Tissue transglutaminase (TGM2; also known as TG2 or tTG) localizes to the syncytial microvillous membrane (MVM) of the human placenta, the primary interface between maternal and fetal tissue. To identify TGM2 substrates in the MVM, membrane vesicles were prepared and labeled with biotinylated acyl donor or acceptor probes. Biotinylated species were selected on an avidin affinity matrix and identified by mass spectrometry of tryptic peptides. The most abundant were cytoskeletal (actin, tubulin, and cytokeratin) and membrane-associated (annexins, integrins, and placental alkaline phosphatase) proteins. During pregnancy, apoptotic particulate material, the end product of the trophoblast life cycle, is shed from the MVM into maternal circulation. Shed material was isolated from primary trophoblast cultures in which syncytial-like masses develop by fusion. A substantial fraction of actin in the particles was in the form of covalent polymeric aggregates, in contrast to cellular actin, which dissociated completely into monomer in SDS-PAGE. When cells were cultured in the presence of transglutaminase inhibitors, actin in the shed particles remained exclusively in monomeric form, and a reduction in trophoblast intercellular fusion and differentiation was observed. These findings suggest that transglutaminase-mediated cross-linking stabilizes the particulate material shed from the placenta.
"For example, shedding of cytoplasmic and nuclear remnants, under the form of cytoplasmic microvesicles or DNA containing microparticles, during cell death of placental multinucleated syncytiotrophoblast is associated with preeclampsia [33–35]. Transglutaminase was shown to normally crosslink F-actin during cell death of multinucleated syncytiotrophoblast creating a large scaffold of polymerized actin that retained cell remnants of dead syncytium masses and prevented shedding of microvesicles . It would be of interest to evaluate if a similar cross-linking system based on transglutaminase is present in LGLs and if it contributes to harden the F-actin scaffold of the clot and retain LGL remnants within the clot structure. "
[Show abstract][Hide abstract] ABSTRACT: Clot formation in the sipunculid Themiste petricola, a coelomate nonsegmented marine worm without a circulatory system, is a cellular response that creates a haemostatic mass upon activation with sea water. The mass with sealing properties is brought about by homotypic aggregation of granular leukocytes present in the coelomic fluid that undergo a rapid process of fusion and cell death forming a homogenous clot or mass. The clot structure appears to be stabilized by abundant F-actin that creates a fibrous scaffold retaining cell-derived components. Since preservation of fluid within the coelom is vital for the worm, clotting contributes to rapidly seal the body wall and entrap pathogens upon injury, creating a matrix where wound healing can take place in a second stage. During formation of the clot, microbes or small particles are entrapped. Phagocytosis of self and non-self particles shed from the clot occurs at the clot neighbourhood, demonstrating that clotting is the initial phase of a well-orchestrated dual haemostatic and immune cellular response.
International Journal of Cell Biology 03/2012; 2012:280675. DOI:10.1155/2012/280675
"The actin and keratin cytoskeleton networks, which are present in all trophoblast cell subtypes of the mouse and human placenta, are required to maintain the structural integrity of these cells. During pregnancy, apoptotic cell material, the end product of the trophoblast life cycle, is permanently shed from the microvilli into the maternal blood . Therefore, STBs require a highly dynamic cytoskeleton rearrangement and permanent repair processes of the plasma membrane. "
[Show abstract][Hide abstract] ABSTRACT: We want to identify proteins that are part of or associated with the plasma membrane of the human feto-maternal barrier, which is crucially important for nutrient, gas, and waste exchange between the mother and the fetus. All transfer processes occur through one specialized endothelial cell layer, the multinuclear syncytiotrophoblast (STB). Specifically, the apical plasma membrane of the STB interacts with the maternal blood and is the site of initial transport processes across the placenta.
We used a proteomic approach that employed the enrichment of apical STB membranes isolated from healthy placentae by ultracentrifugation and saccharose gradient centrifugation steps in combination with 1-D SDS-PAGE and ESI-MS analysis.
We identified 296 different proteins, 175 of which were integral and peripheral membrane proteins, partially containing 1-12 transmembrane domains or lipid anchors. One hundred and sixty-one proteins (54%) were allocated to the plasma membrane.
A high number of transporters, receptors, and proteins involved in signal transduction processes and vesicular trafficking were identified for the first time at the feto-maternal barrier. Our results are valuable sources for further studies of the cell physiology of the healthy placenta at the time of birth or the pathophysiology of several pregnancy disorders.
"Previous results from animal cells suggested the involvement of TGase in the modification of cytoskeleton proteins showing that actin is a substrate of TGase (Lorand and Graham 2003). Recently, Robinson et al. (2007) summarized the cytoskeleton substrates of TGase 2, one of the nine TGases identified in animals. In plants, the crude extract of Malus domestica pollen contained TGase activity and catalyzed the incorporation of PAs into proteins with molecular mass of 43 kDa and 52–58 kDa, identified as actin and a-tubulin. "
[Show abstract][Hide abstract] ABSTRACT: The polyamine (PA) content and the transglutaminase (TGase) activity have been investigated in Pyrus communis pollination with compatible and self-incompatible (SI) pollen in order to deepen their possible involvement in the progamic phase of plant reproduction. The PA distribution as free, perchloric acid (PCA)-soluble and PCA-insoluble fractions in ungerminated (UGP), germinating pollen (GP), styles and pollinated styles with compatible and SI pollens is discussed in the light of a possible role during pollination. Generally, the conjugated PAs both in PCA-soluble and PCA-insoluble fractions were higher than the free form. Within the conjugated PAs, the PCA-insoluble ones were the highest with the exception of the not pollinated styles. As TGase mediates some of the effects of PAs by covalently binding them to proteins, the activity of this enzyme, never checked before in styles and pollinated styles, was examined. In the SI styles, the TGase activity is higher in comparison to style-pollinated with compatible pollen, and high molecular mass cross-linked products were formed, suggesting an involvement of TGase in SI response. This is the first evidence on the presence of this enzyme activity in not pollinated and pollinated styles.
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