Journal of Insect Physiology 49 (2003) 545–550 www.elsevier.com/locate/jinsphys Is cell surface calreticulin involved in phagocytosis by insect hemocytes? S. Asgari * , O. Schmidt Department of Applied and Molecular Ecology, The University of Adelaide, Waite Campus, Glen Osmond, South Australia 5064, Australia Received 13 December 2002; received in revised form 5 February 2003; accepted 5 February 2003 Abstract The innate immune system of insects consists of humoral and cellular components involved in the recognition of and responses to intruding foreign micro- or macroorganisms. Several molecules have been identified so far that recognize molecular patterns present on microorganisms, such as lipopolysaccharides, peptidoglycans and lipoteichonic acid. These molecules, acting as opsonins, trigger immune responses such as phagocytosis, nodule formation, melanization and encapsulation. Here, we investigated the role of calreticulin (CRT) present on the surface of Pieris rapae hemocytes in phagocytosis. Comparative phagocytosis assays using yeast cells showed that hemocytes from different insects exhibit significant variation in their phagocytosing potential and relative CRT involvement.  2003 Elsevier Science Ltd. All rights reserved. Keywords: Phagocytosis; Calreticulin; Pieris rapae; Galleria mellonella; Ephestia kuhniella 1. Introduction Insects occupy a diverse range of habitats indicating that they have an effective defence system against poten- tial pathogens and parasites. Although insects are cap- able of inactivating most intruding micro- and macroor- ganisms, they lack the acquired immunity potential of an antigen–antibody recognition system. Proteins enco- ded by an insects’ germline are utilized to recognize molecular patterns found in pathogen-specific features, such as the cell wall of microorganisms (Hoffmann et al., 1999). These include lipopolysaccharide (LPS) bind- ing proteins, peptidoglycan recognition proteins, β-1,3- glucan recognition proteins and C-type lectins (Yu et al., 2002). These components may be associated with hemo- cytes or circulating in the hemolymph as soluble recog- nition molecules. Hemocyte-mediated immune responses include phagocytosis, nodule formation and encapsul- ation of foreign objects (Lavine and Strand, 2002). How- ever, the distinction between humoral and cellular * Corresponding author. Tel.: +61-8-8303-6565; fax: +61-8-8379- 4095. E-mail address: sassan.asgari@adelaide.edu.au (S. Asgari). 0022-1910/03/$ - see front matter  2003 Elsevier Science Ltd. All rights reserved. doi:10.1016/S0022-1910(03)00025-8 immunity is rather arbitrary as components from both interact with each other. Calreticulin (CRT) is a ubiquitous Ca 2+ binding pro- tein with multi-functional properties. It was originally identified as a molecular chaperone associated with the endoplasmic reticulum (Michalak et al., 1992). How- ever, it has since been isolated from various sources as an intra- or extracellular component. CRTs can be sub- divided into three domains each with multiple functions: a N-terminal, a proline-rich (P-rich) and a C-terminal domain (reviewed in Nakhasi et al., 1998). The N-ter- minal domain is involved in protein–protein interactions, metal binding, RNA binding and autoantibody binding. The P-rich domain contains the Ca 2+ binding site and also has lectin binding and chaperone functions. In con- trast to the N-terminal and P-rich domains, the C-ter- minal domain is poorly conserved among CRTs. CRT is also involved in the regulation of cell adhesion (Malhotra et al., 1993), removal of apoptotic cells (Ogden et al., 2001) and is enriched in capsules formed by Galleria mellonella larvae (Choi et al., 2002). CRT has mainly been studied in vertebrates. In this paper, we show that CRT is present on the surface of Pieris rapae hemocytes and its antibody- mediated blocking decreases phagocytosis. However, we