Brought to you by Portland Press Ltd.
Published on behalf of the International Federation for Cell Biology
Cancer Cell death Cell cycle Cytoskeleton Exo/endocytosis Differentiation Division Organelles Signalling Stem cells Trafficking
Cell Biology International (2001) 25, 809–813 (Printed in Great Britain)
Asmarani Kusumawatif2, Jean‑Pierre Liautard and Joannes Sri Widadaf1
INSERM U-431, Université Montpellier II, Case no. 100, Place E. Bataillon, 34095, Montpellier, Cedex 5, France


Directed mutagenesis, in the form of deletions and point mutations, was used to investigate the regulatory importance of the N-terminal domain of annexin 1. Wild-type and mutant forms were fused to green fluorescent protein (GFP) to track their localization and introduced in to J-774A.1 cells by transfection. The fusion of annexin 1 to GFP at the N- or C-terminal end did not alter the cellular distribution or co-localization with phagosomes. The effects of mutations were determined according to these characteristics. The prominent effect resulted from S27E mutation which mimics the phosphorylated state of Ser-27. Although still retaining the granular structures in the cytoplasm, S27E annexin 1 failed to associate with the phagosomal protein complex. This suggests an essential regulatory role of the phosphorylation of residue 27 in annexin 1 function.

Keywords: annexin 1, J-774A.1, phagocytosis, phosphorylation.

f2Permanent address: Faculty of Veterinary Medicine, University Gadjah Mada, Yogyakarta, Indonesia.

f1To whom correspondence should be addressed.


1 Diakonova, M, Gerke, V, Ernst, J, Liautard, JP, van der Vusse, G and Griffith, G (1997) Localization of five annexins in J774 macrophages and on isolated phagosomes. J Cell Sci 110, 1199-213

2 Harricane, MC, Caron, E, Porte, F and Liautard, JP (1996) Distribution of annexin I during non-pathogen or pathogen phagocytosis by confocal imaging and immunogold electron microscopy. Cell Biol Int 20, 193-203

3 Hoekstra, D, Buist-Arkema, R, Klappe, K and Reitelingsperger, CPM (1993) Interaction of annexins with membranes: the N-terminus as a governing parameter as revealed with a chimeric annexin. Biochemistry 32, 14194-202

4 Kusumawati, A, Commes, T, Liautard, JP and Sri Widada, J (1999) Transfection of myelomonocytic cell lines: cellular response to a lipid-based reagent and electroporation. Anal Biochem 269, 219-21

5 Kusumawati, A, Cazevieille, C, Porte, F, Bettache, S, Liautard, JP and Sri Widada, J (2000) Early events and implication of F-actin and annexin I associated structures in the phagocytic uptake of Brucella suis by the J-774A.1 murine cell line and human monocytes. Microb Pathog 28, 343-52

6 Moss, SE, Edwards, HC and Crumpton, MJ (1991) Diversity in the annexin family. Novel Calcium Binding Proteins. Fundamental and Clinical Applications 535-66

7 Pittis, MG and Garcia, RC (1999) Annexins VII and XI are present in a human macrophage like cell line. Differential translocation on FcR-mediated phagocytosis. J Leukoc Biol 66, 845-50

8 Porte, F, De Santa Barbara, P, Phalipou, S, Liautard, JP and Sri Widada, J (1996) Change in the N-terminal domain conformation of annexin I that correlates with liposomes aggregation is impaired by Ser27 to Glu mutation that mimics phosphorylation. Biochim Biophys Acta 1293, 177-84

9 Raynal, P and Pollard, HB (1994) Annexins: the problem of assessing the biological role for a gene family of multifunctional calcium- and phospholipid-binding proteins. Biochim Biophys Acta 1197, 63-93

10 Seemann, J, Weber, K, Osborn, M, Parton, RG and Gerke, V (1996) The association of annexin I with early endosomes is regulated by Ca2+and requires an intact N-terminal domain. Mol Biol Cell 7, 1359-74

11 Stendahl, O, Krause, KH, Krischer, J, Jerström, J, Theler, JM, Clark, RA et al. (1994) Redistribution of intracellular Ca2+stores during phagocytosis in human neutrophils. Science 265, 1439-41

12 Wang, W and Creutz, CE (1994) Role of the amino-terminal domain in regulating interactions of annexin I with membranes: effects of amino-terminal truncation and mutagenesis of the phosphorylation sites. Biochemistry 33, 275-82

Received 15 August 2000; accepted 16 November 2000


ISSN Print: 1065-6995
ISSN Electronic: 1095-8355
Published by Portland Press Limited on behalf of the International Federation for Cell Biology (IFCB)