Cinnamoyl-CoA reductase (CCR) catalyzes the penultimate step in monolignol biosynthesis. We show that downregulation of CCR in transgenic poplar (Populus tremula 3 Populus alba) was associated with up to 50% reduced lignin content and an orange-brown, often patchy, coloration of the outer xylem. Thioacidolysis, nuclear magnetic resonance (NMR), immunocytochemistry of lignin epitopes, and oligolignol profiling indicated that lignin was relatively more reduced in syringyl than in guaiacyl units. The cohesion of the walls was affected, particularly at sites that are generally richer in syringyl units in wild-type poplar. Ferulic acid was incorporated into the lignin via ether bonds, as evidenced independently by thioacidolysis and by NMR. A synthetic lignin incorporating ferulic acid had a red-brown coloration, suggesting that the xylem coloration was due to the presence of ferulic acid during lignification. Elevated ferulic acid levels were also observed in the form of esters. Transcript and metabolite profiling were used as comprehensive phenotyping tools to investigate how CCR downregulation impacted metabolism and the biosynthesis of other cell wall polymers. Both methods suggested reduced biosynthesis and increased breakdown or remodeling of noncellulosic cell wall polymers, which was further supported by Fourier transform infrared spectroscopy and wet chemistry analysis. The reduced levels of lignin and hemicellulose were associated with an increased proportion of cellulose. Furthermore, the transcript and metabolite profiling data pointed toward a stress response induced by the altered cell wall structure. Finally, chemical pulping of wood derived from 5-year-old, field-grown transgenic lines revealed improved pulping characteristics, but growth was affected in all transgenic lines tested.

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-, or -did not inhibit these effects. Gö 6976 enhanced promoter activity, providing further evidence that PKC␣ was involved. Various CYP11B2 promoter constructs were used to identify the area associated with TPA and PKC inhibition. TPA and CA-PKC␣, -⑀, or -abolished the effects of AII, KCl, and Bt 2 cAMP on the activity of ؊102 and longer constructs. In summary, our findings suggest that the hamster CYP11B2 gene is under differential control by conventional (␣) and atypical () PKC.Aldosterone synthase is the product of the CYP11B2 gene and is responsible for the final step of aldosterone synthesis in most mammalian adrenal glands (1). This key regulatory enzyme is under the control of many factors including angiotensin II (AII), 1 adrenocorticotropin (ACTH), and changes in blood Na ϩ and K ϩ concentrations (1). The hamster adrenal CYP11B2 gene has been cloned and its promoter activity studied (2, 3). In transfected human adrenocortical carcinoma NCI-H295 cells, the first 161 base pairs of the 5Ј-flanking region of the hamster CYP11B2 were shown to be sufficient for maximal promoter activity and for the stimulatory effect of AII, KCl, and Bt 2 cAMP (4). In comparison, the first 129 base pairs of the 5Ј-flanking region of the human CYP11B2 gene were also shown to be responsive to the stimulatory action of AII, KCl and dibutyryl cyclic adenosine 3Ј:5Ј-monophosphate (Bt 2 cAMP) (5).The involvement of protein kinase C (PKC) in the expression of adrenal steroidogenic enzymes has been studied using activators and inhibitors of PKC. For example, 12-O-tetradecanoylphorbol-13-acetate (TPA) has been reported to increase the mRNA levels of 3-␤-hydroxysteroid dehydrogenase (6) and cytochrome P450 21-hydroxylase in the NCI-H295 cell line (7,8). In addition, phorbol esters have been shown to decrease the mRNA levels of cytochrome P450 17␣-hydroxylase (7) and cytochrome P450 side chain cleavage (7) and to abolish the enhancing effect of BAYK 8644 on cytochrome P450 11␤-hydroxylase (P450C11) and cytochrome P450 aldosterone synthase (P450aldo) (9). Steroidogenesis is also affected by inhibition of PKC. For instance, staurosporine has been reported to stimulate steroidogenesis in Y-1 cells (3, 10), to elevate the levels of mRNA of cytochrome P450 side chain cleavage and P450C11 (10), and to stimulate the activity of the hamster CYP11B2 promoter in transfected Y-1 cells (3). We have reported that GF109203X, a specific PKC inhibitor, has stimulated the output of aldosterone and the activity of the hamster CYP11B2 promoter in NCI-H295-transfected cells (4). In addition, staurosporine has been reported to reverse the inhibitory effect of TPA on aldosterone production in rat adrenal glomerulosa cells and to enhance the stimulatory effects of AII and K ϩ , but not of ACTH, in these cells (11).PKC comprises several isoforms of serine/threonine kinases that can be divided in three families (12)(13)(14). Members of the conventional family (PKC␣, -␤1, -␤2, and -␥) are calcium-dependent and are activated by phosphatidylserine and diacylglyce...