Tumor microenvironment and neurofibromatosis type I: connecting the GAPs.

The human disease von Recklinghausen's neurofibromatosis (Nf1) is one of the most common genetic disorders. It is caused by mutations in the NF1 tumor suppressor gene, which encodes a GTPase activating protein (GAP) that negatively regulates p21-RAS signaling. Dermal and plexiform neurofibromas as well as malignant peripheral nerve sheath tumors and other malignant tumors, are significant complications in Nf1. Neurofibromas are complex tumors and composed mainly of abnormal local cells including Schwann cells, endothelial cells, fibroblasts and additionally a large number of infiltrating inflammatory mast cells. Recent work has indicated a role for the microenvironment in plexiform neurofibroma genesis. The emerging evidence points to mast cells as crucial contributors to neurofibroma tumorigenesis. Therefore, further understanding of the molecular interactions between Schwann cells and their environment will provide tools to develop new therapies aimed at delaying or preventing tumor formation in Nf1 patients.

Lysophosphatidylcholine as a ligand for the immunoregulatory receptor G2A.

Although the biological actions of the cell membrane and serum lipid lysophosphatidylcholine (LPC) in atherosclerosis and systemic autoimmune disease are well recognized, LPC has not been linked to a specific cell-surface receptor. We show that LPC is a high-affinity ligand for G2A, a lymphocyte-expressed G protein-coupled receptor whose genetic ablation results in the development of autoimmunity. Activation of G2A by LPC increased intracellular calcium concentration, induced receptor internalization, activated ERK mitogen-activated protein kinase, and modified migratory responses of Jurkat T lymphocytes. This finding implicates a role for LPC-G2A interaction in the etiology of inflammatory autoimmune disease and atherosclerosis.

Mice lacking the orphan G protein-coupled receptor G2A develop a late-onset autoimmune syndrome.

Mice with a targeted disruption of the gene encoding a lymphoid-expressed orphan G protein-coupled receptor, G2A, demonstrate a normal pattern of T and B lineage differentiation through young adulthood. As G2A-deficient animals age, they develop secondary lymphoid organ enlargement associated with abnormal expansion of both T and B lymphocytes. Older G2A-deficient mice (>1 year) develop a slowly progressive wasting syndrome, characterized by lymphocytic infiltration into various tissues, glomerular immune complex deposition, and anti-nuclear autoantibodies. G2A-deficient T cells are hyperresponsive to TCR stimulation, exhibiting enhanced proliferation and a lower threshold for activation. Our findings demonstrate that G2A plays a critical role in controlling peripheral lymphocyte homeostasis and that its ablation results in the development of a novel, late-onset autoimmune syndrome.

Direct genetic demonstration of G alpha 13 coupling to the orphan G protein-coupled receptor G2A leading to RhoA-dependent actin rearrangement.

G2A is an orphan G protein-coupled receptor (GPCR), expressed predominantly in T and B cells and homologous to a small group of GPCRs of unknown function expressed in lymphoid tissues. G2A is transcriptionally induced in response to diverse stimuli, and its ectopic expression suppresses transformation of B lymphoid precursors by BCR-ABL. G2A induces morphological transformation of NIH 3T3 fibroblasts. Microinjection of constructs encoding G2A into Swiss 3T3 fibroblasts induces actin reorganization into stress fibers that depends on RhoA, but not CDC42 or RAC. G2A elicits RhoA-dependent transcriptional activation of serum response factor. Direct evaluation of RhoA activity demonstrates elevated levels of RhoA-GTP in G2A-expressing cells. Microinjection of embryonic fibroblasts derived from various G alpha knockout mice establishes a requirement for G alpha 13 but not G alpha 12 or G alpha q/11 in G2A-induced actin rearrangement. In conclusion, G2A represents a family of GPCRs expressed in lymphocytes that may link diverse stimuli to cytoskeletal reorganization and transcriptional activation through a pathway involving G alpha 13 and RhoA.

A DNA damage and stress inducible G protein-coupled receptor blocks cells in G2/M.

Cell cycle progression is monitored by highly coordinated checkpoint machinery, which is activated to induce cell cycle arrest until defects like DNA damage are corrected. We have isolated an anti-proliferative cell cycle regulator named G2A (for G2 accumulation), which is predominantly expressed in immature T and B lymphocyte progenitors and is a member of the seven membrane-spanning G protein-coupled receptor family. G2A overexpression attenuates the transformation potential of BCR-ABL and other oncogenes, and leads to accumulation of cells at G2/M independently of p53 and c-Abl. G2A can be induced in lymphocytes and to a lesser extent in nonlymphocyte cell lines or tissues by multiple stimuli including different classes of DNA-damaging agents and serves as a response to damage and cellular stimulation which functions to slow cell cycle progression.

Regional gene therapy with a BMP-2-producing murine stromal cell line induces heterotopic and orthotopic bone formation in rodents.

The ability to continuously deliver osteoinductive proteins to a specific anatomic site would facilitate the treatment of fracture nonunions and other clinical problems associated with bone loss. We have developed a murine model of regional gene therapy. A bone-marrow stromal cell line infected with an adenovirus expressing recombinant bone morphogenetic protein-2 cDNA secreted biologically active bone morphogenetic protein-2. These bone morphogenetic protein-2-producing cells were able to induce abundant heterotopic bone formation when implanted into the quadriceps muscle of severe combined immune deficient mice and also successfully healed large segmental femoral defects in nude rats. These studies demonstrate that regional gene therapy with continuous delivery of osteoinductive factors to a specific anatomic site can enhance the formation and repair of bone.