Identification of novel interaction between annexin A2 and keratin 17: evidence for reciprocal regulation

Keratins are cytoplasmic intermediate filament proteins providing crucial structural support in epithelial cells. Keratin expression has diagnostic and even prognostic value in disease settings, and recent studies have uncovered modulatory roles for select keratin proteins in signaling pathways regulating cell growth and cell death. Elevated keratin expression in select cancers is correlated with higher expression of EGF receptor (EGFR), whose overexpression and/or mutation give rise to cancer. To explore the role of keratins in oncogenic signaling pathways, we examined the regulation of epithelial growth-associated keratin 17 (K17) in response to EGFR activation. K17 is specifically up-regulated in detergent-soluble fraction upon EGFR activation, and immunofluorescence analysis revealed alterations in K17-containing filaments. Interestingly, we identified AnxA2 as a novel interacting partner of K17, and this interaction is antagonized by EGFR activation. K17 and AnxA2 proteins show reciprocal regulation. Modulating expression of AnxA2 altered K17 stability, and AnxA2 overexpression delays EGFR-mediated change in K17 detergent solubility. Down-regulation of K17 expression, in turn, results in decreased AnxA2 phosphorylation at Tyr-23. These findings uncover a novel interaction involving K17 and AnxA2 and identify AnxA2 as a potential regulator of keratin filaments.     

Stress-induced rise in body temperature is repeatable in free-ranging Eastern chipmunks (<Emphasis Type="Italic">Tamias striatus</Emphasis>)

In response to handling or other acute stressors, most mammals, including humans, experience a temporary rise in body temperature (T _b). Although this stress-induced rise in T _b has been extensively studied on model organisms under controlled environments, individual variation in this interesting phenomenon has not been examined in the field. We investigated the stress-induced rise in T _b in free-ranging eastern chipmunks (Tamias striatus) to determine first if it is repeatable. We predicted that the stress-induced rise in T _b should be positively correlated to factors affecting heat production and heat dissipation, including ambient temperature (T _a), body mass (M _b), and field metabolic rate (FMR). Over two summers, we recorded both T _b within the first minute of handling time (T _b1) and after 5 min of handling time (T _b5) 294 times on 140 individuals. The mean ∆T _b (T _b5 – T _b1) during this short interval was 0.30 ± 0.02°C, confirming that the stress-induced rise in T _b occurs in chipmunks. Consistent differences among individuals accounted for 40% of the total variation in ∆T _b (i.e. the stress-induced rise in T _b is significantly repeatable). We also found that the stress-induced rise in T _b was positively correlated to T _a, M _b, and mass-adjusted FMR. These results confirm that individuals consistently differ in their expression of the stress-induced rise in T _b and that the extent of its expression is affected by factors related to heat production and dissipation. We highlight some research constraints and opportunities related to the integration of this laboratory paradigm into physiological and evolutionary ecology.     

The Mitotic Exit Network and Cdc14 phosphatase initiate cytokinesis by counteracting CDK phosphorylations and blocking polarised growth

Polarisation of the actin cytoskeleton must cease during cytokinesis, to support efficient assembly and contraction of the actomyosin ring at the site of cell division, but the underlying mechanisms are still understood poorly in most species. In budding yeast, the Mitotic Exit Network (MEN) releases Cdc14 phosphatase from the nucleolus during anaphase, leading to the inactivation of mitotic forms of cyclin-dependent kinase (CDK) and the onset of septation, before G1-CDK can be reactivated and drive re-polarisation of the actin cytoskeleton to a new bud. Here, we show that premature inactivation of mitotic CDK, before release of Cdc14, allows G1-CDK to divert the actin cytoskeleton away from the actomyosin ring to a new site of polarised growth, thereby delaying progression through cytokinesis. Our data indicate that cells normally avoid this problem via the MEN-dependent release of Cdc14, which counteracts all classes of CDK-mediated phosphorylations during cytokinesis and blocks polarised growth. The dephosphorylation of CDK targets is therefore central to the mechanism by which the MEN and Cdc14 initiate cytokinesis and block polarised growth during late mitosis.     

Impact of oncogenic driver mutations on feedback between the PI3K and MEK pathways in cancer cells

Inhibition of the PI3K (phosphoinositide 3-kinase)/Akt/mTORC1 (mammalian target of rapamycin complex 1) and Ras/MEK [MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) kinase]/ERK pathways for cancer therapy has been pursued for over a decade with limited success. Emerging data have indicated that only discrete subsets of cancer patients have favourable responses to these inhibitors. This is due to genetic mutations that confer drug insensitivity and compensatory mechanisms. Therefore understanding of the feedback mechanisms that occur with respect to specific genetic mutations may aid identification of novel biomarkers that predict patient response. In the present paper, we show that feedback between the PI3K/Akt/mTORC1 and Ras/MEK/ERK pathways is cell-line-specific and highly dependent on the activating mutation of K-Ras or overexpression c-Met. We found that cell lines exhibited differential signalling and apoptotic responses to PD184352, a specific MEK inhibitor, and PI103, a second-generation class I PI3K inhibitor. We reveal that feedback from the PI3K/Akt/mTORC1 to the Ras/MEK/ERK pathway is present in cancer cells harbouring either K-Ras activating mutations or amplification of c-Met but not the wild-type counterparts. Moreover, we demonstrate that inhibition of protein phosphatase activity by OA (okadaic acid) restored PI103-mediated feedback in wild-type cells. Together, our results demonstrate a novel mechanism for feedback between the PI3K/Akt/mTORC1 and the Ras/MEK/ERK pathways that only occurs in K-Ras mutant and c-Met amplified cells but not the isogenic wild-type cells through a mechanism that may involve inhibition of a specific endogenous phosphatase(s) activity. We conclude that monitoring K-Ras and c-Met status are important biomarkers for determining the efficacy of PI103 and other PI3K/Akt inhibitors in cancer therapy.     

John Featherstone Witty

Plant and Soil - Multiple functions in the human body are affected by zinc deficiency. Here the etiology, assessment, health consequences, and intervention strategies for human zinc deficiency are...     

CD109, a TGF-β co-receptor, attenuates extracellular matrix production in scleroderma skin fibroblasts

Hemophagocytic lymphohistiocytosis (HLH) is a rare life-threatening disease of severe hyperinflammation caused by uncontrolled proliferation of activated lymphocytes and macrophages secreting high amounts of inflammatory cytokines. It is a frequent manifestation in patients with predisposing genetic defects, but can occur secondary to various infectious, malignant, and autoimmune triggers in patients without a known genetic predisposition. Clinical hallmarks are prolonged fever, cytopenias, hepatosplenomegaly, and neurological symptoms, but atypical variants presenting with signs of chronic immunodeficiency are increasingly recognized. Impaired secretion of perforin is a key feature in several genetic forms of the disease, but not required for disease pathogenesis. Despite progress in diagnostics and therapy, mortality of patients with severe HLH is still above 40%. Reference treatment is an etoposide-based protocol, but new approaches are currently explored. Key for a favorable prognosis is the rapid identification of an underlying genetic cause, which has been facilitated by recent immunological and genetic advances. In patients with predisposing genetic disease, hematopoietic stem cell transplantation is performed increasingly with reduced intensity conditioning regimes. Current research aims at a better understanding of disease pathogenesis and evaluation of more targeted approaches to therapy, including anti-cytokine antibodies and gene therapy.     

The challenges of implementing pathogen control strategies for fishes used in biomedical research

Over the past several decades, a number of fish species, including the zebrafish, medaka, and platyfish/swordtail, have become important models for human health and disease. Despite the increasing prevalence of these and other fish species in research, methods for health maintenance and the management of diseases in laboratory populations of these animals are underdeveloped. There is a growing realization that this trend must change, especially as the use of these species expands beyond developmental biology and more towards experimental applications where the presence of underlying disease may affect the physiology animals used in experiments and potentially compromise research results. Therefore, there is a critical need to develop, improve, and implement strategies for managing health and disease in aquatic research facilities. The purpose of this review is to report the proceedings of a workshop entitled "Animal Health and Disease Management in Research Animals" that was recently held at the 5th Aquatic Animal Models for Human Disease in September 2010 at Corvallis, Oregon to discuss the challenges involved with moving the field forward on this front.     

Direct maximization of protein identifications from tandem mass spectra

The goal of many shotgun proteomics experiments is to determine the protein complement of a complex biological mixture. For many mixtures, most methodological approaches fall significantly short of this goal. Existing solutions to this problem typically subdivide the task into two stages: first identifying a collection of peptides with a low false discovery rate and then inferring from the peptides a corresponding set of proteins. In contrast, we formulate the protein identification problem as a single optimization problem, which we solve using machine learning methods. This approach is motivated by the observation that the peptide and protein level tasks are cooperative, and the solution to each can be improved by using information about the solution to the other. The resulting algorithm directly controls the relevant error rate, can incorporate a wide variety of evidence and, for complex samples, provides 18-34% more protein identifications than the current state of the art approaches.