The Journal of Immunology, Vol 145, Issue 9 2967-2973, Copyright © 1990 by American Association of Immunologists

ARTICLES

The lymphocyte-specific protein LSP1 is associated with the cytoskeleton and co-caps with membrane IgM

DP Klein, S Galea and J Jongstra
Department of Immunology, University of Toronto, Ontario, Canada.

LSP1 is a lymphocyte-specific intracellular Ca2(+)-binding protein. We found previously that a fraction of the total cellular pool of LSP1 protein accumulates at or near the cytoplasmic face of the plasma membrane. LSP1 protein was also shown to be present in the cytoplasm. Here we report that approximately 10% of the total intracellular LSP1 protein is associated with the Nonidet P-40 insoluble cytoskeleton of the mIgM+, mIgD+ B lymphoma cell line BAL17. Variation in conditions of extraction did not alter this value. To rule out the possibility that LSP1 associates with the nucleus that is also present in the detergent insoluble pellet, we prepared a separate nuclear fraction essentially free of cytoskeletal material and found only trace amounts of LSP1 protein. After accounting for yield losses during subcellular fractionation by measuring the recovery of 125I-labeled membrane IgM, or of the cytoplasmic marker enzyme lactate dehydrogenase activity, the LSP1 in membrane fractions was calculated to represent approximately 30% of the total cellular LSP1 and cytoplasmic LSP1 accounted for approximately 55% of the total. Approximately 75% of the plasma membrane LSP1 protein was soluble in 1% Nonidet P-40 containing buffer, indicating that the majority of the LSP1 in the plasma membrane fraction was distinct from the cytoskeletal LSP1 protein. The preparation of membrane fractions in the presence of 1 M NaCl, or washing of membranes in 3 M KCl did not diminish the levels of membrane LSP1. These results show the existence of three discrete intracellular LSP1 pools. Double label immunofluorescence studies showed that the peripheral ring-like distribution of LSP1 in BAL17 cells became a distinct cap upon cross-linking the mIgM. These intracellular LSP1 caps were always found to be located directly underneath the mIgM caps.

This article has been cited by other articles:

				L. Liu, D. C. Cara, J. Kaur, E. Raharjo, S. C. Mullaly, J. Jongstra-Bilen, J. Jongstra, and P. Kubes LSP1 is an endothelial gatekeeper of leukocyte transendothelial migration J. Exp. Med., February 7, 2005; 201(3): 409 - 418. [Abstract] [Full Text] [PDF]

				R. E. Harrison, B. A. Sikorski, and J. Jongstra Leukocyte-specific protein 1 targets the ERK/MAP kinase scaffold protein KSR and MEK1 and ERK2 to the actin cytoskeleton J. Cell Sci., April 15, 2004; 117(10): 2151 - 2157. [Abstract] [Full Text] [PDF]

				C. Wang, H. Hayashi, R. Harrison, B. Chiu, J. R. Chan, H. L. Ostergaard, R. D. Inman, J. Jongstra, M. I. Cybulsky, and J. Jongstra-Bilen Modulation of Mac-1 (CD11b/CD18)-Mediated Adhesion by the Leukocyte-Specific Protein 1 Is Key to Its Role in Neutrophil Polarization and Chemotaxis J. Immunol., July 1, 2002; 169(1): 415 - 423. [Abstract] [Full Text] [PDF]

				M. Hannigan, L. Zhan, Y. Ai, and C.-K. Huang Leukocyte-specific gene 1 protein (LSP1) is involved in chemokine KC-activated cytoskeletal reorganization in murine neutrophils in vitro J. Leukoc. Biol., March 1, 2001; 69(3): 497 - 504. [Abstract] [Full Text]

				J. Jongstra-Bilen, V. L. Misener, C. Wang, H. Ginzberg, A. Auerbach, A. L. Joyner, G. P. Downey, and J. Jongstra LSP1 modulates leukocyte populations in resting and inflamed peritoneum Blood, September 1, 2000; 96(5): 1827 - 1835. [Abstract] [Full Text] [PDF]

				Y. Yamashita, K. Oritani, E. K. Miyoshi, R. Wall, M. Bernfield, and P. W. Kincade Syndecan-4 Is Expressed by B Lineage Lymphocytes and Can Transmit a Signal for Formation of Dendritic Processes J. Immunol., May 15, 1999; 162(10): 5940 - 5948. [Abstract] [Full Text] [PDF]

				T. H. Howard, J. Hartwig, and C. Cunningham Lymphocyte-Specific Protein 1 Expression in Eukaryotic Cells Reproduces the Morphologic and Motile Abnormality of NAD 47/89 Neutrophils Blood, June 15, 1998; 91(12): 4786 - 4795. [Abstract] [Full Text] [PDF]

				C.-K. Huang, L. Zhan, Y. Ai, and J. Jongstra LSP1 Is the Major Substrate for Mitogen-activated Protein Kinase-activated Protein Kinase 2in Human Neutrophils J. Biol. Chem., January 3, 1997; 272(1): 17 - 19. [Abstract] [Full Text] [PDF]

				V. Misener, C Hui, I. Malapitan, M. Ittel, A. Joyner, and J Jongstra Expression of mouse LSP1/S37 isoforms. S37 is expressed in embryonic mesenchymal cells J. Cell Sci., January 12, 1994; 107(12): 3591 - 3600. [Abstract] [PDF]

				A. Khan, J. Steiner, M. Klein, M. Schneider, and S. Snyder IP3 receptor: localization to plasma membrane of T cells and cocapping with the T cell receptor Science, August 7, 1992; 257(5071): 815 - 818. [Abstract] [PDF]

				M.-Y. Cao, F. Shinjo, S. Heinrichs, J.-W. Soh, J. Jongstra-Bilen, and J. Jongstra Inhibition of Anti-IgM-induced Translocation of Protein Kinase C beta I Inhibits ERK2 Activation and Increases Apoptosis J. Biol. Chem., June 29, 2001; 276(27): 24506 - 24510. [Abstract] [Full Text] [PDF]