Edited by John M. Denu EPHB6 is a member of the erythropoietin-producing hepatocellular kinase (EPH) family and a receptor tyrosine kinase with a dead kinase domain. It is involved in blood pressure regulation and adrenal gland catecholamine (CAT) secretion, but several facets of EPHB6-mediated CAT regulation are unclear. In this study, using biochemical, quantitative RT-PCR, immunoblotting, and gene microarray assays, we found that EPHB6 up-regulates CAT biosynthesis in adrenal gland chromaffin cells (AGCCs). We observed that epinephrine content is reduced in the AGCCs from male Ephb6-KO mice, caused by decreased expression of tyrosine hydroxylase, the rate-limiting enzyme in CAT biosynthesis. We demonstrate that the signaling pathway from EPHB6 to tyrosine hydroxylase expression in AGCCs involves Rac family small GTPase 1 (RAC1), MAP kinase kinase 7 (MKK7), c-Jun N-terminal kinase (JNK), proto-oncogene c-Jun, activator protein 1 (AP1), and early growth response 1 (EGR1). On the other hand, signaling via extracellular signalregulated kinase (ERK1/2), p38 mitogen-activated protein kinase, and ELK1, ETS transcription factor (ELK1) was not affected by EPHB6 deletion. We further report that EPHB6's effect on AGCCs was via reverse signaling through ephrin B1 and that EPHB6 acted in concert with the nongenomic effect of testosterone to control CAT biosynthesis. Our findings elucidate the mechanisms by which EPHB6 modulates CAT biosynthesis and identify potential therapeutic targets for diseases, such as hypertension, caused by dysfunctional CAT biosynthesis.EPHB6 is a member of erythropoietin-producing hepatocellular kinases (EPHs), 3 the largest family of receptor tyrosine kinases (1, 2). The ligands of EPH are called ephrins (EFNs), which are also membrane proteins. EFNs can trigger EPH signaling by canonical forward signaling, i.e. from ligand EFNs to receptor EPHs. However, EFNs can also receive signaling from EPHs and transduce signals into cells, and such noncanonical action (i.e. from EPHs to EFNs) is called reverse signaling (2). The interaction between EPHs and EFNs is promiscuous; one EPH can bind to multiple EFNs and one EFN to multiple EPHs. In general, EPHA family members bind to EFNAs, and EPHB family members binds to EFNBs (2).EPHs/EFNs function in many organs and systems (2). We first reported the critical involvement of EPHs and EFNs in the immune system (3-15). In the past 5 years, we have demonstrated in a series of publications (16 -25) that EPHs/EFNs are involved in regulating blood pressure (BP), which was previously unknown. We reported, using gene knockout mouse models, that although EPHB6, EFNB1, EFNB3, and EPHA4 deletion results in BP elevation (16,17,20,25), EPHB4 and EFNB2 deletion lowers it (18,19). Thus, members of EPHBs/ EFNBs are a novel yin and yang system that fine-tunes BP homeostasis. In all such cases, sex hormones act in concert with these EPHs/EFNs to control BP. Some of the findings from the mouse model have been corroborated by human genetic studies, in which we revealed that s...