Characterization of mouse brain-specific angiogenesis inhibitor 1 (BAI1) and phytanoyl-CoA alpha-hydroxylase-associated protein 1, a novel BAI1-binding protein

Previously, PAHX-API (PAHX-associated protein 1) was isolated as a novel protein to interact with Refsum disease gene product (phytanoyl-CoA alpha-hydroxylase, PAHX) and specifically expressed in mouse brain. PAHX-API is also suggested to be involved in the development of the central neurologic deficits of Refsum disease. To clarify its function, we have searched for proteins that associate with PAHX-AP1 via yeast two-hybrid system. We found that PAHX-AP1 interacts with the cytoplasmic region of human brain-specific angiogenesis inhibitor 1 (hBAI1), and isolated murine homolog of hBAII. Structural analysis of the PAHX-API with three reported hBAI-associated proteins (BAP) revealed no homology among them, and we designated PAHX-API as BAP4. The ability of BAP4 to interact with BAI1 was confirmed by pulling-down BAI1 with GST-BAP4 protein and immunoprecipitation study using brain lysate. Northern and Western blot analyses demonstrated a unique pattern of BAI1 expression in the brain. The peak level of BAI1 was observed 10 days after birth. In situ hybridization analyses of the brain showed the same localization of BAI1 as BAP4, such as most neurons of cerebral cortex, hippocampus, and V,VI, VII, VIII, and XII nuclei. Because BAI1 possessed thrombospondin-type I repeats: in its extracellular region, changes of BAI1 expression were examined in the focal cerebral ischemia model. The BAI1 expression decreased on the ischemic side after 24 h but BAP4 was not changed after the time-course of ischemia. Our results indicate that expression and localization of BAI1 in the brain is correlated with BAP4, and that BAI1 is involved in inhibition of angiogenesis and neuronal differentiation. (C) 2001 Elsevier Science B.V. All rights reserved.