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www.sbsonline.org 789 INTRODUCTION G-protein-coupled receptors (Gpcrs) constitute the larGest family of cell surface receptors that respond to a wide range of extracellular stimuli and are involved in virtually all known physiological processes in mammals.1 Although they respond to diverse agonists, the structural and functional features of these receptors are remarkably conserved. All GPCRs have 7 transmembrane domains and activate a variety of G-protein signaling pathways upon agonist stimulation.1,2 There are 4 major Gα protein subfamilies: Gα s , Gα i/o , Gα q , and Gα 12/13 . Gα s stimulates adenylyl cyclase and increases cyclic AMP (cAMP) production, whereas Gα i inhibits cAMP production. Activation of Gα q stimulates phospholipase C β followed by production of diacylglycerol and inositol phosphate, which initiates calcium release from intracellular stores.Even though GPCRs constitute only a small fraction (15%) of total druggable genes, they account for more than 30% of targets of marketed small-molecule drugs.3 Due to the enormous historical success of GPCR drug targets, the search for new drugs targeting GPCRs is still the prime focus of many pharmaceutical companies. The completion of the human genome sequence project revealed more than 800 GPCR sequences, of which around 140 GPCRs, excluding olfactory receptors, are still classified as orphan receptors, (i.e., receptors with ligand and physiological functions unknown).4,5 Some of these uncharacterized orphan receptors are likely to be novel therapeutic targets of the future.6,7 Over the past 2 decades, more than 100 receptors have been deorphanized (i.e., paired with their endogenous ligands). 6,8 The deorphanization of once-orphaned GPCRs has proven to be fruitful and already unveiled several exciting novel drug targets for treating human diseases, including neurological and neurodegenerative disorders, sleep disorders, inflammation, and rheumatoid arthritis. 6,7,[9][10][11] Human GPR139 (also called hGPCR12, GPRg1, and PGR3) is an orphan receptor that was first described as a receptor homologous to thyrotropin-releasing hormone receptor. 12,13 GPR139 is extremely well conserved, as the protein sequence of the human receptor is 96% identical to both mouse and rat orthologs. GPR139 shows distant homology to a variety of peptide and chemokine receptors with identities in the range of

Transfer of sunflower (Helianthus annuus L. cv Russian Mammoth) seedlings from complete nutrient solution to solutions deficient in either boron or calcium resulted in a steady decline in the rate of auxin transport, compared to seedlings that remained in the complete solution. In seedlings transferred to solutions deficient in both B and Ca, the decline in auxin transport was greater than seedlings deficient in only one element. The transfer of B-or Ca-deficient seedlings back to the complete solution prevented further decline in auxin transport, but auxin transport did not increase to the same level as seedlings maintained in complete solution. The significant reduction in auxin transport during the early stages of B or Ca deficiency was not related to (a) reduced growth rate of the hypocotyl, (b) increased acropetal movement of auxin, or (c) lack of respiratory substrates in the hypocotyl. In addition, no difference was found in the water-extractable total and ionic Ca in B-deficient and control nondeficient hypocotyls, indicating a direct effect of B on auxin transport, rather than indirectly by affecting Ca absorption. The rate of auxin transport in hypocotyls deficient in either B or Ca, was inversely correlated with K' leakage and rate of respiration. The data presented strongly support the view that there are separate sites for B and Ca in the basipetal transport of the plant hormone indoleacetic acid.Recently it was demonstrated that Ca is essential in the basipetal transport or secretion of auxin in sunflower hypocotyl segments (5, 7). It was hypothesized that Ca probably functions in the same way that this element does in the secretion of many kinds of substances in animal cells (21 the hypocotyl 1800 relative to gravity. Also, the acropetal efflux of Ca was inhibited by cyanide and low temperature, just as the basipetal transport of auxin (4).Although these findings support our hypothesis, it can be argued that our observations regarding the Ca-IAA transport relationship still appears to be more apparent than real. To probe this hypothesis further, we sought the use of other model systems. The main question is whether the findings of a Ca requirement in auxin transport, and the auxin promotion of Ca efflux at the hypocotyl level, is an accurate representation ofthe phenomenon at the cell level. It is possible that these findings are the direct effect of the overall disturbance in the hypocotyl, such as apical necrosis, increase membrane permeability, etc., resulting from Ca deficiency, and thus only indirectly to the deficiency of Ca. To this end we used the B-deficient seedling for the following reasons.Both B and Ca are considered immobile elements in plant systems (19,20). Deficiency or withdrawal of B or Ca from the root medium leads to necrosis of young tissues in roots and shoots, leaving the older portions of the plant relatively unaffected. The avid binding ofCa to the cell wall (23), and of H3BO3 to membrane and cell wall components with cis-hydroxyl configurations (14), together wit...

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Sunflower (Helianthus annuus L. cv Russian Mammoth) hypocotyl segments deficient in either B or Ca exhibited a higher rate of potassium leakage, compared to nondeficient segments. Potassium leakage, used here as an indication of membrane integrity, was completely reversed by the addition of H3BO3 or Ca(NO3)2 to the incubation medium of the Bdeficient or Ca-deficient hypocotyl segments, respectively. This role of B and Ca in membrane integrity, which may be important in the entry and exit of auxin in cells, is identified as the first site of action for each of these two essential elements in the basipetal secretion of auxin. A second site for B is postulated because auxin transport was not restored, even when K' leakage has been completely reversed to the nondeficient level, when B-deficient hypocotyls were incubated in B solution. This lack of reversibility of auxin transport implied that the incubation for 2 h in B solution was not enough to restore the auxin transport process. However, since the transfer of B-deficient seedlings to B solutions prevented further deterioration of auxin transport, these observations suggest that: (a) either an intact seedling, or a longer period of incubation of the hypocotyl in B solution, is required for the synthesis or maintenance of the functional second site for B; (b) B is probably essential in the synthesis of a ligand, which may or may not be needed to bind B, but which is essential in the basipetal transport of auxin. The second site for Ca in auxin transport, is indicated by the complete reversal of its inhibition in Ca-deficient hypocotyl, when incubated in Ca solution. The second site for Ca is thought to be directly involved in the secretion of auxin, in which Ca probably plays the role of a second messenger, as in stimulus-response coupling. The two sites for Ca can be distinguished from each other by their cation specificity. The requirement for Ca in the first site can be substituted by other divalent cations, while the second site is highly specific for Ca.In the preceding article (17) it was shown that sunflower hypocotyl segments from seedlings deficient in either B or Ca had (a) a higher rate ofK+ leakage, (b) a higher rate ofrespiration, and (c) a lower rate of basipetal auxin transport than hypocotyls from control, nondeficient seedlings. Although similar processes were being affected by the deficiency of either element, separate sites of action for B and for Ca were postulated since the only instance wherein any of the inhibited process can be reversed was by the transfer of the seedling to a solution containing the deficient element. (17). Basically, the seeds were germinated between paper towels moistened with distilled H20 and after 1 d the seeds with good radicle break were transferred to one-fourth strength Hoagland solution A (6) containing 0.023 mM H3BO3 (0.25 gg/ml B). After the 4th d, the seedlings were transferred to fresh Hoagland solution, or to solutions lacking B, Ca, or both elements.Auxin transport was measured in 5 mm hypocotyl se...