Molecular characterization of a thrombospondin-related anonymous protein homologue in Neospora caninum

Thrombospondin-related anonymous protein (TRAP) family members participate in attachment and invasion of host cells by apicomplexan parasites. A TRAP homologue in Neospora caninum strain Nc-l (NcMIC2) was cloned, sequenced an d found to be 61% identical (75% similar) at the amino acid level to Toxopl asma gondii MIC2 (TgMIC2). Similar to TgMIC2, the predicted amino acid sequ ence of NcMIC2 contains one integrin-like domain (I or A domain), five thro mbospondin (TSP) repeats, a putative transmembrane spanning region and intr acellular C-terminus, and was localized to micronemes by cryo-immunoelectro n microscopy. The secretion of NcMIC2 was temperature dependent and was ind uced at or above 25 degrees C. The secreted form of NcMIC2 released into th e medium was found to be proteolytically processed such that it lacked the C-terminal domain. Secretion of NcMIC2 was regulated by calcium, since seve ral agents which raise intracellular calcium levels were shown to promote N cMIC2 secretion and chelation of [Ca2+](i) abrogated release. As a member o f the growing family of apicomplexan TRAP proteins, NcMIC2 may play an impo rtant role in attachment and invasion by N. caninum into host cells. (C) 20 00 Elsevier Science B.V. All rights reserved.