The cardiac phosphoprotein phospholemman (PLM) regulates the cardiac sodium pump, activating the pump when phosphorylated and inhibiting it when palmitoylated. Protein palmitoylation, the reversible ...attachment of a 16 carbon fatty acid to a cysteine thiol, is catalyzed by the Asp-His-His-Cys (DHHC) motif-containing palmitoyl acyltransferases. The cell surface palmitoyl acyltransferase DHHC5 regulates a growing number of cellular processes, but relatively few DHHC5 substrates have been identified to date. We examined the expression of DHHC isoforms in ventricular muscle and report that DHHC5 is among the most abundantly expressed DHHCs in the heart and localizes to caveolin-enriched cell surface microdomains. DHHC5 coimmunoprecipitates with PLM in ventricular myocytes and transiently transfected cells. Overexpression and silencing experiments indicate that DHHC5 palmitoylates PLM at two juxtamembrane cysteines, C40 and C42, although C40 is the principal palmitoylation site. PLM interaction with and palmitoylation by DHHC5 is independent of the DHHC5 PSD-95/Discs-large/ZO-1 homology (PDZ) binding motif, but requires a ∼120 amino acid region of the DHHC5 intracellular C-tail immediately after the fourth transmembrane domain. PLM C42A but not PLM C40A inhibits the Na pump, indicating PLM palmitoylation at C40 but not C42 is required for PLM-mediated inhibition of pump activity. In conclusion, we demonstrate an enzyme–substrate relationship for DHHC5 and PLM and describe a means of substrate recruitment not hitherto described for this acyltransferase. We propose that PLM palmitoylation by DHHC5 promotes phospholipid interactions that inhibit the Na pump.
Significance Dynamic palmitoylation at the cell surface by the acyl transferase DHHC5 regulates a plethora of physiological processes, from endocytosis to synaptic plasticity. Here we report that DHHC5 is abundantly expressed in cell surface caveolar microdomains in cardiac muscle, and that the Na pump regulator phospholemman is a substrate for DHHC5. Palmitoylation of phospholemman C40 by DHHC5 reduces Na pump activity. DHHC5 interacts with phospholemman independent of its PSD-95/Discs-large/ZO-1 homology (PDZ) binding motif, via a region of its intracellular carboxyl tail that has not hitherto been implicated in substrate recognition. This suggests that it may be possible to selectively manipulate DHHC5 activity by blocking the recruitment of specific substrates to the active site by the carboxyl tail.
Purpose Bosutinib is a potent dual SRC/ABL kinase inhibitor approved for adults with Philadelphia chromosome-positive chronic myeloid leukemia (CML) resistant and /or intolerant to prior therapy. We ...assessed the efficacy and safety of bosutinib versus imatinib for first-line treatment of chronic-phase CML. Methods In this ongoing, multinational, phase III study, 536 patients with newly diagnosed chronic-phase CML were randomly assigned 1:1 to receive 400 mg of bosutinib once daily (n = 268) or imatinib (n = 268). Per protocol, efficacy was assessed in patients who were Philadelphia chromosome-positive with typical (e13a2/e14a2) transcripts (bosutinib, n = 246; imatinib, n = 241). Patients with Philadelphia chromosome-negative-/ BCR-ABL1-positive status and those with unknown Philadelphia chromosome status and/or atypical BCR-ABL1 transcript type were excluded from this population. Results The major molecular response (MMR) rate at 12 months (primary end point) was significantly higher with bosutinib versus imatinib (47.2% v 36.9%, respectively; P = .02), as was complete cytogenetic response (CCyR) rate by 12 months (77.2% v 66.4%, respectively; P = .0075). Cumulative incidence was favorable with bosutinib (MMR: hazard ratio, 1.34; P = .0173; CCyR: hazard ratio, 1.38; P < .001), with earlier response times. Four patients (1.6%) receiving bosutinib and six patients (2.5%) receiving imatinib experienced disease progression to accelerated/blast phase. Among treated patients, 22.0% of patients receiving bosutinib and 26.8% of patients receiving imatinib discontinued treatment, most commonly for drug-related toxicity (12.7% and 8.7%, respectively). Grade ≥ 3 diarrhea (7.8% v 0.8%) and increased ALT (19.0% v 1.5%) and AST (9.7% v 1.9%) levels were more common with bosutinib. Cardiac and vascular toxicities were uncommon. Conclusion Patients who received bosutinib had significantly higher rates of MMR and CCyR and achieved responses faster than those who received imatinib. Consistent with the known safety profile, GI events and transaminase elevations were more common with bosutinib. Results indicate bosutinib may be an effective first-line treatment for chronic-phase CML.
Oral and intestinal mucositis are debilitating inflammatory diseases observed in cancer patients undergoing chemo-radiotherapy. These are devastating clinical conditions which often lead to treatment ...disruption affecting underlying malignancy management. Although alimentary tract mucositis involves the entire gastrointestinal tract, oral and intestinal mucositis are often studied independently utilizing distinct organ-specific pre-clinical models. This approach has however hindered the development of potentially effective whole-patient treatment strategies. We now characterize a murine model of alimentary tract mucositis using 5-Fluorouracil (5-FU). Mice were given 5-FU intravenously (50 mg/kg) or saline every 48 h for 2 weeks. Post initial injection, mice were monitored clinically for weight loss and diarrhea. The incidence and extent of oral mucositis was assessed macroscopically. Microscopical and histomorphometric analyses of the tongue and intestinal tissues were conducted at 3 interim time points during the experimental period. Repeated 5-FU treatment caused severe oral and intestinal atrophy, including morphological damage, accompanied by body weight loss and mild to moderate diarrhea in up to 77.8% of mice. Oral mucositis was clinically evident throughout the observation period in 88.98% of mice. Toluidine blue staining of the tongue revealed that the ulcer size peaked at day-14. In summary, we have developed a model reproducing the clinical and histologic features of both oral and intestinal mucositis, which may represent a useful in vivo pre-clinical model for the study of chemotherapy-induced alimentary tract mucositis and the development of preventative therapies.
Little is known about clinical features associated with the risk of recurrence in patients with bipolar disorder receiving treatment according to contemporary practice guidelines. The authors looked ...for the features associated with risk of recurrence.
The authors examined prospective data from a cohort of patients with bipolar disorder participating in the multicenter Systematic Treatment Enhancement Program for Bipolar Disorder (STEP-BD) study for up to 24 months. For those who were symptomatic at study entry but subsequently achieved recovery, time to recurrence of mania, hypomania, mixed state, or a depressive episode was examined with Cox regression.
Of 1,469 participants symptomatic at study entry, 858 (58.4%) subsequently achieved recovery. During up to 2 years of follow-up, 416 (48.5%) of these individuals experienced recurrences, with more than twice as many developing depressive episodes (298, 34.7%) as those who developed manic, hypomanic, or mixed episodes (118, 13.8%). The time until 25% of the individuals experienced a depressive episode was 21.4 weeks and until 25% experienced a manic/hypomanic/mixed episode was 85.0 weeks. Residual depressive or manic symptoms at recovery and proportion of days depressed or anxious in the preceding year were significantly associated with shorter time to depressive recurrence. Residual manic symptoms at recovery and proportion of days of elevated mood in the preceding year were significantly associated with shorter time to manic, hypomanic, or mixed episode recurrence.
Recurrence was frequent and associated with the presence of residual mood symptoms at initial recovery. Targeting residual symptoms in maintenance treatment may represent an opportunity to reduce risk of recurrence.
Numerous definitions have been proposed for the diagnosis of myocardial infarction (MI) after coronary revascularization. The universal definition for MI designates post procedural biomarker ...thresholds for defining percutaneous coronary intervention (PCI)-related MI (type 4a) and coronary artery bypass grafting (CABG)-related MI (type 5), which are of uncertain prognostic importance. In addition, for both the MI types, cTn is recommended as the biomarker of choice, the prognostic significance of which is less well validated than CK-MB. Widespread adoption of a MI definition not clearly linked to subsequent adverse events such as mortality or heart failure may have serious consequences for the appropriate assessment of devices and therapies, may affect clinical care pathways, and may result in misinterpretation of physician competence. Rather than using an MI definition sensitive for small degrees of myonecrosis (the occurrence of which, based on contemporary large-scale studies, are unlikely to have important clinical consequences), it is instead recommended that a threshold level of biomarker elevation which has been strongly linked to subsequent adverse events in clinical studies be used to define a "clinically relevant MI." The present document introduces a new definition for "clinically relevant MI" after coronary revascularization (PCI or CABG), which is applicable for use in clinical trials, patient care, and quality outcomes assessment.
Identification of regulators of Toxoplasma gondii bradyzoite development and cyst formation is the most direct way to address the importance of parasite development in long-term persistence and ...reactivation of this parasite. Here we show that a T. gondii gene (named Regulator of Cystogenesis 1; ROCY1) is sufficient for T. gondii bradyzoite formation in vitro and in vivo. ROCY1 encodes an RNA binding protein that has a preference for 3' regulatory regions of hundreds of T. gondii transcripts, and its RNA-binding domains are required to mediate bradyzoite development. Female mice infected with ΔROCY1 parasites have reduced (>90%) cyst burden. While viable parasites can be cultivated from brain tissue for up to 6 months post-infection, chronic brain-resident ΔROCY1 parasites have reduced oral infectivity compared to wild type. Despite clear defects in bradyzoite formation and oral infectivity, ΔROCY1 parasites were able to reactivate with similar timing and magnitude as wild type parasites for up to 5 months post-infection. Therefore while ROCY1 is a critical regulator of the bradyzoite developmental pathway, it is not required for parasite reactivation, raising new questions about the persisting life stage responsible for causing recrudescent disease.
Genetic variation in the FAM13A (Family with Sequence Similarity 13 Member A) locus has been associated with several glycemic and metabolic traits in genome-wide association studies (GWAS). Here, we ...demonstrate that in humans, FAM13A alleles are associated with increased FAM13A expression in subcutaneous adipose tissue (SAT) and an insulin resistance-related phenotype (e.g. higher waist-to-hip ratio and fasting insulin levels, but lower body fat). In human adipocyte models, knockdown of FAM13A in preadipocytes accelerates adipocyte differentiation. In mice, Fam13a knockout (KO) have a lower visceral to subcutaneous fat (VAT/SAT) ratio after high-fat diet challenge, in comparison to their wild-type counterparts. Subcutaneous adipocytes in KO mice show a size distribution shift toward an increased number of smaller adipocytes, along with an improved adipogenic potential. Our results indicate that GWAS-associated variants within the FAM13A locus alter adipose FAM13A expression, which in turn, regulates adipocyte differentiation and contribute to changes in body fat distribution.
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