BART Inhibits Pancreatic Cancer Cell Invasion by PKCα Inactivation through Binding to ANX7

A novel function for the binder of Arl two (BART) molecule in pancreatic cancer cells is reported. BART inhibits invasiveness of pancreatic cancer cells through binding to a Ca2+-dependent, phosphorylated, guanosine triphosphatase (GTPase) membrane fusion protein, annexin7 (ANX7). A tumor suppressor function for ANX7 was previously reported based on its prognostic role in human cancers and the cancer-prone mouse phenotype ANX7(+/−). Further investigation demonstrated that the BART–ANX7 complex is transported toward cell protrusions in migrating cells when BART supports the binding of ANX7 to the protein kinase C (PKC) isoform PKCα. Recent evidence has suggested that phosphorylation of ANX7 by PKC significantly potentiates ANX7-induced fusion of phospholipid vesicles; however, the current data suggest that the BART–ANX7 complex reduces PKCα activity. Knocking down endogenous BART and ANX7 increases activity of PKCα, and specific inhibitors of PKCα significantly abrogate invasiveness induced by BART and ANX7 knockdown. These results imply that BART contributes to regulating PKCα activity through binding to ANX7, thereby affecting the invasiveness of pancreatic cancer cells. Thus, it is possible that BART and ANX7 can distinctly regulate the downstream signaling of PKCα that is potentially relevant to cell invasion by acting as anti-invasive molecules

Import of cytochrome c into mitochondria

The import of cytochrome c into mitochondria can be resolved into a number of discrete steps. Here we report on the covalent attachment of heme to apocytochrome c by the enzyme cytochrome c heme lyase in mitochondria from Neurospora crassa. A new method was developed to measure directly the linkage of heme to apocytochrome c. This method is independent of conformational changes in the protein accompanying heme attachment. Tryptic peptides of [35S]cysteine-labelled apocytochrome c, and of enzymatically formed holocytochrome c, were resolved by reverse-phase HPLC. The cysteine-containing peptide to which heme was attached eluted later than the corresponding peptide from apocytochrome c and could be quantified by counting 35S radioactivity as a measure of holocytochrome c formation. Using this procedure, the covalent attachment of heme to apocytochrome c, which is dependent on the enzyme cytochrome c heme lyase, could be measured. Activity required heme (as hemin) and could be reversibly inhibited by the analogue deuterohemin. Holocytochrome c formation was stimulated 5–10-fold by NADH > NADPH > glutathione and was independent of a potential across the inner mitochondrial membrane. NADH was not required for the binding of apocytochrome c to mitochondria and was not involved in the reduction of the cysteine thiols prior to heme attachment. Holocytochrome c formation was also dependent on a cytosolic factor that was necessary for the heme attaching step of cytochrome c import. The factor was a heat-stable, protease-insensitive, low-molecular-mass component of unknown function. Cytochrome c heme lyase appeared to be a soluble protein located in the mitochondrial intermembrane space and was distinct from the previously identified apocytochrome c binding protein having a similar location. A model is presented in which the covalent attachment of heme by cytochrome c heme lyase also plays an essential role in the import pathway of cytochrome c

Structure of 55Sc and development of the N=34 subshell closure

The low-lying structure of $^{55}$Sc has been investigated using in-beam $\gamma$-ray spectroscopy with the $^{9}$Be($^{56}$Ti,$^{55}$Sc+$\gamma$)$X$ one-proton removal and $^{9}$Be($^{55}$Sc,$^{55}$Sc+$\gamma$)$X$ inelastic-scattering reactions at the RIKEN Radioactive Isotope Beam Factory. Transitions with energies of 572(4), 695(5), 1539(10), 1730(20), 1854(27), 2091(19), 2452(26), and 3241(39) keV are reported, and a level scheme has been constructed using $\gamma\gamma$ coincidence relationships and $\gamma$-ray relative intensities. The results are compared to large-scale shell-model calculations in the $sd$-$pf$ model space, which account for positive-parity states from proton-hole cross-shell excitations, and to it ab initio shell-model calculations from the in-medium similarity renormalization group that includes three-nucleon forces explicitly. The results of proton-removal reaction theory with the eikonal model approach were adopted to aid identification of positive-parity states in the level scheme; experimental counterparts of theoretical $1/2^{+}_{1}$ and $3/2^{+}_{1}$ states are suggested from measured decay patterns. The energy of the first $3/2^{-}$ state, which is sensitive to the neutron shell gap at the Fermi surface, was determined. The result indicates a rapid weakening of the $N=34$ subshell closure in $pf$-shell nuclei at $Z>20$, even when only a single proton occupies the $\pi f_{7/2}$ orbital

New data on the systematics and interrelationships of sawfishes (Elasmobranchii, Batoidea, Pristiformes)

New characters based on the arrangement and morphology of dermal denticles show that sawfishes can be divided into two distinctive groups. The first group, comprising the knifetooth sawfish Anoxypristis cuspidata, is characterized by tricuspid denticles variably located on both dorsal and ventral parts of the body. The second group is represented by species of the genus Pristis, showing an uniform and homogenous dermal covering of monocuspidate denticles on both dorsal and ventral sides of the body and within the buccopharyngeal cavity. Pristis is further divided into two subgroups: the first comprises species with denticles lacking any keels and furrows (the smalltooth sawfish Pristis pectinata, the green sawfish Pristis zijsron and the dwarf sawfish Pristis clavata); the second comprises species with denticles presenting keels and furrows well differentiated on their anterior part (the common sawfish Pristis pristis, the largetooth sawfish Pristis perotteti and the greattooth sawfish Pristis microdon). This investigation of the dermal covering provides results which agree with studies that separate the same two species groups of Pristis on the basis of other morphological data

Multi-parallel qPCR provides increased sensitivity and diagnostic breadth for gastrointestinal parasites of humans: field-based inferences on the impact of mass deworming

BACKGROUND: Although chronic morbidity in humans from soil transmitted helminth (STH) infections can be reduced by anthelmintic treatment, inconsistent diagnostic tools make it difficult to reliably measure the impact of deworming programs and often miss light helminth infections. METHODS: Cryopreserved stool samples from 796 people (aged 2-81 years) in four villages in Bungoma County, western Kenya, were assessed using multi-parallel qPCR for 8 parasites and compared to point-of-contact assessments of the same stools by the 2-stool 2-slide Kato-Katz (KK) method. All subjects were treated with albendazole and all Ascaris lumbricoides expelled post-treatment were collected. Three months later, samples from 633 of these people were re-assessed by both qPCR and KK, re-treated with albendazole and the expelled worms collected. RESULTS: Baseline prevalence by qPCR (n = 796) was 17 % for A. lumbricoides, 18 % for Necator americanus, 41 % for Giardia lamblia and 15% for Entamoeba histolytica. The prevalence was <1% for Trichuris trichiura, Ancylostoma duodenale, Strongyloides stercoralis and Cryptosporidium parvum. The sensitivity of qPCR was 98% for A. lumbricoides and N. americanus, whereas KK sensitivity was 70% and 32%, respectively. Furthermore, qPCR detected infections with T. trichiura and S. stercoralis that were missed by KK, and infections with G. lamblia and E. histolytica that cannot be detected by KK. Infection intensities measured by qPCR and by KK were correlated for A. lumbricoides (r = 0.83, p < 0.0001) and N. americanus (r = 0.55, p < 0.0001). The number of A. lumbricoides worms expelled was correlated (p < 0.0001) with both the KK (r = 0.63) and qPCR intensity measurements (r = 0.60). CONCLUSIONS: KK may be an inadequate tool for stool-based surveillance in areas where hookworm or Strongyloides are common or where intensity of helminth infection is low after repeated rounds of chemotherapy. Because deworming programs need to distinguish between populations where parasitic infection is controlled and those where further treatment is required, multi-parallel qPCR (or similar high throughput molecular diagnostics) may provide new and important diagnostic information

Nanolitre real-time PCR detection of bacterial, parasitic, and viral agents from patients with diarrhoea in Nunavut, Canada

Background. Little is known about the microbiology of diarrhoeal disease in Canada&#x0027;s Arctic regions. There are a number of limitations of conventional microbiology testing techniques for diarrhoeal pathogens, and these may be further compromised in the Arctic, given the often long distances for specimen transport. Objective. To develop a novel multiple-target nanolitre real-time reverse transcriptase (RT)-PCR platform to simultaneously test diarrhoeal specimens collected from residents of the Qikiqtani (Baffin Island) Region of Nunavut, Canada, for a wide range of bacterial, parasitic and viral agents. Study design/methods. Diarrhoeal stool samples submitted for bacterial culture to Qikiqtani General Hospital in Nunavut over an 18-month period were tested with a multiple-target nanolitre real-time PCR panel for major diarrhoeal pathogens including 8 bacterial, 6 viral and 2 parasitic targets. Results. Among 86 stool specimens tested by PCR, a total of 50 pathogens were detected with 1 or more pathogens found in 40 (46.5%) stool specimens. The organisms detected comprised 17 Cryptosporidium spp., 5 Clostridium difficile with toxin B, 6 Campylobacter spp., 6 Salmonella spp., 4 astroviruses, 3 noroviruses, 1 rotavirus, 1 Shigella spp. and 1 Giardia spp. The frequency of detection by PCR and bacterial culture was similar for Salmonella spp., but discrepant for Campylobacter spp., as Campylobacter was detected by culture from only 1/86 specimens. Similarly, Cryptosporidium spp. was detected in multiple samples by PCR but was not detected by microscopy or enzyme immunoassay. Conclusions. Cryptosporidium spp., Campylobacter spp. and Clostridium difficile may be relatively common but possibly under-recognised pathogens in this region. Further study is needed to determine the regional epidemiology and clinical significance of these organisms. This method appears to be a useful tool for gastrointestinal pathogen research and may also be helpful for clinical diagnostics and outbreak investigation in remote regions where the yield of routine testing may be compromised