnitude substrate strain (HMS, ≥2%) effectively suppresses adipogenesis via induction of focal adhesion (FA) kinase (FAK)/mTORC2/Akt signaling generated at FAs. Physiologic systems also rely on a persistent barrage of low-level signals to regulate behavior. Exposing MSC to extremely low-magnitude mechanical signals (LMS) suppresses adipocyte formation despite the virtual absence of substrate strain (<0.001%), suggesting that LMS-induced dynamic accelerations can generate force within the cell. Here, we show that MSC response to LMS is enabled through mechanical coupling between the cytoskeleton and the nucleus, in turn activating FAK and Akt signaling followed by FAK-dependent induction of RhoA. While LMS and HMS synergistically regulated FAK activity at the FAs, LMS-induced actin remodeling was concentrated at the perinuclear domain. Preventing nuclear-actin cytoskeleton mechanocoupling by disrupting linker of nucleoskeleton and cytoskeleton (LINC) complexes inhibited these LMS-induced signals as well as prevented LMS repression of adipogenic differentiation, highlighting that LINC connections are critical for sensing LMS. In contrast, FAK activation by HMS was unaffected by LINC decoupling, consistent with signal initiation at the FA mechanosome. These results indicate that the MSC responds to its dynamic physical environment not only with "outside-in" signaling initiated by substrate strain, but vibratory signals enacted through the LINC complex enable matrix independent "inside-inside" signaling.
Selenocysteine (Sec) is translated from the codon UGA, typically a termination signal. Codon duality extends the genetic code; however, the coexistence of two competing UGA-decoding mechanisms ...immediately compromises proteome fidelity. Selenium availability tunes the reassignment of UGA to Sec. We report a CRL2 ubiquitin ligase–mediated protein quality-control system that specifically eliminates truncated proteins that result from reassignment failures. Exposing the peptide immediately N-terminal to Sec, a CRL2 recognition degron, promotes protein degradation. Sec incorporation destroys the degron, protecting read-through proteins from detection by CRL2. Our findings reveal a coupling between directed translation termination and proteolysis-assisted protein quality control, as well as a cellular strategy to cope with fluctuations in organismal selenium intake.
Abstract
Protein complexes are the fundamental units of many biological functions. Despite their many advantages, one major adverse impact of protein complexes is accumulations of unassembled ...subunits that may disrupt other processes or exert cytotoxic effects. Synthesis of excess subunits can be inhibited via negative feedback control or they can be degraded more efficiently than assembled subunits, with this latter being termed cooperative stability. Whereas controlled synthesis of complex subunits has been investigated extensively, how cooperative stability acts in complex formation remains largely unexplored. To fill this knowledge gap, we have built quantitative models of heteromeric complexes with or without cooperative stability and compared their behaviours in the presence of synthesis rate variations. A system displaying cooperative stability is robust against synthesis rate variations as it retains high dimer/monomer ratios across a broad range of parameter configurations. Moreover, cooperative stability can alleviate the constraint of limited supply of a given subunit and makes complex abundance more responsive to unilateral upregulation of another subunit. We also conducted an in silico experiment to comprehensively characterize and compare four types of circuits that incorporate combinations of negative feedback control and cooperative stability in terms of eight systems characteristics pertaining to optimality, robustness and controllability. Intriguingly, though individual circuits prevailed for distinct characteristics, the system with cooperative stability alone achieved the most balanced performance across all characteristics. Our study provides theoretical justification for the contribution of cooperative stability to natural biological systems and represents a guideline for designing synthetic complex formation systems with desirable characteristics.
Background
Thawed Plasma (TP), plasma thawed and refrigerated for up to 5 days, is a commonly transfused plasma product. This pilot study was conducted to determine whether Thawed ...Solvent/Detergent‐treated Plasma stored refrigerated for up to 5‐days post‐thaw (T‐S/D) was as efficacious as TP.
Study Design and Methods
This single institution retrospective cohort analysis evaluated the efficacy of T‐S/D in reversing coagulopathies in comparison to TP. Utilizing the institution's electronic medical records, transfusion data were collected in adult patients who received either TP or T‐S/D. The primary outcome was the incidence of subsequent transfusions within 24 hours after first dose of either type of plasma. Secondary outcomes included the number of blood products transfused within 24 hours of first‐dose plasma, correction of pre‐transfusion coagulation laboratory values, volume transfused, and clinical outcomes.
Results
TP was received by 301 patients and 137 received T‐S/D during the first 32 months post‐implementation of T‐S/D. There was no difference in incidence of subsequent transfusions or number of blood products given. The median pre‐INR of both the TP and T‐S/D cohorts was 1.9, with a similar decrease in INR of 0.2 and 0.3 (p = 0.36), respectively, post plasma transfusion. There was no difference in correction of PT/aPTT, mortality, transfusion reactions, readmission rates, length of stay, or inpatient deep venous thrombosis. The median volume of T‐S/D plasma transfused for the first dose was 126 mL less than TP (p = .0001).
Conclusion
T‐S/D was as efficacious as TP for the treatment of coagulopathies and the reversal of coagulation laboratory values.
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