Synaptic clustering of GABA A receptors is important for the function of inhibitory synapses, influencing synapse strength and, consequently, the balance of excitation and inhibition in the brain. Presynaptic terminals are known to induce GABA A receptor clustering during synaptogenesis, but the mechanisms of cluster formation and maintenance are not known. To study how presynaptic neurons direct the formation of GABA A receptor clusters, we have investigated GABA A receptor localization in postsynaptic cells that fail to receive presynaptic contacts in Caenorhabditis elegans. Postsynaptic muscles in C. elegans receive acetylcholine and GABA motor innervation, and GABA A receptors cluster opposite GABA terminals. Selective loss of GABA inputs caused GABA A receptors to be diffusely distributed at or near the muscle cell surface, confirming that GABA presynaptic terminals induce GABA A receptor clustering. In contrast, selective loss of acetylcholine innervation had no effect on GABA A receptor localization. However, loss of both GABA and acetylcholine inputs together caused GABA A receptors to traffic to intracellular autophagosomes. Autophagosomes normally transport bulk cytoplasm to the lysosome for degradation. However, we show that GABA A receptors traffic to autophagosomes after endocytic removal from the cell surface and that acetylcholine receptors in the same cells do not traffic to autophagosomes. Thus, autophagy can degrade cell-surface receptors and can do so selectively. Our results show that presynaptic terminals induce GABA A receptor clustering by independently controlling synaptic localization and surface stability of GABA A receptors. They also demonstrate a novel function for autophagy in GABA A receptor degradative trafficking.

Melanoma growth stimulatory activity factor (MGSA) is a polypeptide which was initially isolated from Hs294 human melanoma cells. Its sequence is identical to the deduced amino acid sequence of the human gro cDNA, isolated from a human tumor cell line. MGSA stimulates the proliferation of malignant melanoma cells, but its function for normal cells has not been defined. Here we report that human umbilical vein endothelial cells are capable of synthesizing and secreting MGSA. The expression and secretion of MGSA are strongly induced by factors often involved in inflammation such as IL‐1, TNF, LPS and thrombin. The induction of MGSA mRNA is dose and time dependent and is independent of new protein synthesis. This stimulation could be mimicked by TPA, suggesting that the action could be mediated through activation of protein kinase C. Furthermore, addition of MGSA to the endothelial cell cultures induces gro/MGSA gene expression, implying that an autocrine mechanism exists. Our data suggest that the protein encoded by gro/MGSA mRNA may play a role in inflammation and exert its effects on endothelial cells in an autocrine fashion.

ABSTRACT:Cytochromes P450 (P450s) contribute to the metabolic activation and inactivation of various endogenous substrates. Despite years of research, the physiological role of CYP2S1 remains unknown. CYP2S1 has demonstrated NADPH P450-reductase-independent metabolism of cyclooxygenase (COX)-derived prostaglandins [e.g., prostaglandin G 2 (PGG 2 )] at nanomolar concentrations. Arachidonic acid is converted to prostaglandin precursors [PGG 2 and prostaglandin H 2 (PGH 2 )] through COX. These precursors are used to synthesize numerous prostanoids, including PGE 2 . Prostaglandin E 2 (PGE 2 ) promotes cell proliferation and cell migration and inhibits apoptosis. CYP2S1 metabolism of PGG 2 presumably sequesters PGG 2 and PGH 2 , making them unavailable for synthesis of prostanoids such as PGE 2 . Whether CYP2S1 contributes to prostaglandin metabolism and influences cell physiological remains to be determined. The purpose of this study was to evaluate the physiological role of CYP2S1, if any, in human bronchial epithelial cells [SV40-derived bronchial epithelial cell line (BEAS-2B)]. To do this, we used small interfering RNA to deplete CYP2S1 mRNA and protein by approximately 75% and evaluated the impact of CYP2S1 depletion on cell proliferation and migration. CYP2S1 depletion enhanced both cell proliferation and migration in BEAS-2B cells. Consistent with the proposed role of CYP2S1 in PGE 2 synthesis, the reduction in CYP2S1 expression doubled intracellular PGE 2 levels. Pharmacological administration of PGE 2 enhanced cell proliferation in BEAS-2B cells but failed to promote migration. Our data reveal an important role for CYP2S1 in the regulation of cell proliferation and migration, occurring in part through modulation of prostaglandin synthesis.