Developmental expression and biochemical analysis of conulin, a protein secreted from a subset of neuronal growth cones

Abstract

In this report, we analyze the developmental pattern of expression of a new grasshopper protein, Conulin, using the monoclonal antibody 7D2 on whole-mount embryos and dissociated neurons. We also have examined its biochemical properties by immunoblot analysis. Conulin is a protein expressed by a subset of neurons in the grasshopper embryo. The monoclonal antibody 7D2 recognizes Conulin as an M(r) 190 x 10(3) protein that is found in both the soluble and membrane-bound fractions of embryonic proteins. The membrane association is disrupted by alkaline pH and high ionic strength. Conulin first is expressed and stored in vesicles inside the cell bodies and axons of central and peripheral neurons. Later, Conulin is detected on the cell surface, but exclusively in the central nervous system neuropil. This expression is confined to a subset of nerve growth cones. Conulin is detected on growth cones only after pioneer neurons have outlined the axonal scaffold. Immunocytochemistry on cultured embryonic neurons demonstrates that the neurons have the autonomous ability to target Conulin to the growth cones. The protein is secreted but remains transiently associated with the growth cone plasma membrane. The discovery of Conulin confirms the existence of proteins specific for the nerve growth cone. Its transitory presence during axonogenesis in only a subset of follower growth cones suggests that Conulin is involved in guidance through selective fasciculation with pre-existing axons within the ganglionic neuropil.