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.

Sphingosylphosphorylcholine (SPC) and lysophosphatidylcholine (LPC) are bioactive lipid molecules involved in numerous biological processes. We have recently identified ovarian cancer G protein-coupled receptor 1 (OGR1) as a specific and high affinity receptor for SPC, and G2A as a receptor with high affinity for LPC, but low affinity for SPC. Among G protein-coupled receptors, GPR4 shares highest sequence homology with OGR1 (51%). In this work, we have identified GPR4 as not only another high affinity receptor for SPC, but also a receptor for LPC, albeit of lower affinity. Both Taken together, our data indicate that GPR4 is a receptor with high affinity to SPC and low affinity to LPC, and that multiple cellular functions can be transduced via this receptor.

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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.

The G2A receptor was originally identified by virtue of its transcriptional induction in murine B lymphoid cells in response to oncogenic transformation and treatment with various DNA-damaging agents. While preliminary characterization of cellular responses to G2A overexpression in fibroblastic cell lines suggested that this receptor may negatively regulate cell growth under conditions of proliferative and genotoxic stress, subsequent studies driven by the discovery of lysophosphatidylcholine (LPC) as a regulator of G2A signaling in immunoregulatory cells point to an important role for this receptor in innate and adaptive immunity.

BTB/POZ-domain C 2 H 2 zinc(Zn)-®nger proteins are encoded by a subfamily of genes related to the Drosophila gap gene kruÈppel. To date, two such proteins, PLZF and LAZ-3/BCL-6, have been implicated in oncogenesis. We have now identi®ed a new member of this gene subfamily which encodes a 62 kDa Zn-®nger protein, termed LRF, with a BTB/POZ domain highly similar to that of PLZF. Both human and mouse LRF genes, which localized to syntenic chromosomal regions (19p13.3 and 10B5.3, respectively), were widely expressed in adult tissues and cell lines. At approximately 9.5 ± 10.0 days of embryonic development, the mouse LRF gene was expressed in the limb buds, pharyngeal arches, tail bud, placenta and neural tube. The LRF protein associated in vivo with LAZ-3/BCL-6, but not with PLZF to which it was more related. Although the LRF, or LAZ-3/BCL-6, BTB/POZ domain could readily homodimerize, no heterodimerization was detected in vivo between the LRF and LAZ-3/BCL-6 BTB/POZ domains and interaction between full length LRF and LAZ-3/BCL-6 required the presence of both the BTB/POZ domain and Zn-®ngers in each partner protein. As expected from the above results, LRF and LAZ-3/BCL-6 also colocalized with each other in the nucleus. Taken together, our ®ndings suggest that BTB/ POZ-domain Zn-®nger proteins may function as homo and heterodimeric complexes whose formation, and hence the resultant eect on transcription of their downstream target genes, is determined by the levels and expression domains of a given partner protein.

In acute promyelocytic leukemia (APL), the typical t(15;17) and the rare t(11;17) translocations express, respectively, the PML͞RAR␣ and PLZF͞RAR␣ fusion proteins (where RAR␣ is retinoic acid receptor ␣). Herein, we demonstrate that the PLZF and PML proteins interact with each other and colocalize onto nuclear bodies (NBs). Furthermore, induction of PML expression by interferons leads to a recruitment of PLZF onto NBs without increase in the levels of the PLZF protein. PML͞RAR␣ and PLZF͞RAR␣ localize to the same microspeckled nuclear domains that appear to be common targets for the two fusion proteins in APL. Although PLZF͞RAR␣ does not affect the localization of PML, PML͞ RAR␣ delocalizes the endogenous PLZF protein in t(15;17)-positive NB4 cells, pointing to a hierarchy in the nuclear targeting of these proteins. Thus, our results unify the molecular pathogenesis of APL with at least two different RAR␣ gene translocations and stress the importance of alterations of PLZF and RAR␣ nuclear localizations in this disease.Acute promyelocytic leukemia (APL) represents approximately 10% of all adult acute myeloid leukemias (1). The molecular pathogenesis of APL is, at least in part, associated with the disruption of the retinoic acid receptor ␣ (RAR␣) gene through its fusion to one of four different loci (2-7). These translocations result in the expression of chimeric RAR␣ fusion proteins that retain the DNA and ligand binding domains of the receptor and gain a dimerization domain from the fusion partner. Paradoxically, APL is the first human malignancy that may undergo complete remission in response to differentiation therapy with all-trans-retinoic acid (RA). The molecular basis of these remissions is still disputed.The majority of APL cases, and all cases that consistently respond to RA treatment, possess the t(15;17) translocation that fuses the PML and RAR␣ genes (3,(8)(9)(10)(11)(12). PML is a member of a functionally diverse gene family that encodes proteins characterized by the presence of a N-terminal C 3 HC 4 RING-finger motif (13), followed by one or two cysteine-rich regions (B boxes) and a coiled-coil protein dimerization interface. The function of PML is unknown, but up-regulation of its expression by interferons (IFNs) (14-16) and its negative effect on cell growth and cellular transformation by cooperating oncogenes (17-19) suggest a role in growth control. The product of the wild-type PML gene is a phosphoprotein (20) that localizes both in the nucleoplasm and in the specific multiprotein structures called PML nuclear bodies (NBs) (20-24). The PML͞RAR␣ fusion protein, which is expressed in APL cells as a result of t(15;17), contains all predicted PML structural motifs and is able to delocalize the wild-type PML and other NB components onto discrete microspeckled nuclear structures (21-24). It is still unclear which role, if any, disruption of NBs and͞or establishment of microspeckled structures play in cellular transformation. Nevertheless, complete restoration of NBs upon RA treatment in NB4 ...

The Philadelphia translocation commonly observed in chronic myeloid leukaemia (CML) and a proportion of cases of acute leukaemia results in the creation of a chimeric fusion protein, BCR‐ABL. The fusion protein exhibits an elevated tyrosine kinase activity as compared to normal ABL. Using a temperature sensitive mutant of p210 BCR‐ABL (ts‐p210) we find that the primary effect of BCR‐ABL expression in an IL‐3 dependent cell line is to prolong survival following growth factor withdrawal; only a small proportion of cells remain viable and rapidly evolve to complete growth factor independence. During passage in the presence of IL‐3 at the temperature permissive for kinase activity, ts‐p210 expressing cultures become dominated by completely growth factor independent cells within 10‐30 days. There is also a significant difference between BCR‐ABL and IL‐3 mediated signalling with respect to the MAP kinase pathway; in contrast to IL‐3 stimulation or v‐ABL expression, BCR‐ABL does not signal ERK 2 (MAP 2 kinase) activation, underlining the apparent inability of BCR‐ABL to deliver an immediate proliferative signal in Ba/F3 cells. Our data suggest that growth factor independence does not simply reflect the convergence of BCR‐ABL and IL‐3 mediated signalling pathways and its development, at least in Ba/F3 cells, requires prolonged exposure to BCR‐ABL kinase activity. We suggest that the myeloid expansion characteristic of CML may result from the prolongation of survival of myeloid progenitor cells under conditions of limiting growth factor rather than their uncontrolled proliferation.