Question: What and exactly where are plasmodesmata?

Answer: Plasmodesmata are pores that form connections between two adjoining plant cells. They are lined by plasma membranes and allow for cytoplasmic continuity between cells. They are the pathways through which sugar moves from cell to cell in the symplastic pathway of sugar transport.

Question: What is in situ hybridization and how it is carried out in the case of expansin and SCR ? Are we looking for the presence of a gene expression using this technique?

Answer: You are correct: in situ hybridization is a technique in which labeled probes specific for a given messenger RNA hybridize to that RNA in tissue sections. The position of the hybridization can then be easily detected and that position reveals which cells are most active in expressing that RNA.

Question: What is CAB stand for (in senescence, comparing mRNA of CAB to SARK) and what does it do?

Answer: CAB is chlorophyll a/b protein. The loss of this protein is an early indicator of the onset of senescence.

Question: In the Zn tolerance handout what is the significance of the Mg-GTP treatment? Also, why is Mg needed along with ATP in the function of the proton pump?

Answer: The authors are trying to see whether the zinc transporter in the tolerant ecotype is different from the one in the sensitive ecotype. Testing whether the transporter requires SPECIFICALLY ATP or can use GTP equally well is just one way to discriminate the two transporters. When used by active transporters, ATP almost always has to be complexed with Mg; it doesn't work without Mg.

Question: Is this a true statement? - Rubisco switches to oxygenase activity when the oxygen concentration increases due the closing of the stoma in a plant living in a hot, sunny environment. The high oxygen concentration is caused by oxygen let off during photosynthesis and its inability to escape through the stomatal openings.

Answer: The rise in oxygen concentration under the conditions you describe certainly plays a role in switching rubisco to an oxygenase mode, but at least equally important is the depletion of CO2 in the same spaces as carbon fixation procedes.

Question: Which direction does the proton pump pump H+ ions in the sucrose symport in apoplastic loading? Phloem? Source cell?

Answer: The proton pump pumps protons from the cytoplasm of the cell that will be loading sucrose into the apoplasm (wall space) of that cell. Then, when sugar "loads" into that cell, protons enter along with the sugar in a symport process.

Question: I'm not understanding the purpose of a low affinity high capacity sucrose transport protein used in minor veins of the vascular system. I think I may not fully understand the terms affinity and capacity with respect to an protein's function.

Answer: If a transporter has a low affinity for its cargo, that means it will bind to it only at relatively high concentrations of the cargo, and its capacity for binding would not saturate until relatively high concentrations of the cargo were reached. High affinity transporters, in contrast, saturate at much lower concentrations of their cargo, and so they have a lower transport capacity. The new sucrose transporter described in the August 2000 issue of Plant Cell has a low affinity and high capacity for its cargo, sucrose, and it is expressed predominately in zones with high rates of phloem loading. It appears that plants optimize the capacity and affinity of sucrose uptake into the sieve elements in different regions of leaves, depending on the rates of phloem loading needed and the level of sugar available for loading.

Question: In regard to the Figure relating pressure to polar ratio in Chara cells. I understand what's happening with the vertical cells, but am a little confused about the horizontal cells.
Answer: If no pressure is added, cells in the horizontal position should have a polar ratio of 1.0 [rate of flow to the left = rate of flow to the right]. If positive pressure is added, the polar ratio goes to 1.1, that is flow in one direction is 10% faster than flow in the other direction), but this change can be blocked by nifedipine (calcium-channel blocker); if negative pressure is added, ratio goes to near 0.9, and this change is also blocked by nifedipine. Thus, whether tension and compression forces are generated by gravity (when cells are in vertical orientation) or by pressure (when cells are in horizontal orientation), the polar ratio becomes 10% faster in one direction than in the other, and this change can be blocked by calcium channel blockers.

Question: In the integrin experiments, what exactly is RGDS? I know integrins bind to the RGD sequence. Is RGDS just an integrin like protein that competes for that RGD sequence, thus inhibiting the attachment of integrins?
Answer: RGDS is a peptide [Arg-Gly-Glu-Ser] containing the RGD sequence present at the site on ECM proteins where integrins bind. So, you are right, it competes for the RGD binding site, thus inhibiting the attachment of integrins to proteins in the ECM.

Question: Regarding the article on "A Role for Ectophosphatase in Xenobiotic Resistance," I don't understand the purpose of the first set of illustrations shown under E. Better yet, what is this figure showing?.

Answer: The illustration shows two rows of petri dishes with tiny Arabidopsis seedlings growing in them. The first row (MDR1 OE) shows transgenic plants that are overexpressing the MDR transporter gene. The second row (Wt) shows wild type plants (not transformed). Beneath the two rows of petri dishes, there are numbers comparing the % germination of seeds in the two rows, with the numerator denoting the % MDR plants that germinated, and the denominator denoting the % Wt plants that germinated.

There are 4 columns showing different treatments: Col. 1 is No Treatment, Col. 2 is treatment with the poison cycloheximide, Col. 3 is treatment with an inhibitor of extracellular ATPase activity (which would inhibit apyrase), and Col 4 is treatment with a combination of Cycloheximide and ATPase inhibitor.

What the results show is that:

(1) Plants overexpressing MDR are more resistant to cycloheximide than Wt plants (50% germination vs. 2 % germination)

(2) The ATPase inhibitor by itself is not very toxic (83% germination & 90% germination)

(3) Inhibiting the extracellular ATPase completely nullifies the resistance of the MDR plants to cycloheximide (0% germination).

Question: Regarding the November 2000 article by Bauly et al. on the topic of ABP1, what is this article trying to prove? I know that the experiment involved genetically modifying ABP1 so that it did not have its normal KDEL sequence, which is a signal that sends the ABP1 to the ER. The results were that the responsiveness of cells to auxin did not change whether the KDEL signal was altered or not. But what is this proving? In addition, if KDEL is removed, shouldn't the cell end up more localized somewhere else in the cell, such as the plasma membrane? Was this shown or not?

Answer: You are correct, the key result of the November 2000 article is that when plants were were engineered to overexpress ABP-1, their auxin sensitivity was enhanced equally whether they were transformed with a wild-type form of ABP-1 (which has a KDEL targeting signal that confines it mainly to the ER) or with a mutated form of ABP-1 in which the KDEL signal was altered so that ABP-1 was NOT confined to the ER. Although the mutated form of ABP-1 was not confined to the ER, plants expressing this mutated form did not have more ABP-1 on the plasma membrane than plants expressing the wild-type form of ABP-1.

The key conclusions are that:

1) Overexpressing ABP-1 confers hypersensitivity to auxin whether it is confined mostly to the ER or not and whether more of it is located on the plasma membrane or not, and

2) Engineering ABP-1 so that it is no longer confined mainly to the ER does not make more of it go to the plasma membrane, nor does it make ABP-1 more active as a receptor (i.e., make the plants more responsive to auxin).

The results and conclusions of this paper are important because they appear inconsistent with these two widely-held hypotheses about ABP-1:

1. Redirecting ABP-1 out of the ER (by altering its KDEL address) should increase, or at least alter, its receptor activity.

2. Increasing the receptor activity of ABP-1 should require that more ABP-1 be directed to the plasma membrane.

According to the rules of Strong Inference, these findings advance the auxin field by rendering certain widely-help hypotheses unlikely and thus forcing the field to devise alternate explanations for relating the subcellular locale of ABP-1 to its ability to confer auxin hypersensitivity to plants.

Question: In reference to the Ecto-Apyrase paper--are the apyrases functioning as ATPases?

Ans.: Yes; apyrases hydrolyze ATP to ADP.

Question.: What is "xenobiotic" defined as?

Ans.: A biologically active compound that is "foreign" to the cell. Often xenobiotics are poisons.

Question: Are the wasps homing in on the scent from the caterpillar's saliva, or is the saliva inducing the plant to release their attractants? Also, are these attractants congruent to human pheromones?

Ans.: Volicitin in the saliva of the feeding army worm induces the plant to release the attractants. The attractants resemble insect pheromones in that they are volatile attractants. Normally, however, insect pheromones function to attract mates for reproduction.

Question: If the saliva does induce the plant to release attractants, why does it remain localized instead of flowing through the plant like the proteinases do?

Ans.: It is likely that the feeding injury imposed by the army worm does induce the hormone systemin (= PIIF, proteinase inhibitor inducing factor), which would flow through the plant to induce proteinase inhibitors. The volicitin-induced response is over and above the systemin response. [Note: The proteinases do not flow through the plant; what flows through the plant is systemin, the hormone that induces proteinase inhibitors].

Question: You had said that it is environmentally better if you contain the pesiticide in the plant through genetic modification instead of spraying pesticide over an entire field. But I was thinking about it later and I don't understand how this would be better for the environment. If you spray pesticide over a crop, then some of the pesticide will drain into our water supply, right? But if the plant is always making the pesticide, then there will be a constant flow of pesticide into our water. So that is clearly more harmful for us. What do scientists say about this situation?

Answer: The Bt toxin that is engineered to be produced by the plant is a protein made in (some of) the cells of the plant. So none of it actually flows out. The insect encounters this poison only when it eats plant tissue. Presumably when the leaves of the plant die, the Bt toxin peptides are broken down to their component amino acids, which are non-toxic, and recycled to make other proteins.

Question: Are phyA-GFP and phyB-GFP fused complexes that act like the photoreceptors in native plants? And are phyA-GFP and phyB-GFP localized in the cytosol of all dark-adapted plants, meaning that when the plant is kept in the dark, then phyA-GFP and phyB-GFP are found in the cytosol? And, when introduced to red light, both phyA-GFP and phyB-GFP move to the nucleus?

Answer: Yes

Question: The movement of PhyB-GFP to the nucleus is inhibited in far red light, but the movement of phyA-GFP to the nucleus is promoted by far-red light. Does this prove that nuclear import of phyA-GFP is controlled by a very low fluence response, whereas translocation of phyB-GFP is regulated by a low fluence response of phytochrome?

Ans: Exactly right, the irradiation results testing the effects of red and far-red light prove that the movement of phyA-GFP to the nucleus is a very low fluence response, whereas the movement of phyB-GFP to the nucleus is a low-fluence response.

Question: I don't understand the difference between very low and low fluence responses.

Answer: By definition "Very Low" fluence phytochrome responses are triggered by irradiations that establish only VERY low levels of Pfr. Because of the overlap in the absorbance spectra of Pr and Pfr, even far-red light can shift the equilibrium between Pr and Pfr from 100% Pr (in darkness) to levels of Pfr that are at least 0.1% [= VERY low levels]. Levels of Pfr that are as low as 0.1% can promote very low fluence responses, and because these responses cannot be reversed by far-red light, they are not photoreversible.

In contrast, "Low Fluence" responses require significantly higher levels of Pfr, and these levels cannot be obtained using far-red light. Thus "low-fluence" responses ARE photoreversible; i.e., the inducing effects of red light can be reversed by far-red light.

The movement of phyA-GFP to the nucleus is a very-low fluence response, because it can be induced by far-red light. The movement of phyB-GFP is a low-fluence response because far-red light can reverse the ability of red light to induce the movement of phyB-GFP to the nucleus.