High Aspect Ratio Polymer Nanocarriers for Gene Delivery and Expression in Plants.

Plant genetic engineering methods are critical for food security and biofuel production and to enable molecular farming. Here, we elucidated how polymeric high aspect ratio nanocarriers can enable DNA delivery to plants and transient expression. We demonstrated that a nanocarrier with 20 nm width, 80 nm length, and a polymer-to-DNA ratio of N/P = 3.

ABA-activated low-nanomolar Ca-CPK signalling controls root cap cycle plasticity and stress adaptation.

Abscisic acid (ABA) regulates plant stress adaptation, growth and reproduction. Despite extensive ABA-Ca signalling links, imaging ABA-induced increases in Ca concentration has been challenging, except in guard cells. Here we visualize ABA-triggered [Ca] dynamics in diverse organs and cell types of Arabidopsis thaliana using a genetically encoded Ca ratiometric sensor with a low-nanomolar Ca-binding affinity and a large dynamic range.

Polymeric Nanocarriers Autonomously Cross the Plant Cell Wall and Enable Protein Delivery for Stress Sensing.

Delivery of proteins in plant cells can facilitate the design of desired functions by modulation of biological processes and plant traits but is currently limited by narrow host range, tissue damage, and poor scalability. Physical barriers in plants, including cell walls and membranes, limit protein delivery to desired plant tissues. Herein, a cationic high aspect ratio polymeric nanocarriers (PNCs) platform is developed to enable efficient protein delivery to plants.

Mitigating growth-stress tradeoffs via elevated TOR signaling in rice.

Rice production accounts for approximately half of the freshwater resources utilized in agriculture, resulting in greenhouse gas emissions such as methane (CH) from flooded paddy fields. To address this challenge, environmentally friendly and cost-effective water-saving techniques have become widely adopted in rice cultivation. However, the implementation of water-saving treatments (WSTs) in paddy-field rice has been associated with a substantial yield loss of up to 50% as well as a reduction in nitrogen use efficiency (NUE).

Dynamic Proximity Tagging in Living Plant Cells with Pupylation-Based Interaction Tagging.

Identification of protein-protein interactions (PPIs) and protein kinase substrates is fundamental for understanding how proteins exert biological functions with their partners and targets. However, it is still technically challenging, especially for transient and weak interactions involved in most cellular processes. The proximity-tagging systems enable capturing snapshots of both stable and transient PPIs.

Glucose-driven TOR-FIE-PRC2 signalling controls plant development.

Nutrients and energy have emerged as central modulators of developmental programmes in plants and animals. The evolutionarily conserved target of rapamycin (TOR) kinase is a master integrator of nutrient and energy signalling that controls growth. Despite its key regulatory roles in translation, proliferation, metabolism and autophagy, little is known about how TOR shapes developmental transitions and differentiation.

ARF2-PIF5 interaction controls transcriptional reprogramming in the ABS3-mediated plant senescence pathway.

One of the hallmarks of plant senescence is the global transcriptional reprogramming coordinated by a plethora of transcription factors (TFs). However, mechanisms underlying the interactions between different TFs in modulating senescence remain obscure. Previously, we discovered that plant ABS3 subfamily MATE transporter genes regulate senescence and senescence-associated transcriptional changes.

TOR kinase, a GPS in the complex nutrient and hormonal signaling networks to guide plant growth and development.

To survive and sustain growth, sessile plants have developed sophisticated internal signalling networks that respond to various external and internal cues. Despite the central roles of nutrient and hormone signaling in plant growth and development, how hormone-driven processes coordinate with metabolic status remains largely enigmatic. Target of rapamycin (TOR) kinase is an evolutionarily conserved master regulator that integrates energy, nutrients, growth factors, hormones, and stress signals to promote growth in all eukaryotes.

ConducTORs of a Signaling Symphony: Metabolic and Hormone Responses Converge on TOR and EIN2 in plants.

Development is coordinated by dozens of signals that act in overlapping pathways to orchestrate multicellular growth. Understanding how signaling pathways intersect and diverge at a molecular level is critical to predicting how organisms will react to dynamic environmental conditions. In plants, two antagonistic signaling hubs are strictly required to sense and respond to many nutrients and hormones: TARGET OF RAPAMYCIN (TOR) and ETHYLENE INSENSITIVE 2 (EIN2).

DNA-free CRISPR-Cas9 gene editing of wild tetraploid tomato Solanum peruvianum using protoplast regeneration.

Wild tomatoes (Solanum peruvianum) are important genomic resources for tomato research and breeding. Development of a foreign DNA-free clustered regularly interspaced short palindromic repeat (CRISPR)-Cas delivery system has potential to mitigate public concern about genetically modified organisms. Here, we established a DNA-free CRISPR-Cas9 genome editing system based on an optimized protoplast regeneration protocol of S.

A Versatile and Efficient Plant Protoplast Platform for Genome Editing by Cas9 RNPs.

The ultimate goal of technology development in genome editing is to enable precisely targeted genomic changes in any cells or organisms. Here we describe protoplast systems for precise and efficient DNA sequence changes with preassembled Cas9 ribonucleoprotein (RNP) complexes in , and . Cas9 RNP-mediated gene disruption with dual gRNAs could reach ∼90% indels in Arabidopsis protoplasts.

Efficient and Economical Targeted Insertion in Plant Genomes via Protoplast Regeneration.

Versatile genome editing can be facilitated by the insertion of DNA sequences into specific locations. Current protocols involving CRISPR and Cas proteins rely on low efficiency homology-directed repair or non-homologous end joining with modified double-stranded DNA oligonucleotides as donors. Our simple protocol eliminates the need for expensive equipment, chemical and enzymatic donor DNA modification, or plasmid construction by using polyethylene glycol-calcium to deliver non-modified single-stranded DNA oligonucleotides and CRISPR-Cas9 ribonucleoprotein into protoplasts.

Primary nitrate responses mediated by calcium signalling and diverse protein phosphorylation.

Nitrate, the major source of inorganic nitrogen for plants, is a critical signal controlling nutrient transport and assimilation and adaptive growth responses throughout the plant. Understanding how plants perceive nitrate and how this perception is transduced into responses that optimize growth are important for the rational improvement of crop productivity and for mitigating pollution from the use of fertilizers. This review highlights recent findings that reveal key roles of cytosolic-nuclear calcium signalling and dynamic protein phosphorylation via diverse mechanisms in the primary nitrate response (PNR).

Model-driven discovery of calcium-related protein-phosphatase inhibition in plant guard cell signaling.

The plant hormone abscisic acid (ABA) promotes stomatal closure via multifarious cellular signaling cascades. Our previous comprehensive reconstruction of the stomatal closure network resulted in an 81-node network with 153 edges. Discrete dynamic modeling utilizing this network reproduced over 75% of experimental observations but a few experimentally supported results were not recapitulated.

Default Activation and Nuclear Translocation of the Plant Cellular Energy Sensor SnRK1 Regulate Metabolic Stress Responses and Development.

Energy homeostasis is vital to all living organisms. In eukaryotes, this process is controlled by fuel gauging protein kinases: AMP-activated kinase in mammals, Sucrose Non-Fermenting1 (SNF1) in yeast (), and SNF1-related kinase1 (SnRK1) in plants. These kinases are highly conserved in structure and function and (according to this paradigm) operate as heterotrimeric complexes of catalytic-α and regulatory β- and γ-subunits, responding to low cellular nucleotide charge.

Integration of nutrient, energy, light, and hormone signalling via TOR in plants.

The multidomain target of rapamycin (TOR) is an atypical serine/threonine protein kinase resembling phosphatidylinositol lipid kinases, but retains high sequence identity and serves a remarkably conserved role as a master signalling integrator in yeasts, plants, and humans. TOR dynamically orchestrates cell metabolism, biogenesis, organ growth, and development transitions in response to nutrient, energy, hormone, and environmental cues. Here we review recent findings on the versatile and complex roles of TOR in transcriptome reprogramming, seedling, root, and shoot growth, and root hair production activated by sugar and energy signalling.

Noncanonical ATG8-ABS3 interaction controls senescence in plants.

Protein homeostasis is essential for cellular functions and longevity, and the loss of proteostasis is one of the hallmarks of senescence. Autophagy is an evolutionarily conserved cellular degradation pathway that is critical for the maintenance of proteostasis. Paradoxically, autophagy deficiency leads to accelerated protein loss by unknown mechanisms.

Mitogen-activated protein kinases MPK3 and MPK6 are required for stem cell maintenance in the Arabidopsis shoot apical meristem.

CLV3p-mediated phosphorylation of MPK3 and MPK6 occurs via CLV1 and BAM1 receptors to regulate the maintenance of SAM development. The CLAVATA peptide-receptor (CLV3p-CLV1) pathway modulates a homeodomain master regulator WUSCHEL (WUS) transcription factor in the shoot apical meristem (SAM) with poorly defined signaling mechanisms. Here, we report that mitogen-activated protein kinases (MAPKs, also known as MPKs in plants) act in an intracellular signaling cascade to play an important role in the maintenance of SAM development.

TOR and RPS6 transmit light signals to enhance protein translation in deetiolating seedlings.

Deetiolation is an essential developmental process transforming young plant seedlings into the vegetative phase with photosynthetic activities. Light signals initiate this important developmental process by triggering massive reprogramming of the transcriptome and translatome. Compared with the wealth of knowledge of transcriptional regulation, the molecular mechanism underlying this light-triggered translational enhancement remains unclear.

Dual CLAVATA3 Peptides in Arabidopsis Shoot Stem Cell Signaling.

Plant shoot stem cell pool is constantly maintained by a negative feedback loop through peptide-receptor mediated signaling pathway. () encode a 96 amino-acid protein which is processed to 12-amino-acid or arabinosylated 13-amino-acid peptides, acting as a ligand signal to regulate stem cell homeostasis in the shoot apical meristem (SAM). Although arabinosylated 13-amino-acid CLV3 peptide (CLV3p) shows more significant binding affinity to its receptors and biological activities in the SAM, the physiological function of two mature forms of CLV3p remained an unresolved puzzle in the past decade due to the technical difficulties of arabinosylation modification in the peptide synthesis.

TOR signaling in plants: conservation and innovation.

Target of rapamycin (TOR) is an evolutionarily conserved protein kinase that plays a central role in both plants and animals, despite their distinct developmental programs and survival strategies. Indeed, TOR integrates nutrient, energy, hormone, growth factor and environmental inputs to control proliferation, growth and metabolism in diverse multicellular organisms. Here, we compare the molecular composition, upstream regulators and downstream signaling relays of TOR complexes in plants and animals.