PMB 3005W Lecture Notes - 5 Feb 2024 PDF

PMB 3005W Feb 5, 2024 Reminders/Announcements: Do not share your data as a good drive link. All data must be uploaded on eLabJournal either as an attachment or image. All group members must participate and contribute to weekly assignments. Writing Assignment 1 is due next Monday by noon. Undergradua...

PMB 3005W Feb 5, 2024 Reminders/Announcements: Do not share your data as a good drive link. All data must be uploaded on eLabJournal either as an attachment or image. All group members must participate and contribute to weekly assignments. Writing Assignment 1 is due next Monday by noon. Undergraduate Research Opportunity: The Barney and Olszewski laboratories study the association between pennycress and the nitrogen-fixing microbe Gluconacetobacter diazotrophicus. We have developed improved strains of G. diazotrophicus that produces and releases more nitrogen. We are initiating a greenhouse study to determine the effectiveness of these strains in providing nitrogen to pennycress and the impact on yield. We will be hiring an Undergraduate Research Assistant to work between 10 and 20 hours per week on this project. The project will run for one year. If you are interested in this project contact: Neil Olszewski Email: [email protected] What We Learned Lab 2 – Stomatal Regulation Recap Stomatal regulation recap What effect did light have on stomatal aperture? Stomatal regulation recap Dark Light Stomatal regulation recap Overexpression of plasma membrane H+-ATPase in guard cells promotes light-induced stomatal opening and enhances plant growth Yin Wanga, Ko Noguchib, Natsuko Onoa, Shin-ichiro Inouea, Ichiro Terashimab, and Toshinori Kinoshitaa,c,1 a Division of Biological Science, Graduate School of Science, Nagoya University, Chikusa, Nagoya 464-8602, Japan; bDepartment of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; and cInstitute of Transformative Bio-Molecules (WPI-ITbM), Nagoya University, Chikusa, Nagoya 464-8602, Japan www.pnas.org/cgi/doi/10.1073/pnas.1305438111 Blue light Edited by Sarah M. Assmann, Penn State University, University Park, PA, and accepted by the Editorial Board November 18, 2013 (received for review March 21, 2013) Stomatal pores surrounded by a pair of guard cells in the plant epidermis control gas exchange between plants and the atmosphere in response to light, CO2, and the plant hormone abscisic acid. Light-induced stomatal opening is mediated by at least three key components: the blue light receptor phototropin (phot1 and phot2), plasma membrane H+-ATPase, and plasma membrane inward-rectifying K+ channels. Very few attempts have been made to enhance stomatal opening with the goal of increasing photosynthesis and plant growth, even though stomatal resistance is thought to be the major limiting factor for CO2 uptake by plants. Here, we show that transgenic Arabidopsis plants overexpressing H+-ATPase using the strong guard cell promoter GC1 showed enhanced light-induced stomatal opening, photosynthesis, and plant growth. The transgenic plants produced larger and increased numbers of rosette leaves, with ∼42–63% greater fresh and dry weights than the wild type in the first 25 d of growth. The dry weights of total flowering stems of 45-d-old transgenic plants, including seeds, siliques, and flowers, were ∼36–41% greater than those of the wild type. In addition, stomata in the transgenic plants closed normally in response to darkness and abscisic acid. In contrast, the overexpression of phototropin or inward-rectifying K+ channels in guard cells had no effect on these phenotypes. These results demonstrate that stomatal aperture is a limiting factor in photosynthesis and plant growth, and that manipulation of stomatal opening by overexpressing H+-ATPase in guard cells is useful for the promotion of plant growth. Light is one of the principal factors that stimulates stomatal opening, and various mechanisms underlie stomatal opening in response to different light wavelengths (6–8). Red light is thought to induce stomatal opening via photosynthesis in the mesophyll and guard cell chloroplasts, as well as the reduction of the intercellular CO2 concentration (Ci) (5, 9, 10). However, the detailed mechanisms of stomatal responses to red light are under debate (11, 12). In contrast, blue light acts as a signal and exerts the most pronounced effect on stomatal opening. The blue light receptors phototropins (phot1 and phot2) activate plasma membrane H+ -ATPase through the phosphorylation of the penultimate threonine and subsequent binding of the 14-3-3 protein to the phosphorylated threonine (13–15). Blue lightactivated H+ -ATPase induces hyperpolarization of the plasma membrane, which allows K+ uptake through inward-rectifying K+ (K+ in) channels (16–21). Accumulation of K+ induces the swelling of guard cells and stomatal opening. Thus, these three components (phototropins, plasma membrane H+-ATPase, and K+in channels) have important roles in blue light-induced stomatal opening. In addition to these components, FLOWERING LOCUS T (FT) is suggested to be a positive regulator for stomatal opening via its effect on the activation status of the plasma membrane H+-ATPase (22). Required for stomatal opening: Blue light receptor H+ ATPAse K+ uptake channel Hypothesis: Enhancing stomatal opening will increase photosynthesis and plant growth. Arabidopsis thaliana biomass https://doi.org/10.3389/fpls.2013.00138 I | stomatal conductance | photosynthetic rate | n the present era of global climate changes and the threat of food insufficiency, finding ways to improve the uptake of CO2 Significance Stomata are microscopic pores surrounded by two guard cells and play an important role in the uptake of CO2 for photosynthesis. Recent researches revealed that light-induced sto- F Stomatal regulation recap 4 WT GC1::AHA2 #1 GC1::AHA2 #2 A *** *** B GC1::AHA2 #1 WT GC1::AHA2 # 3 2 ******** 1 0 F C Dark Light 80 WT G Light + ABA Figure 1 – Overexpressing an 120 H+-ATPase promotes stomatal 100 opening. f initial weight) WT D Scale bar: 2 GC1::AHA2 #1 #1 WT GC1::AHA2 Figure 3 – Plant phenotypes of overexpression 2.0 lines compared to wild type – 40-60% more * * 1.5 fresh weight 6 ive stem weight 1 5 ) Stomatal aperture (µm) C Scale bar: 2 1.0 E www.pnas.org/cgi/doi/10.1073/pnas.1305438111 2.5 Fresh weight Stomatal regulation recap What effect did ABA have on stomatal aperture? Stomatal aperture recap ABA Dark Light Basal ABA Basal Plant Hormones https://www.pathwayz.org/Tree/Plain/PLANT+HORMONES Small molecules made in plants that act as signals to evoke some response in the plant. Act at very low concentrations - micromolar ABA are essential for maintaining plant growth and development. Indeed, ABA affects leaf and Stomatal aperture recap Role of ABA in plant growth Abscisic acid (ABA) is a stress responsive hormone that accumulates under drought conditions. One of its effects is the closing of stomata to reduce water loss. ABA is synthesized in the root and leaf vasculature and moves to the site of action. https://doi.org/10.1016/j.tplants.2019.04.008 General role in control development. In roots, ABA is involved in processes stimulating water uptake, including hydrotropism and xylem of water status. Trends in Plant Science Figure 3. Physiological Roles of Basal ABA. Basal abscisic acid (ABA) is involved in water homeostasis, growth, and formation. Deposition of hydrophobic suberin layers and primary root growth are promoted by ABA. In the shoot, Stomatal aperture recap Hypothesis: Potassium transporters are required for the ABA response (closure of stomatal apertures). Method: Use mutants lacking various transporters in experiments with and w/out ABA Null Hypothesis: Mutants and wild-type plants will show the same stomatal responses Stomatal aperture recap Experimental Design: Similar procedure to Stomatal Regulation lab but with a few key exceptions - Epidermal peels underwent stomatal pre-opening in the light using a 2h incubation in 10mM KCl, 0.2mM CaCl2, 10mM MES-KOH pH 6.15. - Peeled strips were treated with or without ABA for 1h. - 50 stomatal apertures were photographed. - Experiments were repeated three times Plant Cellaperture recap Stomatal Asterisks = statistically significant difference b/w wild-type (Co) and mutants using a Student’s t-test: *P < 0.001 **P