Serpin1 of Arabidopsis thaliana is a suicide inhibitor for metacaspase 9

Metacaspases are distant relatives of animal caspases found in plants, fungi and protozoa. We demonstrated previously that two type II metacaspases of Arabidopsis thaliana, AtMC4 and AtMC9 are Arg/Lys-specific cysteine-dependent proteases. We screened a combinatorial tetrapeptide library of 130,321 substrates with AtMC9. Here, we show that AtMC9 is a strict Arg/Lys-specific protease. Based on the position-specific scoring matrix derived from the substrate library results, the tetrapeptide Val-Arg-Pro-Arg was identified as an optimized substrate. AtMC9 had a kcat/KM of 4.6x10(5) M-1 s-1 for Ac-Val-Arg-Pro-Arg-amido-4-methyl-coumarin, representing a more than 10-fold improvement over existing fluorogenic substrates. A yeast two-hybrid screen with catalytically inactive AtMC9 as bait identified a serine protease inhibitor, designated AtSerpin1, which was found to be a potent inhibitor of AtMC9 activity in vitro through cleavage of its reactive center loop and covalent binding to AtMC9. On the basis of the substrate profiling of AtMC9 and confirmation through site-directed mutagenesis, the inhibitory P4-P1 cleavage site of AtSerpin1 was determined to be Ile-Lys-Leu-Arg351. Further mutagenesis of the AtSerpin1 inhibitory cleavage site modulated AtMC9 inhibition positively or negatively. Both AtMC9 and AtSerpin1 were localized in the extracellular space, suggesting an in vivo interaction as well. To our knowledge, this is the first report of plant protease inhibition by a plant serpin.     

Deletion of mFICD AMPylase alters cytokine secretion and affects visual short-term learning in vivo

Fic domain-containing AMP transferases (fic AMPylases) are conserved enzymes that catalyze the covalent transfer of AMP to proteins. This posttranslational modification regulates the function of several proteins, including the ER-resident chaperone Grp78/BiP. Here we introduce a mouse FICD (mFICD) AMPylase knockout mouse model to study fic AMPylase function in vertebrates. We find that mFICD deficiency is well tolerated in unstressed mice. We also show that mFICD-deficient mouse embryonic fibroblasts are depleted of AMPylated proteins. mFICD deletion alters protein synthesis and secretion in splenocytes, including that of IgM, an antibody secreted early during infections, and the proinflammatory cytokine IL-1β, without affecting the unfolded protein response. Finally, we demonstrate that visual nonspatial short-term learning is stronger in old mFICD^−/− mice than in wild-type controls while other measures of cognition, memory, and learning are unaffected. Together, our results suggest a role for mFICD in adaptive immunity and neuronal plasticity in vivo.     

Induced malignant genome reprogramming in somatic cells by testis-specific factors

Germline cell differentiation is controlled by a specific set of genes whose expression is tightly locked into the repressed state in somatic cells. Large-scale epigenome alterations, now evidenced in nearly all cancers, lead to aberrant activation of these normally silenced genes, as attested by the many reports describing the expression of testis-specific factors, known as cancer-testis genes, in various cancer cells. Here, based on the literature, we argue that off-context activity of some of the testis-specific epigenome regulators can reprogram the somatic cell epigenome toward a malignant state by favoring self-renewal and sustaining cell proliferation under stressful conditions, thereby constituting a major oncogenic mechanism.     

Shifts in coastal Antarctic marine microbial communities during and after melt water-related surface stratification

Antarctic coastal waters undergo major physical alterations during summer. Increased temperatures induce sea-ice melting and glacial melt water input, leading to strong stratification of the upper water column. We investigated the composition of micro-eukaryotic and bacterial communities in Ryder Bay, Antarctic Peninsula, during and after summertime melt water stratification, applying community fingerprinting (denaturing gradient gel electrophoresis) and sequencing analysis of partial 18S and 16S rRNA genes. Community fingerprinting of the eukaryotic community revealed two major patterns, coinciding with a period of melt water stratification, followed by a period characterized by regular wind-induced breakdown of surface stratification. During the first stratified period, we observed depth-related differences in eukaryotic fingerprints while differences in bacterial fingerprints were weak. Wind-induced breakdown of the melt water layer caused a shift in the eukaryotic community from an Actinocyclus sp.- to a Thalassiosira sp.-dominated community. In addition, a distinct transition in the bacterial community was found, but with a few days' delay, suggesting a response to the changes in the eukaryotic community rather than to the mixing event itself. Sequence analysis revealed a shift from an Alpha- and Gammaproteobacteria to a Cytophaga-Flavobacterium-Bacteroides-dominated community under mixed conditions. Our results show that melt water stratification and the transition to nonstabilized Antarctic surface waters may have an impact not only on micro-eukaryotic but also bacterial community composition.     

Self-Esteem in Hearing-Impaired Children: The Influence of Communication,Education, and Audiological Characteristics

Objective

Sufficient self-esteem is extremely important for psychosocial functioning. It is hypothesized that hearing-impaired (HI) children have lower levels of self-esteem, because, among other things, they frequently experience lower language and communication skills. Therefore, the aim of this study was to compare HI children''s self-esteem across different domains with those of normal hearing (NH) children and to investigate the influence of communication, type of education, and audiological characteristics.

Methods

This large (N = 252) retrospective, multicenter study consisted of two age- and gender-matched groups: 123 HI children and 129 NH controls (mean age  = 11.8 years). Self-reports were used to measure self-esteem across four domains: perceived social acceptance by peers, perceived parental attention, perceived physical appearance, and global self-esteem.

Results

HI children experienced lower levels of self-esteem regarding peers and parents than NH controls. Particularly HI children who attended special education for the deaf were at risk, even after correcting for their language development and intelligence. Yet, levels of global self-esteem and self-esteem involving physical appearance in HI children equalled those of NH controls. Furthermore, younger age at implantation and longer duration of having cochlear implants (CIs) were related to higher levels of self-esteem.

Conclusion

HI children experience lower levels of self-esteem in the social domains. Yet, due to the heterogeneity of the HI population, there is high variability in levels of self-esteem.

Discussion

Clinicians must always be aware of the risk and protective factors related to self-esteem in order to help individual patients reach their full potential.     

Ovarian Cancer Cell Line Panel (OCCP): Clinical Importance of In Vitro Morphological Subtypes

Epithelial ovarian cancer is a highly heterogeneous disease and remains the most lethal gynaecological malignancy in the Western world. Therapeutic approaches need to account for inter-patient and intra-tumoural heterogeneity and detailed characterization of in vitro models representing the different histological and molecular ovarian cancer subtypes is critical to enable reliable preclinical testing. There are approximately 100 publicly available ovarian cancer cell lines but their cellular and molecular characteristics are largely undescribed. We have characterized 39 ovarian cancer cell lines under uniform conditions for growth characteristics, mRNA/microRNA expression, exon sequencing, drug response for clinically-relevant therapeutics and collated all available information on the original clinical features and site of origin. We tested for statistical associations between the cellular and molecular features of the lines and clinical features. Of the 39 ovarian cancer cell lines, 14 were assigned as high-grade serous, four serous-type, one low-grade serous and 20 non-serous type. Three morphological subtypes: Epithelial (n = 21), Round (n = 7) and Spindle (n = 12) were identified that showed distinct biological and molecular characteristics, including overexpression of cell movement and migration-associated genes in the Spindle subtype. Comparison with the original clinical data showed association of the spindle-like tumours with metastasis, advanced stage, suboptimal debulking and poor prognosis. In addition, the expression profiles of Spindle, Round and Epithelial morphologies clustered with the previously described C1-stromal, C5-mesenchymal and C4 ovarian subtype expression profiles respectively. Comprehensive profiling of 39 ovarian cancer cell lines under controlled, uniform conditions demonstrates clinically relevant cellular and genomic characteristics. This data provides a rational basis for selecting models to develop specific treatment approaches for histological and molecular subtypes of ovarian cancer.     

Myocardial Metastases on 6-[18F] fluoro-L-DOPA PET/CT: A Retrospective Analysis of 116 Serotonin Producing Neuroendocrine Tumour Patients

Purpose

This study evaluates the prevalence of cardiac metastases in patients with serotonin producing neuroendocrine tumours (NET), examined with ¹⁸F-FDOPA PET/CT, and the relationship of these metastases to the presence of carcinoid heart disease (CHD) based on echocardiography.

Background

CHD occurs in patients with serotonin producing NET. The diagnostic method of choice remains echocardiography. The precise prevalence of cardiac metastases is unknown given the limitations of standard technologies. Nuclear medicine modalities have the potential to visualize metastases of NET.

Methods

All patients who underwent ¹⁸F-FDOPA PET/CT because of serotonin producing NET between November 2009 and May 2012 were retrospectively analyzed. The presence of cardiac metastasis was defined as myocardial tracer accumulation higher than the surrounding physiological myocardial uptake. Laboratory tests and transthoracic echocardiography (TTE) results were digitally collected.

Results

116 patients (62 male) underwent ¹⁸F-FDOPA PET/CT, mean age was 61±13 years. TTE was performed in 79 patients. Cardiac metastases were present in 15 patients, of which 10 patients also underwent TTE. One patient had both cardiac metastasis (only on ¹⁸F-FDOPA PET/CT) and echocardiographic signs of CHD. There were no differences in echocardiographic parameters for CHD between patients with and without cardiac metastases. TTE in none of the 79 patients showed cardiac metastases.

Conclusion

The prevalence of cardiac metastases detected with ¹⁸F-FDOPA PET/CT in this study is 13%. ¹⁸F-FDOPA PET/CT can visualize cardiac metastases in serotonin producing NET patients. There appears to be no relationship between the presence of cardiac metastases and TTE parameters of CHD.     

Coactivation during gait as an adaptive behavior after stroke

The aims of the present study were to quantify the impairment in ankle coactivation on the paretic and non-paretic sides of subjects with hemiparesis and to examine the relationship of ankle coactivation with postural instability, motor deficit of the paretic lower extremity and locomotor performance. Electromyography of the medial gastrocnemius (MG) and tibialis anterior (TA) muscles were recorded bilaterally during gait in 30 subjects (62.1±9.9 years) who had suffered a recent stroke (<6 months) as well as on one side of 17 healthy controls (59.3±9.1 years) walking at very slow speed. Ankle muscle coactivation was calculated by dividing the time of overlap between MG and TA signals (threshold of 20 μV) by the duration of the gait phases of interest: stance, swing, first and second double support sub-phases and single support sub-phase. The time spent in single support and the peak plantarflexor moment of force on the paretic side were used to measure, respectively, postural stability and dynamic strength of the paretic plantarflexors. The subjects with hemiparesis demonstrated less coactivation on the paretic side during the single support sub-phase (p<0.01) and more coactivation during first and second double support sub-phases on the non-paretic side (p<0.001) compared to control values. The patients with coactivation patterns that differed the most from controls were the patients with the more severe impairments and disabilities. While the reduced coactivation on the paretic side may contribute to poor postural stability and poor locomotor performance, the presence of excessive coactivation on the non-paretic side when both limbs were in ground contact may be an adaptation to help maintain postural stability during gait.