Altered placental expression of PAPPA2 does not affect birth weight in mice

Background  

Pregnancy-associated plasma protein A2 (PAPPA2) is an insulin-like growth factor binding protein (IGFBP) protease expressed in the placenta and upregulated in pregnancies complicated by pre-eclampsia. The mechanism linking PAPPA2 expression and pre-eclampsia and the consequences of altered PAPPA2 expression remain unknown. We previously identified PAPPA2 as a candidate gene for a quantitative trait locus (QTL) affecting growth in mice and in the present study examined whether this QTL affects placental PAPPA2 expression and, in turn, placental or embryonic growth.     

Association of serum markers with improvement in clinical response measures after treatment with golimumab in patients with active rheumatoid arthritis despite receiving methotrexate: results from the GO-FORWARD study

Introduction  

The goal of this study was to identify serum markers that are modulated by treatment with golimumab with or without methotrexate (MTX) and are associated with clinical response.     

Connective Tissue Growth Factor in Regulation of RhoA Mediated Cytoskeletal Tension Associated Osteogenesis of Mouse Adipose-Derived Stromal Cells

Background

Cytoskeletal tension is an intracellular mechanism through which cells convert a mechanical signal into a biochemical response, including production of cytokines and activation of various signaling pathways.

Methods/Principal Findings

Adipose-derived stromal cells (ASCs) were allowed to spread into large cells by seeding them at a low-density (1,250 cells/cm²), which was observed to induce osteogenesis. Conversely, ASCs seeded at a high-density (25,000 cells/cm²) featured small cells that promoted adipogenesis. RhoA and actin filaments were altered by changes in cell size. Blocking actin polymerization by Cytochalasin D influenced cytoskeletal tension and differentiation of ASCs. To understand the potential regulatory mechanisms leading to actin cytoskeletal tension, cDNA microarray was performed on large and small ASCs. Connective tissue growth factor (CTGF) was identified as a major regulator of osteogenesis associated with RhoA mediated cytoskeletal tension. Subsequently, knock-down of CTGF by siRNA in ASCs inhibited this osteogenesis.

Conclusions/Significance

We conclude that CTGF is important in the regulation of cytoskeletal tension mediated ASC osteogenic differentiation.     

Specific visualization of the distribution of the calcium dependent regulatory protein of cyclic nucleotide phosphodiesterase (modulator protein) in tissue culture cells by immunofluorescence microscopy: mitosis and intercellular bridge.

Monospecific antibodies against the homogeneous Ca++ dependent regulatory protein of cyclic nucleotide phosphodiesterase (CDR protein) from bovine brain were used in indirect immunofluorescence microscopy to visualize the cytoplasmic organization of this key regulatory protein in growing tissue culture cells. Although cells during interphase reveal only a weak general cytoplasmic fluorescence, a dramatic reorganization of CDR protein occurs with the onset of mitosis. Throughout the different mitotic stages CDR protein is strongly concentrated in the two polar parts of each half spindle. After completion of telophase (CDR protein appears at both cytoplasmic ends of the intercellular bridge which still connects the two daughter cells. Parallel use of monospecific antibodies against CDR protein and tubulin emphasizes the spatial restriction in the localization of CDR protein during mitosis and early G1 phase of the cell cycle.     

Immunological analysis of hemoglobin transition during metamorphosis of normal and isogenicXenopus

Summary Antisera against larval and adultXenopus hemoglobins as well as adult human hemoglobin showed no cross-reaction when tested by immunodiffusion against each heterologous antigen. In this test hemoglobin of a single animal produced two precipitation lines for larvae, but only one for adult stages. Immunoelectrophoresis also revealed more complex precipitation patterns for larval than for adult hemoglobins. Hemoglobin of the isogenic hybrid cloneXenopus laevis/X. gilli also reacted with antisera against normalXenopus hemoglobin.Quantitation of hemoglobins, analyzed by radial immunodiffusion showed fewer than 1% of adult hemoglobin in red cells of larvae, but 30% at completion of metamorphosis. Two weeks later adult hemoglobin attained over 90%, and in red cells of adultXenopus an average of 1% larval hemoglobin were detected.The relatively short transition period suggests that the loss of larval hemoglobin may be due to the elimination of larval red cells, and that the increase in adult hemoglobin may be indicative of a new cell line.     

Characterization of a Ca(2+)-binding site in human annexin II by site-directed mutagenesis.

Annexin II, a major cytoplasmic substrate of the src tyrosine kinase, is a member of the annexin family of Ca2+/phospholipid-binding proteins. It is composed of a short N-terminal tail (30 residues) followed by four so-called annexin repeats (each 70-80 residues in length) which share sequence homologies and are thought to form (a) new type(s) of Ca(2+)-binding site(s). We have produced wild-type and site specifically mutated annexin II molecules to compare their structure and biochemistry. The recombinant wild-type annexin II displays biochemical and spectroscopical properties resembling those of the authentic protein purified from mammalian cells. In particular, it shows the Ca(2+)-induced blue shift in fluorescence emission which is typical for this annexin. Replacement of the single tryptophan in annexin II (Trp-212) by a phenylalanine abolishes the fluorescence signal and allows the unambiguous assignment of the Ca(2+)-sensitive spectroscopic properties to Trp-212. This residue is located in the third annexin repeat in a highly conserved stretch of 17 amino acids which are also found in the other repeats and known as the endonexin fold. To study the precise architecture of the Ca2+ site which must reside in close proximity to Trp-212, we changed several residues of the endonexin fold in repeat 3 by site-directed mutagenesis. An analysis of these mutants by fluorescence spectroscopy and Ca(2+)-dependent phospholipid binding reveals that Gly-206 and Thr-207 seem indispensible for a correct folding of this Ca(2+)-binding site.     

Effect of skeletal muscle myosin light chain 2 on the Ca2+-sensitive interaction of myosin and heavy meromyosin with regulated actin

A low-speed centrifugation assay has been used to examine the binding of myosin filaments to F-action and to regulated actin in the presence of MgATP. While the cross-linking of F-actin by myosin was Ca2+ insensitive, much less regulated actin was cross-linked by myosin in the absence of Ca2+ than in its presence. Removal of the 19000-dalton, phosphorylatable light chain from myosin resulted in the loss of this Ca2+ sensitivity. Readdition of this light chain partially restored the Ca2+-sensitive cross-linking of regulated actin by myosin. Urea gel electrophoresis has been used to distinguish that fraction of heavy meromyosin which contains intact phosphorylatable light chain from that which contains a 17000-dalton fragment of this light chain. In the absence of Ca2+, heavy meromyosin which contained digested light chain bound to regulated actin in MgATP about 10-fold more tightly than did heavy meromyosin which contained intact light chain. The regulated actin-activated ATPases of heavy meromyosin also showed that cleavage of this light chain causes a substantial increase in the affinity of heavy meromyosin for regulated actin in the absence of Ca2+. Thus, the binding of both myosin and heavy meromyosin to regulated actin is Ca2+ sensitive, and this sensitivity is dependent on the phosphorylatable light chain.     

Differential Expression of Two Soybean (Glycine max L.) Proline-Rich Protein Genes after Wounding

We have investigated the wound-induced expression of two members of the soybean (Glycine max L.) proline-rich cell wall protein gene family and show that SbPRP1 and SbPRP2 exhibit unique patterns of expression after physical damage. SbPRP1 mRNA can be detected in the hook of soybean seedlings within 2 h after wounding and is present at high levels in the hook and elongating hypocotyl 20 h after wounding. In contrast, SbPRP2 mRNA increases transiently and rapidly throughout the soybean seedling after wounding. SbPRP2 is also induced by wounding in soybean leaves, but the pattern of mRNA accumulation in leaves is distinct from that seen in seedlings and reaches high levels of expression 20 h after physical damage. SbPRP2 mRNA levels were also found to increase in the mature hypocotyl and roots of seedlings in response to treatment with 10 [mu]M indoleacetic acid and naphthalene-1-acetic acid. These data indicate that the wound-induced expression of PRPs in soybean is tissue specific and that the regulation of these genes after physical damage may operate through different signal transduction pathways.