Cellular Microbiology (2004) 6(7), 609–623 doi:10.1111/j.1462-5822.2004.00386.x © 2004 Blackwell Publishing Ltd Blackwell Science, LtdOxford, UKCMICellular Microbiology1462-5814Blackwell Publishing Ltd, 20046 7609623Original ArticleD. Smith et al. targeting of protein L Received 18 December, 2003; accepted 23 December, 2003. *For correspondence. E-mail mary-jo.wick@immuno.gu.se; Tel. (+46) 31 342 4602; Fax (+46) 31 342 4621. † Present address: Department of Clinical Immunology, Göteborg University, Guldhedsgatan 10, 413 46 Göteborg, Sweden. Whole-body autoradiography reveals that the Peptostreptococcus magnus immunoglobulin-binding domains of protein L preferentially target B lymphocytes in the spleen and lymph nodes in vivo David Smith, 1 Roland D'Argy, 2 Mats Nilsson, 2 Ulf Yrlid, 1 James de Jersey, 1 Lars Björck 3 and Mary Jo Wick 1 * † 1 Section for Immunology, Department of Cell and Molecular Biology, Lund University, Lund, Sweden. 2 Active Biotech Research AB, Lund, Sweden. 3 Molecular Pathogenesis, Department of Cell and Molecular Biology, Lund University, Lund, Sweden. Summary Protein L is an immunoglobulin (Ig)-binding protein produced by the Gram-positive bacterium Peptostrep- tococcus magnus that interacts with the variable region of Ig k light chains. The Ig light chain-binding capacity of protein L gives it the potential to interact with cells expressing surface Ig such as B cells. The present study was performed to address the in vivo trafficking of protein L at both the organ and the cellular level. Using the powerful technique of whole- body autoradiography in a murine model system, we demonstrate specific targeting of protein L to second- ary lymphoid tissues in whole-animal analysis. The observed targeting depends on the capacity to inter- act with murine Ig, as tissue targeting was not appar- ent in mice given protein H, an Ig-binding protein produced by Streptococcus pyogenes with affinity for human but not murine Ig. Tissue targeting data were combined with flow cytometry analysis, which dem- onstrated the capacity of protein L to target and acti- vate B lymphocytes in vivo. B cells targeted by protein L had increased surface expression of CD86 and MHC-II, and protein L was present in vacuolar com- partments of B cells. Protein L did not bind T cells or natural killer cells but had some capacity to target dendritic cells and macrophages. The data show that protein L preferentially targets secondary lymphoid organs, and activates and is internalized by B cells in vivo. Furthermore, the observed tissue and cell tar- geting properties require an affinity for murine Ig. These data support the potential use of this Ig- binding protein as a targeting approach to deliver agents to defined cell populations in vivo. Introduction Several genera of Gram-positive bacteria including Staphylococcus aureus, Streptococcus pyogenes and Peptostreptococcus magnus produce surface proteins that bind immunoglobulin (Ig) molecules of several mam- malian species (Boyle, 1990). Protein L produced by the Gram-positive anaerobic species P. magnus is one such Ig-binding protein (Björck, 1988). P. magnus is a member of the indigenous flora of the skin, oral cavities, the gas- trointestinal tract and the urogenital tract. However, this species is also the causative agent of a variety of infec- tions (Sutter et al., 1985). Approximately 10% of clinical isolates of P. magnus carry and express the gene encod- ing protein L, and the majority of these isolates are from patients with gynaecological infections. This suggests a role for protein L in the virulence of P. magnus (Kastern et al., 1990). Protein L is distinct from the other bacterial Ig-binding proteins because of its unique ability to bind Ig light chains and, in particular, to interact with the variable region of the Ig k light chain (Björck, 1988; Kastern et al., 1992; Nilson et al., 1992; de Château et al., 1993; Wikström et al., 1995; Graille et al., 2001). It binds specifically to the outer framework region of Ig k light chains of the I, III and IV subgroups but does not interact with the Vk II subgroup or any l light chain subgroups (Nilson et al., 1992; Graille et al., 2001). Moreover, this binding is independent of the antigen-binding capacity of Ig (Enokizono et al., 1997; Graille et al., 2001). Protein L activates mast cells and basophils by binding to the k light chains of IgE associated with the Fc receptors of these cells, thereby cross-linking IgE on the cell surface. As a result, histamine, interleukin (IL)-4 and IL-13 are released, which could contribute to the virulence of the bacteria (Patella et al., 1990; Genovese et al., 2003).