POLO Kinase Regulates the Drosophila Centromere Cohesion Protein MEI-S332

AbstractAccurate segregation of chromosomes is critical to ensure that each daughter cell receives the full genetic complement. Maintenance of cohesion between sister chromatids, especially at centromeres, is required to segregate chromosomes precisely during mitosis and meiosis. The Drosophila protein MEI-S332, the founding member of a conserved protein family, is essential in meiosis for maintaining cohesion at centromeres until sister chromatids separate at the metaphase II/anaphase II transition. MEI-S332 localizes onto centromeres in prometaphase of mitosis or meiosis I, remaining until sister chromatids segregate. We elucidated a mechanism for controlling release of MEI-S332 from centromeres via phosphorylation by POLO kinase. We demonstrate that POLO antagonizes MEI-S332 cohesive function and that full POLO activity is needed to remove MEI-S332 from centromeres, yet this delocalization is not required for sister chromatid separation. POLO phosphorylates MEI-S332 in vitro, POLO and MEI-S332 bind each other, and mutation of POLO binding sites prevents MEI-S332 dissociation from centromeres

Assessment of the characteristic of nutrients, total metals, and fecal coliform in Sibu Laut River, Sarawak, Malaysia

The concentrations of nutrients (nitrogen and phosphorus), total metals, and fecal coliform (FC) coupling with chlorophyll-a (chl-a), 5-day biochemical oxygen demand (BOD5) and other general environmental parameters were evaluated at the sub-surface and near-bottom water columns of 13 stations in the Sibu Laut River during low and high slack waters. The results indicated that inorganic nitrogen (mainly nitrate) was the primary form of nitrogen whereas organic phosphorus was the major form of phosphorus. The abundance of total heavy metals in Sibu Laut River and its tributaries was in the order of Pb\Cu\Zn\Cd. Fecal coliform concentration was relatively low along Sibu Laut River. The shrimp farm effluents contributed a substantial amount of chl-a, BOD5, nutrients, and FC to the receiving creek except for total metals. Nevertheless, the influence was merely noticeable in the intake creek and amended rapidly along Selang Sibu River and brought minimal effects on the Sibu Laut River. Besides, the domestic sewage effluents from villages nearby also contributed a substantial amount of pollutants

IMI – industry guidelines and ethical considerations for myopia control report

PURPOSE. To discuss guidelines and ethical considerations associated with the development and prescription of treatments intended for myopia control (MC). METHODS. Critical review of published papers and guidance documents was undertaken, with a view to carefully considering the ethical standards associated with the investigation, development, registration, marketing, prescription, and use of MC treatments. RESULTS. The roles and responsibilities of regulatory bodies, manufacturers, academics, eye care practitioners, and patients in the use of MC treatments are explored. Particular attention is given to the ethical considerations for deciding whether to implement a MC strategy and how to implement this within a clinical trial or practice setting. Finally, the responsibilities in marketing, support, and education required to transfer required knowledge and skills to eye care practitioners and academics are discussed. CONCLUSIONS. Undertaking MC treatment in minors creates an ethical challenge for a wide variety of stakeholders. Regulatory bodies, manufacturers, academics, and clinicians all share an ethical responsibility to ensure that the products used for MC are safe and efficacious and that patients understand the benefits and potential risks of such products. This International Myopia Institute report highlights these ethical challenges and provides stakeholders with recommendations and guidelines in the development, financial support, prescribing, and advertising of such treatments.</p

Biochemical Characterization of the Human Cyclin-dependent Protein Kinase Activating Kinase: IDENTIFICATION OF p35 AS A NOVEL REGULATORY SUBUNIT

The activation of cyclin-dependent protein kinases (Cdks) is dependent upon site-specific phosphorylation and dephosphorylation reactions, as well as positive and negative regulatory subunits. The human Cdk-activating protein kinase (Cak1) is itself a Cdc2-related cyclin-dependent protein kinase that associates with cyclin H. The present study utilized specific anti-Cak1 antibodies and immunoaffinity chromatography to identify additional Cak1-associated proteins and potential target substrates. Immunoprecipitation of metabolically labeled human osteosarcoma cells revealed a number of Cak1-associated proteins, including p95, p37 (cyclin H), and a 35-kDa protein that was further characterized herein. Microsequence analysis obtained after limited proteolysis revealed peptide fragments that are similar, but not identical to, human and yeast cyclins, thus identifying p35 as a cyclin-like regulatory subunit. The greatest sequence similarity of human p35 is with Mcs2, a yeast cyclin that is essential for cell cycle progression. Immunoaffinity chromatography performed under nondenaturing conditions afforded the isolation of enzymatically active Cak1 from cell lysates, enabling studies of kinase autophosphorylation and comparative substrate utilization. Immunoaffinity-purified Cak1 phosphorylated monomeric Cdc2 and Cdk2, but not Cdk4; the phosphorylation of both Cdc2 and Cdk2 were increased in the presence of recombinant cyclin A. These studies indicate that the Cak1 catalytic subunit, like Cdc2 and Cdk2, associates with multiple regulatory partners and suggests that subunit composition may be an important determinant of this multifunctional enzyme

The complement system and human autoimmune diseases

Genetic deficiencies of early components of the classical complement activation pathway (especially C1q, r, s, and C4) are the strongest monogenic causal factors for the prototypic autoimmune disease systemic lupus erythematosus (SLE), but their prevalence is extremely rare. In contrast, isotype genetic deficiency of C4A and acquired deficiency of C1q by autoantibodies are frequent among patients with SLE. Here we review the genetic basis of complement deficiencies in autoimmune disease, discuss the complex genetic diversity seen in complement C4 and its association with autoimmune disease, provide guidance as to when clinicians should suspect and test for complement deficiencies, and outline the current understanding of the mechanisms relating complement deficiencies to autoimmunity. We focus primarily on SLE, as the role of complement in SLE is well-established, but will also discuss other informative diseases such as inflammatory arthritis and myositis

Versican G3 Domain Modulates Breast Cancer Cell Apoptosis: A Mechanism for Breast Cancer Cell Response to Chemotherapy and EGFR Therapy

Overexpression of EGFR and versican has been reported in association with breast cancers. Considered oncogenic, these molecules may be attractive therapeutic targets. Possessing anti-apoptotic and drug resistant properties, overexpression of these molecules is accompanied by selective sensitization to the process of apoptosis. In this study, we exogenously expressed a versican G3 construct in breast cancer cell lines and analyzed the effects of G3 on cell viability in fetal bovine serum free conditioned media and evaluated the effects of apoptotic agent C2-ceramide, and chemotherapeutic agents including Docetaxel, Doxorubicin, and Epirubicin. Versican G3 domain enhanced tumor cell resistance to apoptosis when cultured in serum free medium, Doxorubicin, or Epirubicin by up-regulating pERK and GSK-3β (S9P). However, it could be prevented by selective EGFR inhibitor AG 1478 and selective MEK inhibitor PD 98059. Both AG 1478 and PD 98059 enhanced expression of pSAPK/JNK, while selective JNK inhibitor SP 600125 enhanced expression of GSK-3β (S9P). Versican G3 promoted cell apoptosis induced by C2-ceramide or Docetaxel by enhancing expression of pSAPK/JNK and decreasing expression of GSK-3β (S9P), an observation blocked by AG 1478 or SP 6000125. Inhibition of endogenous versican expression by siRNA or reduction of versican G3's expression by linking G3 with 3′UTR prevented G3 modulated cell apoptosis. The dual roles of G3 in modulating breast cancer cell resistance to chemotherapeutic agents may in part explain a potential mechanism for breast cancer cell resistance to chemotherapy and EGFR therapy. The apoptotic effects of chemotherapeutics depend upon the activation and balance of down stream signals in the EGFR pathway. GSK-3β (S9P) appears to function as a key checkpoint in this balance of apoptosis and anti-apoptosis. Investigation and potential consideration of targeting GSK-3β (S9P) merits further study

Multiple Kinases Can Phosphorylate the N-Terminal Sequences of Mitochondrial Proteins in Arabidopsis thaliana

Phosphorylation of the transit peptides of nuclear-encoded preprotein is a well-known regulatory process of protein import in plant chloroplasts. In the Arabidopsis Protein Phosphorylation Site Database, 103 out of 802 mitochondrial proteins were found to contain one or more experimentally proven phosphorylation sites in their first 60 amino acid residues. Analysis of the N-terminal sequences of selected mitochondrial preproteins and their homologs from 64 plant species showed high conservation among phosphorylation sites. The ability of kinases from various sources including leaf extract (LE), root extract (RE), wheat germ lysate (WGL), and STY kinases to phosphorylate N-terminal sequences of several respiratory chain proteins were examined by in vitro kinase assays. The three STY kinases were shown to phosphorylate the N-terminal sequences of some proteins we tested but exhibited different specificities. Interestingly, the N-terminal sequences of two mitochondrial ATP synthase beta subunit 1/3 (pF1 beta-1/3) could be phosphorylated by LE and RE but not by STY kinases, suggesting that there are uncharacterized presequence-phosphorylating kinases other than STY kinases present in RE and LE. Mitochondrial import studies showed that the import of RRL-synthesized pF1 beta s was impeded by the treatment of LE, and the addition of a short SSU transit peptide containing a phosphorylatable 14-3-3 binding site could enhance the import of LE-treated pF1 beta s. Our results suggested that the transit peptide of pSSU can compete with the presequences of pF1 beta s for an uncharacterized kinase(s) in leaf. Altogether, our data showed that phosphorylation of transit peptides/presequences are not uncommon for chloroplast-targeted and mitochondria-targeted proteins, albeit possibly differentially regulated

Impacts of sulfide exposure on juvenile Tor tambroides (Bleeker, 1854): behavioral responses and mortality

Construction of hydroelectric reservoirs had been reported to be the cause of increased sulfide levels resulting from the decomposition of organic matter. As more dams are being built, a better understanding of the impact of sulfide on indigenous species is required. In Sarawak, Tor tambroides is a highly valuable and sought after species which is facing declining population. This study aimed to determine the behavioral responses and mortality of juvenile T. tambroides exposed to sulfide. The three exposure experiments were gradual sulfide exposure, gradual sulfide exposure under lowering DO and gradual sulfide exposure under lowering pH. A modified flow-through design was used to expose the juveniles in containers to sulfide of different concentrations. Actual total sulfide in containers was determined according to standard method. During the duration of the experiment, behavioral responses, DO and pH were monitored. Experimental results show that negative controls recorded no behavioral response and no mortality was observed in all control experiments. However, under all sulfide exposure experiments, the juveniles displayed at least one behavioral response in the progression of huddling together, aquatic surface respiration, loss of equilibrium and turning upside down except for the gradual sulfide exposure experiment where no response was observed with the lowest total sulfide concentration tested (82 µg L-1). For all three exposure experiments, faster responses and mortalities were observed when the concentration of sulfide increased. The LC50 at 6th hour of exposure was estimated to be 306 µg/L total sulfide (138 µg L-1 H2S) at 95% confidence level. Sulfide toxicity was found to be highly related to the decreasing DO and pH levels attributable to intensifying toxicity which led to mortality

Determining the Physical Lens Parameters of the Binary Gravitational Microlensing Event MOA-2009-BLG-016

We report the result of the analysis of the light curve of the microlensing event MOA-2009-BLG-016. The light curve is characterized by a short-duration anomaly near the peak and an overall asymmetry. We find that the peak anomaly is due to a binary companion to the primary lens and the asymmetry of the light curve is explained by the parallax effect caused by the acceleration of the observer over the course of the event due to the orbital motion of the Earth around the Sun. In addition, we detect evidence for the effect of the finite size of the source near the peak of the event, which allows us to measure the angular Einstein radius of the lens system. The Einstein radius combined with the microlens parallax allows us to determine the total mass of the lens and the distance to the lens. We identify three distinct classes of degenerate solutions for the binary lens parameters, where two are manifestations of the previously identified degeneracies of close/wide binaries and positive/negative impact parameters, while the third class is caused by the symmetric cycloid shape of the caustic. We find that, for the best-fit solution, the estimated mass of the lower-mass component of the binary is (0.04 +- 0.01) M_sun, implying a brown-dwarf companion. However, there exists a solution that is worse only by \Delta\chi^2 ~ 3 for which the mass of the secondary is above the hydrogen-burning limit. Unfortunately, resolving these two degenerate solutions will be difficult as the relative lens-source proper motions for both are similar and small (~ 1 mas/yr) and thus the lens will remain blended with the source for the next several decades.Comment: 7 pages, 2 tables, and 5 figure