Serine/arginine-rich splicing factor 3 (SRSF3), a member of the serine/arginine (SR)-rich family of proteins, regulates both alternative splicing of pre-mRNA and export of mature mRNA from the nucleus. Although its role in nuclear mRNA processing is well understood, the mechanism by which it alters the fate of cytoplasmic mRNA molecules remains elusive. Here, we provide evidence that SRSF3 not only regulates the alternative splicing pattern of programmed cell death 4 (PDCD4) mRNA, but also modulates its translational efficiency in the cytoplasm by lowering translation levels. We observed a marked increase in PDCD4 mRNA in translating polysome fractions upon silencing of SRSF3, and, conversely, ectopic overexpression of SRSF3 shifted PDCD4 mRNA into non-translating ribosomal fractions. In live cells, SRSF3 colocalized with PDCD4 mRNA in P-bodies (PBs), where translationally silenced mRNAs are deposited, and this localization was abrogated upon SRSF3 silencing. Furthermore, using two different reporter systems, we showed that SRSF3 interacts directly with PDCD4 mRNA and mediates translational repression by binding to the 5 0 -untranslated region (5 0 -UTR). In summary, our data suggest that the oncogenic potential of SRSF3 might be realized, in part, through the translational repression of PDCD4 mRNA.

To cite this article: Wang J-S, Chow S-E, Chen J-K. Strenuous, acute exercise affects reciprocal modulation of platelet and polymorphonuclear leukocyte activities under shear¯ow in men. J Thromb Haemost 2003; 1: 2031±7.Summary. Vigorous exercise transiently increases the risk of primary cardiac arrest. The reciprocal modulation of platelet and polymorphonuclear leukocyte (PMN) activities is important in the pathogenesis of thrombosis. This study investigates how strenuous, acute exercise affects platelet±PMN reciprocal modulation by closely examining 18 sedentary men who exercised strenuously on a bicycle ergometer. Shear-induced platelet activation, PMN interaction with surface-adherent platelets under shear¯ow, and PMN-dependent inhibition of platelet activation were measured both before and immediately after exercise. Analytical results can be summarized as follows: (i) shear-induced platelet adhesion on ®bronectin-coated surface as well as ADP-induced release of platelet soluble Pselectin release and elevation of [Ca 2 ] i signi®cantly increases after strenuous exercise; (ii) strenuous exercise is associated with higher velocity and percentage of rolling PMNs and lower numbers of PMNs remaining bound to surface-adherent platelets under shear¯ow than at rest; (iii) PMN-dependent inhibition of platelet [Ca 2 ] i elevation and soluble P-selectin release after strenuous exercise is much greater than that at rest; and (iv) strenuous exercise increases PMN-derived nitric oxide metabolite level and reduces oxidized low-density lipoprotein-promoted interaction between platelets and PMNs. Therefore, we conclude that platelet activity may be sensitized by strenuous exercise. However, strenuous exercise can also simultaneously enhance the antiplatelet effect of PMNs. The ®nding provides a new insight into the negative feedback of PMNs against exercise-evoked platelet-related thrombotic risk.

Oxidative stress and inflammation are important processes in the progression of Alzheimer's disease (AD). Recent studies have implicated the role of amyloid β-peptides (Aβ) in mediating these processes. In astrocytes, oligomeric Aβ induces the assembly of NADPH oxidase complexes resulting in its activation to produce anionic superoxide. Aβ also promotes production of proinflammatory factors in astrocytes. Since low energy laser has previously been reported to attenuate oxidative stress and inflammation in biological systems, the objective of this study was to examine whether this type of laser light was able to abrogate the oxidative and inflammatory responses induced by Aβ. Primary rat astrocytes were exposed to Helium-Neon laser (λ=632.8 nm), followed by the treatment with oligomeric Aβ. Primary rat astrocytes were used to measure Aβ-induced production of superoxide anions using fluorescence microscopy of dihydroethidium (DHE), assembly of NADPH oxidase subunits by the colocalization between the cytosolic p47 phox subunit and the membrane gp91 phox subunit using fluorescent confocal microscopy, phosphorylation of cytosolic phospholipase A 2 (cPLA 2 ), and expressions of pro-inflammatory factors including interleukin-1β (IL-1β) and inducible nitric-oxide synthase (iNOS) using Western blot Analysis. Our data showed that laser light at 632.8 nm suppressed Aβ-induced superoxide production, colocalization between NADPH oxidase gp91 phox and p47 phox subunits, phosphorylation of cPLA 2 , and the expressions of IL-1β and iNOS in primary astrocytes. We demonstrated for the first time that 632.8 nm laser was capable of suppressing cellular pathways of oxidative stress and inflammatory responses critical in the pathogenesis in AD. This study should prove to provide the groundwork for further investigations for the potential use of laser therapy as a treatment for AD.

Nuclear processes depend heavily on the organization of chromatin, whose subunits are cylinder-shaped complexes called nucleosomes. A subset of mammalian nucleosomes in situ resemble the canonical structure determined in vitro 24 years ago. The structure of nucleosomes in situ is otherwise poorly understood. Here we use cryo-ET and 3-D classification analysis to study the structure of yeast nucleosomes both in vitro and in situ. We show that the class averages of GFP-tagged yeast nucleosomes in vitro resemble canonical nucleosomes, with additional GFP densities. In contrast, none of the class averages of nucleosome-like particles in situ (inside cells) resemble canonical nucleosomes. The heterogeneous nature of the in situ class averages suggests that the intranuclear environment favors multiple conformations. Using the structural observations here and the results of previous genomics and biochemical studies, we propose a model in which the average yeast nucleosome's DNA is partially detached in situ.

To monitor cellular processes in individual cells, it is important to measure the concentrations of intracellular metabolites and to retrieve them for analysis. The use of functionalized polyelectrolyte microcapsules as intracellular sensors for in vivo reporting is persented. Capsules loaded with streptavidinrhodamine, which was introduced into fibroblasts by electroporation, autonomously escaped from an endocytic compartment and effi ciently recruited biotin-fluorescein from the cytosol. This work demonstrates the utility of polyelectrolyte microcapsules for intracellular capture of metabolites and eventually for drug delivery on an organismic level.