Consensus peptide antibodies reveal a widespread occurrence of Ca2+/lipid-binding proteins of the annexin family

AbstractAntibodies generated against synthetic peptides that correspond to highly conserved sequence motifs in the annexins reacted with a variety of annexins from different species. These include Xenopus laevis and Drosophila melanogaster, which contain cross-reacting polypeptides of apparent Mr, 34000 and 30000. As expected for typical annexins, the two Drosophila proteins interact in a Ca2+-dependent manner with phosphatidylserine liposomes

Annexin A8 displays unique phospholipid and F-actin binding properties

AbstractAnnexin A8 is a poorly characterized member of the annexin family of Ca2+-regulated membrane binding proteins. Initially only identified at the cDNA level it had been tentatively linked to acute promyelocytic leukaemia (APL) due to its high and regulated expression in APL-derived cells. Here we identify unique properties of the annexin A8 protein. We show that it binds Ca2+-dependently and with high specificity to phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P2) and is also capable of interacting with F-actin. In line with these characteristics annexin A8 is recruited to F-actin-associated PtdIns(4,5)P2-rich membrane domains formed in HeLa cells upon infection with non-invading enteropathogenic Escherichia coli. These properties suggest a role of annexin A8 in the organization of certain actin-associated membrane domains

Analysis of Cd44-Containing Lipid Rafts: Recruitment of Annexin II and Stabilization by the Actin Cytoskeleton

CD44, the major cell surface receptor for hyaluronic acid (HA), was shown to localize to detergent-resistant cholesterol-rich microdomains, called lipid rafts, in fibroblasts and blood cells. Here, we have investigated the molecular environment of CD44 within the plane of the basolateral membrane of polarized mammary epithelial cells. We show that CD44 partitions into lipid rafts that contain annexin II at their cytoplasmic face. Both CD44 and annexin II were released from these lipid rafts by sequestration of plasma membrane cholesterol. Partition of annexin II and CD44 to the same type of lipid rafts was demonstrated by cross-linking experiments in living cells. First, when CD44 was clustered at the cell surface by anti-CD44 antibodies, annexin II was recruited into the cytoplasmic leaflet of CD44 clusters. Second, the formation of intracellular, submembranous annexin II–p11 aggregates caused by expression of a trans-dominant mutant of annexin II resulted in coclustering of CD44. Moreover, a frequent redirection of actin bundles to these clusters was observed. These basolateral CD44/annexin II–lipid raft complexes were stabilized by addition of GTPγS or phalloidin in a semipermeabilized and cholesterol-depleted cell system. The low lateral mobility of CD44 in the plasma membrane, as assessed with fluorescent recovery after photobleaching (FRAP), was dependent on the presence of plasma membrane cholesterol and an intact actin cytoskeleton. Disruption of the actin cytoskeleton dramatically increased the fraction of CD44 which could be recovered from the light detergent-insoluble membrane fraction. Taken together, our data indicate that in mammary epithelial cells the vast majority of CD44 interacts with annexin II in lipid rafts in a cholesterol-dependent manner. These CD44-containing lipid microdomains interact with the underlying actin cytoskeleton

Annexin 2 has an essential role in actin-based macropinocytic rocketing

AbstractAnnexin 2 is a Ca2+ binding protein that binds to and aggregates secretory vesicles at physiological Ca2+ levels [1] and that also associates Ca2+ independently with early endosomes [2, 3]. These properties suggest roles in both exocytosis and endocytosis, but little is known of the dynamics of Annexin 2 distribution in live cells during these processes. We have used evanescent field microscopy to image Annexin 2-GFP in live, secreting rat basophilic leukemia cells and in cells performing pinocytosis. Although we found no evidence of Annexin 2 involvement in exocytosis, we observed an enrichment of Annexin 2-GFP in actin tails propeling macropinosomes. The association of Annexin 2-GFP with rocketing macropinosomes was specific because Annexin 2-GFP was absent from the actin tails of rocketing Listeria. This finding suggests that the association of Annexin 2 with macropinocytic rockets requires native pinosomal membrane. Annexin 2 is necessary for the formation of macropinocytic rockets since overexpression of a dominant-negative Annexin 2 construct abolished the formation of these structures. The same construct did not prevent the movement of Listeria in infected cells. These results show that recruitment of Annexin 2 to nascent macropinosome membranes 16656is an essential prerequisite for actin polymerization-dependent vesicle locomotion

Suppression of electrical breakdown phenomena in liquid TriMethyl Bismuth based ionization detectors

Organometallic liquids provide good properties for ionization detectors. TriMethyl Bismuth (TMBi) has been proposed as a detector medium with charge and Cherenkov photon readout for Positron Emission Tomography. In this work, we present studies for the handling of TMBi at different electric fields and under different environmental conditions to find applicable configurations for the suppression of electrical breakdowns in TMBi at room temperature. A simple glass cell with two electrodes filled with TMBi was constructed and tested under different operation conditions. Working at the vapour pressure of TMBi at room temperature of about 40 mbar and electric fields of up to 20 kV/cm in presence of a small oxygen contamination we found the formation of a discharge channel in the liquid and a steady increase in the current. Further reduction of pressure by pumping caused the TMBi to boil and a spontaneous combustion. Eliminating the oxygen contamination led the TMBi under the same condition to only decompose. When operating the setup under an argon atmosphere of 1 bar we did not observe breakdowns of the electrical potential up to field strengths of 20 kV/cm. Still, in presence of a small oxygen contamination fluctuating currents in the nA range were observed, but no decomposition or combustion. We conclude from our experiments that TMBi at room temperature in a pure argon atmosphere of 1 bar remains stable against electrical breakdown at least up to electric field strengths of 20 kV/cm, presumably because the formation of gaseous TMBi was prevented.Comment: 14 page, 8 figure

Detecting neutral hydrogen in emission at redshift z ~ 1

We use a large N-body simulation to examine the detectability of HI in emission at redshift z ~ 1, and the constraints imposed by current observations on the neutral hydrogen mass function of galaxies at this epoch. We consider three different models for populating dark matter halos with HI, designed to encompass uncertainties at this redshift. These models are consistent with recent observations of the detection of HI in emission at z ~ 0.8. Whilst detection of 21 cm emission from individual halos requires extremely long integrations with existing radio interferometers, such as the Giant Meter Radio Telescope (GMRT), we show that the stacked 21 cm signal from a large number of halos can be easily detected. However, the stacking procedure requires accurate redshifts of galaxies. We show that radio observations of the field of the DEEP2 spectroscopic galaxy redshift survey should allow detection of the HI mass function at the 5-12 sigma level in the mass range 10^(11.4) M_sun/h < M_halo < 10^(12.5)M_sun/h, with a moderate amount of observation time. Assuming a larger noise level that corresponds to an upper bound for the expected noise for the GMRT, the detection significance for the HI mass function is still at the 1.7-3 sigma level. We find that optically undetected satellite galaxies enhance the HI emission profile of the parent halo, leading to broader wings as well as a higher peak signal in the stacked profile of a large number of halos. We show that it is in principle possible to discern the contribution of undetected satellites to the total HI signal, even though cosmic variance limitation make this challenging for some of our models.Comment: 14 pages, 9 figures, Submitted To MNRA

The frontotemporal dementia mutation R406W blocks tau’s interaction with the membrane in an annexin A2–dependent manner

The R406W mutation prevents tau from functioning as a linker between the membrane and the microtubule cytoskeleton

Agonists of proteinase-activated receptor-2 affect transendothelial migration and apoptosis of human neutrophils

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/73837/1/j.1600-0625.2007.00605.x.pd

AJAM-A–tetraspanin–αvβ5 integrin complex regulates contact inhibition of locomotion

Contact inhibition of locomotion (CIL) is a process that regulates cell motility upon collision with other cells. Improper regulation of CIL has been implicated in cancer cell dissemination. Here, we identify the cell adhesion molecule JAM-A as a central regulator of CIL in tumor cells. JAM-A is part of a multimolecular signaling complex in which tetraspanins CD9 and CD81 link JAM-A to αvβ5 integrin. JAM-A binds Csk and inhibits the activity of αvβ5 integrin-associated Src. Loss of JAM-A results in increased activities of downstream effectors of Src, including Erk1/2, Abi1, and paxillin, as well as increased activity of Rac1 at cell–cell contact sites. As a consequence, JAM-A-depleted cells show increased motility, have a higher cell–matrix turnover, and fail to halt migration when colliding with other cells. We also find that proper regulation of CIL depends on αvβ5 integrin engagement. Our findings identify a molecular mechanism that regulates CIL in tumor cells and have implications on tumor cell dissemination.publishedVersio