Quantification of Cell-to-Cell Connectivity Using Particle Bombardment

Tee, Estee E-Ling; Samwald, Sebastian; Faulkner, Christine

doi:10.1007/978-1-0716-2132-5_17

Cited by 6 publications

(4 citation statements)

References 14 publications

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“…A SIG5::LUCIFERASE reporter construct (pGREENII0229 SIG5:: LUCIFERASE 24 ) was expressed transiently in Arabidopsis seedlings using methods similar to those used previously 35,36 . Briefly, 5 µg of pGREENII0229 SIG5::LUCIFERASE or pB7WG2.0-GFP (green fluorescent protein (GFP) positive control for transformation 100 ) was combined with 25 µl of 1 nm gold particle suspension (Bio-Rad), 25 µl of 2.5 M CaCl 2 and 10 µl of 0.1 M spermidine (Sigma), and incubated on ice for 30 min. DNA-coated gold particles were washed with 100% ethanol, resuspended in 100% ethanol and stored at −20 °C before particle bombardment.…”

Section: Transient Luciferase Expression By Particle Bombardmentmentioning

confidence: 99%

Low-temperature and circadian signals are integrated by the sigma factor SIG5

et al. 2023

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Chloroplasts are a common feature of plant cells and aspects of their metabolism, including photosynthesis, are influenced by low-temperature conditions. Chloroplasts contain a small circular genome that encodes essential components of the photosynthetic apparatus and chloroplast transcription/translation machinery. Here, we show that in Arabidopsis, a nuclear-encoded sigma factor that controls chloroplast transcription (SIGMA FACTOR5) contributes to adaptation to low-temperature conditions. This process involves the regulation of SIGMA FACTOR5 expression in response to cold by the bZIP transcription factors ELONGATED HYPOCOTYL5 and ELONGATED HYPOCOTYL5 HOMOLOG. The response of this pathway to cold is gated by the circadian clock, and it enhances photosynthetic efficiency during long-term cold and freezing exposure. We identify a process that integrates low-temperature and circadian signals, and modulates the response of chloroplasts to low-temperature conditions.

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Section: Transient Luciferase Expression By Particle Bombardmentmentioning

confidence: 99%

Low-temperature and circadian signals are integrated by the sigma factor SIG5

et al. 2023

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“…Bombardment assays were performed as described in ref. 45 . Bombardment sites were imaged 16 h after bombardment using a confocal microscope (LSM Zeiss 800) with a 20× water dipping objective (W N-Achroplan 20×/0.5).…”

Section: Methodsmentioning

confidence: 99%

A PDLP-NHL3 complex integrates plasmodesmal immune signaling cascades

Tee

Johnston

Papp

et al. 2023

Proc. Natl. Acad. Sci. U.S.A.

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The plant immune system relies on the perception of molecules that signal the presence of a microbe threat. This triggers signal transduction that mediates a range of cellular responses via a collection of molecular machinery including receptors, small molecules, and enzymes. One response to pathogen perception is the restriction of cell-to-cell communication by plasmodesmal closure. We previously found that while chitin and flg22 trigger specialized immune signaling cascades in the plasmodesmal plasma membrane, both execute plasmodesmal closure via callose synthesis at the plasmodesmata. Therefore, the signaling pathways ultimately converge at or upstream of callose synthesis. To establish the hierarchy of signaling at plasmodesmata and characterize points of convergence in microbe elicitor-triggered signaling, we profiled the dependence of plasmodesmal responses triggered by different elicitors on a range of plasmodesmal signaling machinery. We identified that, like chitin, flg22 signals via RESPIRATORY BURST OXIDASE HOMOLOGUE D (RBOHD) to induce plasmodesmal closure. Further, we found that PLASMODESMATA-LOCATED PROTEIN 1 (PDLP1), PDLP5, and CALLOSE SYNTHASE 1 (CALS1) are common to microbe- and salicylic acid (SA)-triggered responses, identifying PDLPs as a candidate signaling nexus. To understand how PDLPs relay a signal to CALS1, we screened for PDLP5 interactors and found NON-RACE SPECIFIC DISEASE RESISTANCE/HIN1 HAIRPIN-INDUCED-LIKE protein 3 (NHL3), which is also required for chitin-, flg22- and SA-triggered plasmodesmal responses and PDLP-mediated activation of callose synthesis. We conclude that a PDLP–NHL3 complex acts as an integrating node of plasmodesmal signaling cascades, transmitting multiple immune signals to activate CALS1 and plasmodesmata closure.

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“…Bombardment assays of cytosolic GFP endoplasmic reticulum RFP (red fluorescent protein) were performed as described in Tee et al (2022). Bombardment sites were imaged 16 hours after bombardment using a confocal microscope (LSM Zeiss 800) with a 20× water dipping objective (W N-Achroplan 20×/0.5).…”

Section: Microprojectile Bombardment Assaysmentioning

confidence: 99%

Cell-specialized chloroplast signaling orchestrates photosynthetic and extracellular reactive oxygen species for stress responses

Tee

Fairweather

et al. 2023

Preprint

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Cellular responses to abiotic stress involve multiple secondary messengers including reactive oxygen species (ROS), Ca2+, phytohormones such as abscisic acid (ABA) and chloroplast-to-nucleus retrograde signals such as 3’-phosphoadenosine 5’-phosphate (PAP). Mechanism(s) by which these messengers, produced in different subcellular compartments, intersect for cell regulation remain enigmatic. Here we demonstrate a mechanistic link enabling ABA and the chloroplast retrograde signal PAP to coordinate both chloroplast and plasma membrane ROS production. In whole leaves, PAP alters various ROS-related processes including plasmodesmal permeability as well as responses to ozone and the bacterial elicitor flg22, but largely quenches ROS during oxidative stress. Conversely, in guard cells, both PAP and ABA induce a ROS burst in both chloroplasts via photosynthetic electron transport, and the apoplast via the RESPIRATORY BURST OXIDASE HOMOLOG (RBOH). Both subcellular ROS sources were necessary for ABA- and PAP-mediated stomatal closure. However, PAP signaling diverges from ABA by activating RBOHD, instead of RBOHF, for apoplastic ROS production. We identify calcium-dependent protein kinases (CPKs) transcriptionally induced by PAP as the post-translational activators of RBOHD-mediated ROS production. CPK13, CPK32, and CPK34 concurrently activate RBOHD and the slow anion channel SLAC1 by phosphorylating two Serine (S) residues, including S120 which is also targeted by the core ABA signaling kinase OPEN STOMATA 1 (OST1). Consequently, overexpressing the PAP-induced CPKs rescues stomatal closure inost1.Our data identify chloroplasts, as sources and mediators of ROS and retrograde signals such PAP, to be critical environmental sensors and focal node in the multifaceted cellular stress response network.Significance StatementThe chloroplast is an important node to coordinate multiple plant signaling pathways in response to stresses such as drought. However, how does it function in specialized cells for which carbon fixation is secondary? Here we show that the chloroplast retrograde signal 3’-phosphoadenosine 5’-phosphate (PAP) plays multiple roles in reactive oxygen species (ROS) signaling and homeostasis. While PAP suppresses ROS in photosynthetic tissue, surprisingly PAP induces ROS in chloroplasts and extracellular space of guard cells to induce stomatal closure. We decipher how PAP-induced proteins activate both extracellular ROS production and anion channels for stomatal closure, thus providing a mechanism by which chloroplasts provide a strategic complement to canonical hormonal pathways in regulating plant physiological responses in specialized cells.

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Quantification of Cell-to-Cell Connectivity Using Particle Bombardment

Cited by 6 publications

References 14 publications

Low-temperature and circadian signals are integrated by the sigma factor SIG5

Low-temperature and circadian signals are integrated by the sigma factor SIG5

A PDLP-NHL3 complex integrates plasmodesmal immune signaling cascades

Cell-specialized chloroplast signaling orchestrates photosynthetic and extracellular reactive oxygen species for stress responses

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