Abi1 mediates airway smooth muscle cell proliferation and airway remodeling via Jak2/STAT3 signaling

Asthma is a complex pulmonary disorder with multiple pathological mechanisms. A key pathological feature of chronic asthma is airway remodeling, which is largely attributed to airway smooth muscle (ASM) hyperplasia that contributes to thickening of the airway wall and further drives asthma pathology. The cellular processes that mediate ASM cell proliferation are not completely elucidated. Using multiple approaches, we demonstrate that the adapter protein Abi1 (Abelson interactor 1) is upregulated in ∼50% of ASM cell cultures derived from patients with asthma. Loss-of-function studies demonstrate that Abi1 regulates the activation of Jak2 (Janus kinase 2) and STAT3 (signal transducers and activators of transcription 3) as well as the proliferation of both nonasthmatic and asthmatic human ASM cell cultures. These findings identify Abi1 as a molecular switch that activates Jak2 kinase and STAT3 in ASM cells and demonstrate that a dysfunctional Abi1-associated pathway contributes to the progression of asthma

Inhibition of PC cell-derived growth factor (PCDGF)/granulin-epithelin precursor (GEP) decreased cell proliferation and invasion through downregulation of cyclin D and CDK 4 and inactivation of MMP-2

BACKGROUND: PC cell-derived growth factor (PCDGF), also called epithelin/granulin precursor (GEP), is an 88-kDa secreted glycoprotein with the ability to stimulate cell proliferation in an autocrine fashion. In addition, some studies indicated that PCDGF participated in invasion, metastasis and survival of cancer cells by regulating cell migration, adhesion and proliferation. Yet the effects of PCDGF on proliferation and invasion of ovarian cancer cells in vitro and the mechanisms by which PCDGF mediates biological behaviors of ovarian cancer have rarely been reported. In the present study we investigated whether and how PCDGF/GEP mediated cell proliferation and invasion in ovarian cancer. METHODS: PCDGF/GEP expression level in three human ovarian cancer cell lines of different invasion potential were detected by RT-PCR and western blot. Effects of inhibition of PCDGF expression on cell proliferation and invasion capability were determined by MTT assay and Boyden chamber assay. Expression levels of cyclin D1 and CDK4 and MMP-2 activity were evaluated in a pilot study. RESULTS: PCDGF mRNA and protein were expressed at a high level in SW626 and A2780 and at a low level in SKOV3. PCDGF expression level correlated well with malignant phenotype including proliferation and invasion in ovarian cancer cell lines. In addition, the proliferation rate and invasion index decreased after inhibition of PCDGF expression by antisense PCDGF cDNA transfection in SW626 and A2780. Furthermore expression of CyclinD1 and CDK4 were downregulated and MMP-2 was inactivated after PCDGF inhibition in the pilot study. CONCLUSION: PCDGF played an important role in stimulating proliferation and promoting invasion in ovarian cancer. Inhibition of PCDGF decreased proliferation and invasion capability through downregulation of cyclin D1 and CDK4 and inactivation of MMP-2. PCDGF could serve as a potential therapeutic target in ovarian cancer

A Comparative Study of DSP Multiprocessor List Scheduling Heuristics

This paper presents a quantitative comparison of a collection of DSP multiprocessor list scheduling heuristics which consider inter-processor communication delays. The following aspects have been addressed: (1) performance in terms of the total execution time (makespan), (2) sensitivity of heuristics in terms of the characteristics of acyclic precedence graphs, including graph size and graph parallelism, (3) sensitivity of heuristics to the number of processors, and (4) compile time efficiency. In addition, the effectiveness of list scheduling performance enhancement techniques is examined. The main contributions of this paper are: ffl Contrary to the belief of some previous authors, our study indicates that no single published list scheduling heuristic consistently produces the best schedules under all possible program structures and DSP multiprocessor configurations. We believe this fact is very important to designers of DSP multiprocessor scheduling heuristics. ffl Based on such o..

Internal Ribosome entry site mediated translation leads to delocalization of <i>DIAPH1</i> mRNA.

<p><b>A.</b><b> </b> Illustration of bicistronic DIAPH1 expression constructs. D-I-M for DIAPH1-IRES-mCherry and M-I-D for mCherry-IRES-DIAPH1. CEF were transfected for 24 hr and processed for <i>DIAPH1</i> mRNA and HA-tag detection. <b>B–G</b>. Representative transfected cells show localizing <i>DIAPH1</i> mRNA (green in <b>D</b>, indicated by arrows) and delocalizing <i>DIAPH1</i> mRNA (green in <b>G</b>, indicated by arrowheads), respectively. Red: mCherry. <b>B–C</b> and <b>E–F</b> are gray scale images for better presentation of the distribution of mCherry protein and <i>DIAPH1</i> mRNA in cells transfected with the localizing and delocalizing constructs, respectively. <b>H.</b> Quantitative results of <i>DIAPH1</i> mRNA localization from analysis of 300–500 cells from three independent experiments for each expression construct. Error bars: sem. ** p<0.01.</p

Delocalized <i>DIAPH1</i> mRNA cannot be re-localized.

<p><b>A–I</b>. Resumption of translation after puromycin wash-off. CEF grown on cover slips were treated with DMSO or 10 µg/ml of puromycin in methionine-free DMEM for 90 min and then followed by 2×10 min washes with Hank’s balanced saline. Newly synthesized proteins were detected using the Click-iT kit (Invitrogen) as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068190#s4" target="_blank">Materials and Methods</a>. <b>A-H</b>. Representative cells showing the fluorescence signal (red) of the newly synthesized proteins. <b>I</b>. Quantitative results of newly synthesized proteins indicate resumption of protein translation after puromycin wash-off (fluorescence per cell, normalized to that of time zero, representing ∼80 cells at each time point per condition from two independent experiments). <b>J–Q</b>. Representative cells for <i>DIAPH1</i> mRNA distribution after the indicated treatments. Images in the left column are gray scale for better display the <i>DIAPH1</i> mRNA signal. CEF were transfected with HA-tagged DIAPH1 expression plasmid for 24 hr and then treated with DMSO (control, <b>J & K</b>), or 5 µg/ml of transcription inhibitor actinomycin D (Act-D) (<b>L & M</b>), or 10 µg/ml of puromycin (N & O) for 90 min before fixed for FISH detection of <i>DIAPH1</i> mRNA localization. In <b>P & Q</b>, the cells were first treated with 10 µg/ml of puromycin for 90 min then followed by 2×10 min washes with growth medium plus 5 µg/ml of Act-D then incubated in normal growth medium for 90 min before fixed for FISH and <i>DIAPH1</i> mRNA localization score. Note that Act-D at this concentration did not affect the normal localization of already transcribed <i>DIAPH1</i> mRNA. In right column, Red: <i>DIAPH1</i> mRNA; green: HA-tagged Dia1 protein; Blue: nucleus. Dotted lines show cell border. Arrows indicate localizing <i>DIAPH1</i> mRNA molecules. Scale bar: 10 µm. <b>R</b>. Quantitative results of <i>DIAPH1</i> mRNA localization. 300–500 cells were scored for each condition. Error bars: sem. n = 3. **. P<0.01.</p

5′-cap-mediated translation is required for perinuclear <i>DIAPH1</i> mRNA localization.

<p><b>A–H,</b> 4E1RCat inhibits the majority of new protein synthesis (assayed with Click-iT kit, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068190#s4" target="_blank">Materials and Methods</a> for details). <b>I</b>. Quantitative results of 4E1RCat inhibition of new protein synthesis. ∼120 cells were analyzed for each time point per condition from three independent experiments. <b>J.</b> Illustration of bicistronic expression plasmid M-I-D (for mCherry-IRES-DIAPH1). <b>K–N</b>. 4E1RCat inhibits cap-mediated but not IRES-mediated translation. Representative images show transfected cells treated with DMSO (<b>K,L</b>) or 10 µM of 4E1RCat (<b>M,N</b>). CEF were first transfected with the bicistronic plasmid for 2 hr and then incubated with DMSO or 10 µM 4E1RCat for 11 hr. The cells were fixed and processed for immunofluorescence staining for the HA tag. Fluorescence images were acquired and quantified. <b>O</b>. Quantitative result of mCherry/HA ratio in single cells. (n = 12–24). ** p<0.01. <b>P–S</b>. Inhibition of cap-mediated translation delocalizes <i>DIAPH1</i> mRNA. CEF were incubated with DMSO or 10 µM of 4E1RCat in growth medium for 3 hr and then fixed for mRNA detection. <b>P–S</b>, representative cells treated with DMSO or 4E1RCat. Images in left column are gray scale for better display of <i>DIAPH1</i> mRNA signal. Dotted lines indicate cell border. Arrows indicate localizing <i>DIAPH1</i> mRNA. In right column, green: <i>DIAPH1</i> mRNA, blue: nucleus. Note that cells treated with 4E1RCat show diffused <i>DIAPH1</i> mRNA. Scale bar: 10 µm. <b>T</b>. Quantitative result of endogenous <i>DIAPH1</i> mRNA localization in treated CEF. 300–500 cells were scored from three independent experiments for each condition. * p<0.05.</p

Localization of <i>DIAPH1</i> mRNA correlates with DIAPH1 protein distribution.

<p><b>A–F</b>. Representative transfected cells show localization of <i>DIAPH1</i> mRNA and protein. <b>A–B</b> and <b>D–E</b> are gray scale images for the distribution of <i>DIAPH1</i> mRNA and protein in NIH3T3 cells transfected with the construct of D-I-M (<b>A–C</b>) or M-I-D (<b>D–F</b>), respectively. Their merged images are shown in <b>C</b> or <b>F</b>. <b>G</b>. Quantitative results of <i>DIAPH1</i> mRNA localization from analysis of 300 cells from three independent experiments for each expression construct. Error bars: sem. ** p<0.01. <b>H–P.</b> Analysis of the relationship of localization of <i>DIAPH1</i> mRNA and its protein distribution in NIH3T3 cells. <b>H–I</b> and <b>K–L</b> are gray scale images for distribution of DIAPH1-HA fusion protein and mCherry in NIH3T3 cells transfected with the construct of D-I-M (<b>H–I</b>) or M-I-D (<b>K–L</b>), respectively. Their merged images are shown in <b>J</b> or <b>M</b>. <b>N</b>. Illustration of a cell with 5 equal-area zones according to their relative distance to the nucleus border. Note that the shape of the zones are listed in the carton is simplified one and is likely vary within a cell (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068190#s4" target="_blank">Methods and Materials</a> for details). <b>O</b>. Scatter plot graphs show two individual quantitative results of intracellular DIAPH1-HA fusion protein distribution in single NIH3T3 cells transfected with the construct of D-I-M and M-I-D, respectively. The red and blue color lines are linear regression for the ratio points of D-I-M or M-I-D transfected cells. <b>P.</b> A bar graph shows average IDI value for D-I-M or M-I-D transfected cells from analysis of 30 cells for each expression construct. Error bars: sem. ** p<0.01.</p

Manipulation of <i>DIAPH1</i> mRNA localization using an Iron ribo-switch.

<p><b>A</b>. Schematic diagram of the IRE riboswitch (See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068190#s4" target="_blank">Materials and Methods</a> for details). Red balls represent 5′-cap. FB: iron biding protein which also binds to the <i>IRE</i> stem-loop. Green arrow indicates translation permission. <b>B</b>. Western blotting result of mCherry reporter for the effect of IRE in fibroblasts. A construct consisting of <i>IRE-mCherry</i> was transfected into CEF. 3 hr post transfection, ferric ammonium citrate (final 100 µM) or iron chelator desferrioxamine mesylate (final 100 µM) was added into the growth medium. 16 hr after transfection, the cells were collected for Western blotting. Quantitative results of Western blotting (n = 4), * p<0.05. <b>C–H</b>. IRE-mediated control of <i>DIAPH1</i> mRNA localization. CEF were transfected with a construct consisting of <i>IRE</i>-<i>DIAPH1</i> and then treated similarly as in <b>B</b>. 16 hr after transfection, the cells were fixed and processed for FISH detection of mRNA localization. <b>C–H</b>. Representative cells. Red: <i>DIAPH1</i> mRNA signal; Green: HA-tagged DIAPH1 protein signal; Blue: nucleus. <b>C</b>, <b>E</b> & <b>G</b> are gray scale images for better presentation of <i>DIAPH1</i> mRNA in the cells. Dotted lines show cell border. Arrows indicate localizing <i>DIAPH1</i> mRNA. <b>I</b>. Quantitative results of <i>DIAPH1</i> mRNA localization from analysis of 300–500 cells from three independent experiments for each condition. Error bars: sem. ** p<0.01.</p

A Dynamically Scheduled Parallel DSP Architecture for Stream Flow Programming

This paper presents a dynamically scheduled parallel DSP architecture for general purpose DSP computations. The architecture consists of multiple DSP processors and of one or more scheduling units. DSP applications are first captured by stream flow graphs, and then stream flow graphs are statically mapped onto a parallel architecture. The ordering and starting time of DSP tasks are determined by the scheduling unit(s) using a dynamic scheduling algorithm. The main contributions of this paper are summarized as follows: ffl A scalable parallel DSP architecture: The parallel DSP architecture proposed in this paper is scalable to meet signal processing requirements. For parallel DSP architectures with large configurations, the scheduling unit may become a performance bottlenecks. A distributed scheduling mechanism is proposed to address this problem. ffl A mapping algorithm: A algorithm is proposed to systematically map a stream flow graph onto a parallel DSP architecture. ffl A dynamic..