@misc{wyatt_tsou_robertson_boss_2004, title={Transgenic plants with increased calcium stores}, volume={6,753,462}, number={2004 June 22}, publisher={Washington, DC: U.S. Patent and Trademark Office}, author={Wyatt, S. and Tsou, P.-L. and Robertson, D. and Boss, W.}, year={2004} } @article{wyatt_tsou_robertson_2002, title={Expression of the high capacity calcium-binding domain of calreticulin increases bioavailable calcium stores in plants}, volume={11}, ISSN={["1573-9368"]}, DOI={10.1023/A:1013917701701}, abstractNote={Modulation of cytosolic calcium levels in both plants and animals is achieved by a system of Ca2+-transport and storage pathways that include Ca2+ buffering proteins in the lumen of intracellular compartments. To date, most research has focused on the role of transporters in regulating cytosolic calcium. We used a reverse genetics approach to modulate calcium stores in the lumen of the endoplasmic reticulum. Our goals were two-fold: to use the low affinity, high capacity Ca2+ binding characteristics of the C-domain of calreticulin to selectively increase Ca2+ storage in the endoplasmic reticulum, and to determine if those alterations affected plant physiological responses to stress. The C-domain of calreticulin is a highly acidic region that binds 20-50 moles of Ca2+ per mole of protein and has been shown to be the major site of Ca2+ storage within the endoplasmic reticulum of plant cells. A 377-bp fragment encoding the C-domain and ER retention signal from the maize calreticulin gene was fused to a gene for the green fluorescent protein and expressed in Arabidopsis under the control of a heat shock promoter. Following induction on normal medium, the C-domain transformants showed delayed loss of chlorophyll after transfer to calcium depleted medium when compared to seedlings transformed with green fluorescent protein alone. Total calcium measurements showed a 9-35% increase for induced C-domain transformants compared to controls. The data suggest that ectopic expression of the calreticulin C-domain increases Ca2+ stores, and that this Ca2+ reserve can be used by the plant in times of stress.}, number={1}, journal={TRANSGENIC RESEARCH}, author={Wyatt, SE and Tsou, PL and Robertson, D}, year={2002}, month={Feb}, pages={1–10} } @article{wyatt_rashotte_shipp_robertson_muday_2002, title={Mutations in the gravity persistence signal loci in arabidopsis disrupt the perception and/or signal transduction of gravitropic stimuli}, volume={130}, ISSN={["1532-2548"]}, DOI={10.1104/pp.102.010579}, abstractNote={Abstract Gravity plays a fundamental role in plant growth and development, yet little is understood about the early events of gravitropism. To identify genes affected in the signal perception and/or transduction phase of the gravity response, a mutant screen was devised using cold treatment to delay the gravity response of inflorescence stems of Arabidopsis. Inflorescence stems of Arabidopsis show no response to gravistimulation at 4°C for up to 3 h. However, when gravistimulated at 4°C and then returned to vertical at room temperature (RT), stems bend in response to the previous, horizontal gravistimulation (H. Fukaki, H. Fujisawa, M. Tasaka [1996] Plant Physiology 110: 933–943). This indicates that gravity perception, but not the gravitropic response, occurs at 4°C. Recessive mutations were identified at three loci using this cold effect on gravitropism to screen for gravity persistence signal (gps) mutants. All three mutants had an altered response after gravistimulation at 4°C, yet had phenotypically normal responses to stimulations at RT. gps1-1 did not bend in response to the 4°C gravity stimulus upon return to RT.gps2-1 responded to the 4°C stimulus but bent in the opposite direction. gps3-1 over-responded after return to RT, continuing to bend to an angle greater than wild-type plants. At 4°C, starch-containing statoliths sedimented normally in both wild-type and the gps mutants, but auxin transport was abolished at 4°C. These results are consistent with GPS loci affecting an aspect of the gravity signal perception/transduction pathway that occurs after statolith sedimentation, but before auxin transport.}, number={3}, journal={PLANT PHYSIOLOGY}, author={Wyatt, SE and Rashotte, AM and Shipp, MJ and Robertson, D and Muday, GK}, year={2002}, month={Nov}, pages={1426–1435} } @article{persson_wyatt_love_thompson_robertson_boss_2001, title={The Ca2+ status of the endoplasmic reticulum is altered by induction of calreticulin expression in transgenic plants}, volume={126}, ISSN={["1532-2548"]}, DOI={10.1104/pp.126.3.1092}, abstractNote={Abstract To investigate the endoplasmic reticulum (ER) Ca2+ stores in plant cells, we generated tobacco (Nicotiana tabacum; NT1) suspension cells and Arabidopsis plants with altered levels of calreticulin (CRT), an ER-localized Ca2+-binding protein. NT1 cells and Arabidopsis plants were transformed with a maize (Zea mays) CRT gene in both sense and antisense orientations under the control of an Arabidopsis heat shock promoter. ER-enriched membrane fractions from NT1 cells were used to examine how altered expression of CRT affects Ca2+uptake and release. We found that a 2.5-fold increase in CRT led to a 2-fold increase in ATP-dependent 45Ca2+accumulation in the ER-enriched fraction compared with heat-shocked wild-type controls. Furthermore, after treatment with the Ca2+ ionophore ionomycin, ER microsomes from NT1 cells overproducing CRT showed a 2-fold increase in the amount of45Ca2+ released, and a 2- to 3-fold increase in the amount of 45Ca2+ retained compared with wild type. These data indicate that altering the production of CRT affects the ER Ca2+ pool. In addition, CRTtransgenic Arabidopsis plants were used to determine if altered CRT levels had any physiological effects. We found that the level of CRT in heat shock-induced CRT transgenic plants correlated positively with the retention of chlorophyll when the plants were transferred from Ca2+-containing medium to Ca2+-depleted medium. Together these data are consistent with the hypothesis that increasing CRT in the ER increases the ER Ca2+ stores and thereby enhances the survival of plants grown in low Ca2+ medium.}, number={3}, journal={PLANT PHYSIOLOGY}, author={Persson, S and Wyatt, SE and Love, J and Thompson, WF and Robertson, D and Boss, WF}, year={2001}, month={Jul}, pages={1092–1104} } @article{collings_carter_rink_scott_wyatt_allen_2000, title={Plant nuclei can contain extensive grooves and invaginations}, volume={12}, DOI={10.2307/3871239}, number={12}, journal={Plant Cell}, author={Collings, D. A. and Carter, C. N. and Rink, J. C. and Scott, A. C. and Wyatt, S. E. and Allen, N. S.}, year={2000}, pages={2425–2439} } @article{scott_wyatt_tsou_robertson_allen_1999, title={Model system for plant cell biology: GFP imaging in living onion epidermal cells}, volume={26}, ISSN={["1940-9818"]}, DOI={10.2144/99266st04}, abstractNote={ The ability to visualize organelle localization and dynamics is very useful in studying cellular physiological events. Until recently, this has been accomplished using a variety of staining methods. However, staining can give inaccurate information due to nonspecific staining, diffusion of the stain or through toxic effects. The ability to target green fluorescent protein (GFP) to various organelles allows for specific labeling of organelles in vivo. The disadvantages of GFP thus far have been the time and money involved in developing stable transformants or maintaining cell cultures for transient expression. In this paper, we present a rapid transient expression system using onion epidermal peels. We have localized GFP to various cellular compartments (including the cell wall) to illustrate the utility of this method and to visualize dynamics of these compartments. The onion epidermis has large, living, transparent cells in a monolayer, making them ideal for visualizing GFP. This method is easy and inexpensive, and it allows for testing of new GFP fusion proteins in a living tissue to determine deleterious effects and the ability to express before stable transformants are attempted. }, number={6}, journal={BIOTECHNIQUES}, author={Scott, A and Wyatt, S and Tsou, PL and Robertson, D and Allen, NS}, year={1999}, month={Jun}, pages={1125-+} } @article{collings_winter_wyatt_allen_1998, title={Growth dynamics and cytoskeleton organization during stem maturation and gravity-induced stem bending in Zea mays L.}, volume={207}, ISSN={["1432-2048"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-0032435367&partnerID=MN8TOARS}, DOI={10.1007/s004250050480}, abstractNote={Characterization of gravitropic bending in the maize stem pulvinus, a tissue that functions specifically in gravity responses, demonstrates that the pulvinus is an ideal system for studying gravitropism. Gravistimulation during the second of three developmental phases of the pulvinus induces a gradient of cell elongation across the non-growing cells of the pulvinus, with the most elongation occurring on the lower side. This cell elongation is spatially and temporally separated from normal internodal cell elongation. The three characterized growth phases in the pulvinus correspond closely to a specialized developmental sequence in which structural features typical of cells not fully matured are retained while cell maturation occurs in surrounding internodal and nodal tissue. For example, the lignification of supporting tissue and rearrangement of transverse microtubules to oblique that occur in the internode when cell elongation ceases are delayed for up to 10 d in the adjacent cells of the pulvinus, and only occurs as a pulvinus loses its capacity to respond to gravistimulation. Gravistimulation does not modify this developmental sequence. Neither wall lignification nor rearrangement of transverse microtubules occurs in the rapidly elongating lower side or non-responsive upper side of the pulvinus until the pulvinus loses the capacity to bend further. Gravistimulation does, however, lead to the formation of putative pit fields within the expanding cells of the pulvinus.}, number={2}, journal={PLANTA}, author={Collings, DA and Winter, H and Wyatt, SE and Allen, NS}, year={1998}, month={Dec}, pages={246–258} }