Cloning and functional studies of a novel gene aberrantly expressed in RB-deficient embryos

The tumor suppressor RE regulates diverse cellular processes such as G1/S t ransition, cell differentiation, and cell survival. Indeed, Rb-knockout mic e exhibit phenotypes including ectopic mitosis, defective differentiation, and extensive apoptosis in the neurons. Using differential display, a novel gene, Rig-1, was isolated based on its elevated expression in the hindbrai n and spinal cord of Rb-knockout embryos. The longest open reading frame of Rig-1 encoded a polypeptide that consists of a putative extracellular segm ent with five immunoglobulin-like domains and three fibronectin III-like do mains, a putative transmembrane domain, and a distinct intracellular segmen t. The Rig-1 sequence was 40% identical to the recently identified roundabo ut protein. Consistent with the predicted transmembrane nature of the prote in, Rig-i protein was present in the membranous fraction. Antisera raised a gainst the putative extracellular and intracellular segments of Rig-i react ed with an similar to 210-kDa protein in mouse embryonic CNS. Rig-i mRNA wa s transiently expressed in the embryonic hindbrain and spinal cord. Elevate d levels of Rig-i mRNA and protein were found in Rb-/- embryos. Ectopic exp ression of a transmembrane form of Rig-1, but not the secreted form, promot ed neuronal cell entrance to S phase and repressed the expression of a mark er of differentiated neuron, T alpha 1 tubulin. Thus Rig-1, a possible dist ant relative of roundabout, may mediate some of the pleiotropic roles of RE in the developing neurons. (C) 1999 Academic Press.