Different classes of plant hormones and different wavelengths of light act through specific signal transduction mechanisms to coordinate higher plant development. A specific prephenate dehydratase protein (PD1) was discovered to have a strong interaction with the sole canonical G-protein Gα-subunit (GPA1) in Arabidopsis (Arabidopsis thaliana). PD1 is a protein located in the cytosol, present in etiolated seedlings, with a specific role in blue light-mediated synthesis of phenylpyruvate and subsequently of phenylalanine (Phe). Insertion mutagenesis confirms that GPA1 and the sole canonical G-protein-coupled receptor (GCR1) in Arabidopsis also have a role in this blue light-mediated event. In vitro analyses indicate that the increase in PD1 activity is the direct and specific consequence of its interaction with activated GPA1. Because of their shared role in the light-mediated synthesis of phenylpyruvate and Phe, because they are iteratively interactive, and because activated GPA1 is directly responsible for the activation of PD1; GCR1, GPA1, and PD1 form all of or part of a signal transduction mechanism responsible for the light-mediated synthesis of phenylpyruvate, Phe, and those metabolites that derive from that Phe. Data are also presented to confirm that abscisic acid can act through the same pathway. An additional outcome of the work is the confirmation that phenylpyruvate acts as the intermediate in the synthesis of Phe in etiolated plants, as it commonly does in bacteria and fungi.

In the mammalian heart, the extracellular matrix plays an important role in regulating cell behavior and adaptation to mechanical stress. In cell culture, a significant number of cells detach in response to mechanical stimulation, limiting the scope of such studies. We describe a method to adhere the synthetic peptides RGD (fibronectin) and YIGSR (laminin) onto silicone for culturing primary cardiac cells and studying responses to mechanical stimulation. We first examined cardiac cells on stationary surfaces and observed the same degree of cellular adhesion to the synthetic peptides as their respective native proteins. However, the number of striated myocytes on the peptide surfaces was significantly reduced. Focal adhesion kinase (FAK) protein was reduced by 50% in cardiac cells cultured on YIGSR peptide compared with laminin, even though beta(1)-integrin was unchanged. Connexin43 phosphorylation increased in cells adhered to RGD and YIGSR peptides. We then subjected the cardiac cells to cyclic strain at 20% maximum strain (1 Hz) for 48 h. After this period, cell attachment on laminin was reduced to approximately 50% compared with the unstretched condition. However, in cells cultured on the synthetic peptides, there was no significant difference in cell adherence after stretch. On YIGSR peptide, myosin protein was decreased by 50% after mechanical stimulation. However, total myosin was unchanged in cells stretched on laminin. These results suggest that RGD and YIGSR peptides promote the same degree of cellular adhesion as their native proteins; however, they are unable to promote the signaling required for normal FAK expression and complete sarcomere formation in cardiac myocytes.

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Various surface modifications have been applied to titanium alloy (Ti-6Al-4V) implants, in an attempt to enhance osseointegration; crucial for ideal prosthetic fixation. Despite the numerous studies demonstrating that peptide-modified surfaces influence in vitro cellular behavior, there is relatively little data reporting their effects on bone remodeling. The objective of this article was to examine the effects of chemically modifying Ti-6Al-4V surfaces with a common RGD sequence, a 15-residue peptide containing GRGDSP (glycine-arginine-glycine-aspartate-serine-proline), on the modulation of bone remodeling. The expression of proteins known to be associated with osseous matrix and bone resorption were studied during the growth of human bone-derived cells (HBDC) on these peptide-modified surfaces. HBDC grown for 7 days on RGD surfaces displayed significantly increased levels of osteocalcin, and pro-collagen Ialpha1 mRNAs, compared with the production by HBDC grown on the native Ti-6Al-4V. A pattern that was also reflected at the protein levels for osteocalcin, type I collagen, and bone sialoprotein. Moreover, HBDC grown for 7 and 14 days on RGD-modified Ti-6Al-4V expressed significantly higher level of osteoclast differentiation factors and lower levels of osteoprotegerin and IL-6 proteins compared with other surfaces tested. These results suggest that different chemical treatments of implant material (Ti-6Al-4V) surface result in differential bone responses, not only their ability to form bone but also to stimulate osteoclastic formation.

Chemical derivatization of peptides allows efficient F2 laser single photon ionization (SPI) of Fmoc-derivatized peptides covalently bound to surfaces. Laser desorption photoionization mass spectrometry using 337-nm pulses for desorption and 157.6-nm pulses for threshold SPI forms large ions identified as common peptide fragments bound to either Fmoc or the surface linker. Electronic structure calculations indicate the Fmoc label is behaving as an ionization tag for the entire peptide, lowering the ionization potential of the complex below the 7.87-eV photon energy. This method should allow detection of many molecular species covalently or electrostatically bound to surfaces.