Distribution Of Annexin Research Articles

The NK1 Receptor Localizes to the Plasma Membrane Microdomains, and Its Activation Is Dependent on Lipid Raft Integrity

The spatial targeting of receptors to discrete domains within the plasma membrane allows their preferential coupling to specific effectors, which is essential for rapid and accurate discrimination of signals. Efficiency of signaling is further increased by protein and lipid segregation within the plasma membrane. We have previously demonstrated the importance of raft-mediated signaling in the regulation of smooth and skeletal muscle cell contraction. Since G protein-coupled receptors (GPCRs) are key components in the regulation of smooth muscle contraction-relaxation cycles, it is important to determine whether GPCR signaling is mediated by lipid rafts and raft-associated molecules. Neurokinin 1 receptor (NK1R) is expressed in central and peripheral nervous system as well as in endothelial and smooth muscle cells and involved in mediation of pain, inflammation, exocrine secretion, and smooth muscle contraction. The NK1 receptor was transiently expressed in HEK293 and HepG2 cell lines and its localization in membrane microdomains investigated using biochemical methods and immunofluorescent labeling. We show that the NK1 receptor, similar to the earlier described beta(2)-adrenergic receptor and G proteins, localizes to lipid rafts and caveolae. Protein kinase C (PKC) is one of the downstream effectors of the NK1 activation. Its active form translocates from the cytoplasm to the plasma membrane. Upon stimulation of the NK1 receptor with Substance P, the activated PKC relocated to lipid rafts. Using cholesterol extraction and replenishment assays we show that activation of NK1 receptor is dependent on the microarchitecture of the plasma membrane: NK1R-mediated signaling was abolished after cholesterol depletion of the receptor-expressing cells with methyl-beta-cyclodextrin. Our results demonstrate that reorganization of the plasma membrane has an effect on the activation of the raft-associated NK1R and the down-stream events such as recruitment of protein kinases.

Distribution of annexin 5 in the testis and variation of expression by different physiological changes

Distribution of annexin 5 in the testis and variation of expression by different physiological changes

LDL-cholesterol modulates the membrane binding and intracellular distribution of annexin 6 in CHO cells

LDL-cholesterol modulates the membrane binding and intracellular distribution of annexin 6 in CHO cells

Immunocytochemical localization of annexin 5, a calcium-dependent phospholipid-binding protein, in rat endocrine organs.

Annexin 5, a unique calcium- and phospholipid-binding protein, has been investigated for its specific distribution in rat endocrine organs by immunocytochemistry with a specific antiserum to recombinant rat annexin 5. Follicular epithelial cells and parafollicular cells of the thyroid gland, adrenocortical cells of the zona fasciculata and zona reticularis, luteal cells, testicular interstitial cells, and Sertoli cells were shown to contain annexin 5. To examine whether the synthesis of annexin 5 would be affected by a change in humoral signal, the distribution of annexin 5 in the anterior pituitary was examined three weeks after ovariectomy. The withdrawal of ovarian hormones induced huge castration cells in the anterior pituitary gland, which contained abundant annexin 5. Annexin 5 was not detected in the pineal gland, the parathyroid gland, the islet of Langerhans, the adrenal medulla, zona glomerulosa cells, and granulosa cells. Since annexin 5 was shown to exist in many of the endocrine tissues examined, to be localized in specific cell types, and to be abundant in castration cells, it is suggested that annexin 5 contributes to secretory cell functions, which may be common to endocrine cells secreting chemically different hormones.

Distribution of annexin I and II in term human fetal membranes, decidua and placenta

Annexin I and II are calcium binding proteins implicated in the regulation of a number of cellular functions, including secretory processes, prolactin release and prostaglandin formation. The cellular distribution of these proteins was examined in human term placenta, fetal membranes (amnion and chorion laeve) and decidua using immunohistochemistry. Annexin I was found in amnion epithelial cells and chorion laeve trophoblast but not in decidua, and was located in the syncytiotrophoblast cells of placenta. Annexin II and annexin II light-chain were located in the amnion epithelial cells, the cells of the mesenchymal layer between the amnion and chorion laeve trophoblast and endothelial cells lining the blood vessels in the decidua. In contrast to annexin I, annexin II was located in the villous core and not the syncytiotrophoblast cells in the placenta. There was no apparent change in the distribution of these annexins during labour. These findings indicate that the cellular distribution of these annexins is different, and may be an important consideration when examining their synthesis or action in tissues and in vitro with mixed cell populations and tissue homogenates.

Membrane-bound annexin V isoforms (CaBP33 and CaBP37) and annexin VI in bovine tissues behave like integral membrane proteins

The distribution of annexin V isoforms (CaBP33 and CaBP37) and of annexin VI in bovine lung, heart, and brain subfractions was investigated with special reference to the fractions of these proteins which are membrane-bound. In addition to EGTA-extractable pools of the above proteins, membranes from lung, heart, and brain contain EGTA-resistant annexins V and VI which can be solubilized with detergents (Triton X-100 or Triton X-114). A strong base like Na 2CO 3, which is usually effective in extracting peripheral membrane proteins, only partially solubilizes the membrane-bound, EGTA-resistant annexins analyzed here. Also, only 50–60% of the Triton X-114-soluble annexins partition in the aqueous phase, the remaining fractions being recovered in the detergent-rich phase. Altogether, these findings suggest that, by an as yet unknown mechanism, following Ca 2+-dependent association of annexin V isoforms and annexin VI with membranes, substantial fractions of these proteins remain bound to membranes in a Ca 2+-independent way and behave like integral membrane proteins. These results further support the possibility that the above annexins might play a role in membrane trafficking and/or in the regulation of the structural organization of membranes.

Immunoelectron microscopic analysis of the intracellular distribution of primer recognition proteins, annexin 2 and phosphoglycerate kinase, in normal and transformed cells

Primer recognition proteins (PRP) are accessory proteins for DNA polymerase alpha in lagging strand DNA replication. We have previously reported that the PRP consist of a complex of two proteins identified as 3-phosphoglycerate kinase (PGK) and the protein-tyrosine kinase substrate, annexin 2 monomer. The physiological role of annexin 2 is not known. Two pools of annexin 2 exist in cells. A majority of annexin 2 is localized with the plasma membrane as a heterotetramer in association with a light chain. Monomer annexin 2 is cytosolic. The identification of annexin 2 monomer as a part of the PRP complex represents one of the physiological roles of this protein in cells. To function as PRP, annexin 2 and PGK would have to be present in the cell nucleus. To investigate whether monomer annexin 2 is indeed associated with nuclear DNA synthesis, we investigated the presence of annexin 2 and PGK in the cell nucleus. In this paper, we demonstrate the presence of annexin 2 and PGK in nuclear extracts. The nuclear fraction of these proteins represents a small subset of the total cellular pools. Immunoelectron-microscopic analyses using anti-PRP antisera demonstrate the distribution of these proteins in HeLa cell nuclei and cytoplasm. Under identical conditions, an anti-cytokeratin monoclonal antibody preferentially labels the plasma membrane without detectable intracellular staining. The distribution of annexin 2 and PGK in both nuclei and cytoplasm is similarly observed in cells from normal tissues such as freshly isolated rat hepatocytes and hamster pancreatic tissue.(ABSTRACT TRUNCATED AT 250 WORDS)