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The metabolism of tetrahydrofolate (H₄PteGlu_n)‐bound one‐carbon (C₁) units (C₁metabolism) is multifaceted and required for plant growth, but it is unclear what of many possible synthesis pathways provide C₁units in specific organelles and tissues. One possible source of C₁units is via formate‐tetrahydrofolate ligase, which catalyzes the reversible ATP‐driven production of 10‐formyltetrahydrofolate (10‐formyl‐H₄PteGlu_n) from formate and tetrahydrofolate (H₄PteGlu_n). Here, we report biochemical and functional characterization of the enzyme fromArabidopsis thaliana(AtFTHFL). We show that the recombinant AtFTHFL has lowerK_mandk_catvalues with pentaglutamyl tetrahydrofolate (H₄PteGlu₅) as compared to monoglutamyl tetrahydrofolate (H₄PteGlu₁), resulting in virtually identical catalytic efficiencies for the two substrates. Stable transformation ofArabidopsisplants with the EGFP‐tagged AtFTHFL, followed with fluorescence microscopy, demonstrated cytosolic signal. Two independent T‐DNA insertion lines with impaired AtFTHFL function had shorter roots compared to the wild type plants, demonstrating the importance of this enzyme for root growth. Overexpressing AtFTHFL led to the accumulation of H₄PteGlu_n + 5,10‐methylene‐H₄PteGlu_nand serine, accompanied with the depletion of formate and glycolate, in roots of the transgenicArabidopsisplants. This metabolic adjustment supports the hypothesis that AtFTHFL feeds the cytosolic C₁network in roots with C₁units originating from glycolate, and that these units are then used mainly for biosynthesis of serine, and not as much for the biosynthesis of 5‐methyl‐H₄PteGlu_n, methionine, andS‐adenosylmethionine. This finding has implications for any future attempts to engineer one‐carbon unit‐requiring products through manipulation of the one‐carbon metabolic network in non‐photosynthetic organs.

 

Typical plant membranes and storage lipids are comprised of five common fatty acids yet over 450 unusual fatty acids accumulate in seed oils of various plant species. Plant oils are important human and animal nutrients, while some unusual fatty acids such as hydroxylated fatty acids (HFA) are used in the chemical industry (lubricants, paints, polymers, cosmetics, etc.). Most unusual fatty acids are extracted from non-agronomic crops leading to high production costs. Attempts to engineer HFA into crops are unsuccessful due to bottlenecks in the overlapping pathways of oil and membrane lipid synthesis where HFA are not compatible.Physaria fendlerinaturally overcomes these bottlenecks through a triacylglycerol (TAG) remodeling mechanism where HFA are incorporated into TAG after initial synthesis. TAG remodeling involves a unique TAG lipase and two diacylglycerol acyltransferases (DGAT) that are selective for different stereochemical and acyl-containing species of diacylglycerol within a synthesis, partial degradation, and resynthesis cycle. The TAG lipase interacts with DGAT1, localizes to the endoplasmic reticulum (with the DGATs) and to puncta around the lipid droplet, likely forming a TAG remodeling metabolon near the lipid droplet-ER junction. Each characterized DGAT and TAG lipase can increase HFA accumulation in engineered seed oils.

 

The plant cytokinetic microtubule array, called the phragmoplast, exhibits higher microtubule dynamics in its center (midzone) than at the periphery (distal zone). This behavior is known as the axial asymmetry. Despite being a major characteristic of the phragmoplast, little is known about regulators of this phenomenon. Here we address the role of microtubule nucleation in axial asymmetry by characterizing MACERATOR (MACET) proteins in Arabidopsis thaliana and Nicotiana benthamiana with a combination of genetic, biochemical, and live-cell imaging assays, using photo-convertible microtubule probes, and modeling. MACET paralogs accumulate at the shrinking microtubule ends and decrease the tubulin OFF rate. Loss of MACET4 and MACET5 function abrogates axial asymmetry by suppressing microtubule dynamicity in the midzone. MACET4 also narrows the microtubule nucleation angle at the phragmoplast leading edge and functions as a microtubule tethering factor for AUGMIN COMPLEX SUBUNIT 7 (AUG7). The macet4 macet5 double mutant shows diminished clustering of AUG7 in the phragmoplast distal zone. Knockout of AUG7 does not affect MACET4 localization, axial asymmetry, or microtubule nucleation angle, but increases phragmoplast length and slows down phragmoplast expansion. The mce4-1 mce5 aug7-1 triple knockout is not viable. Experimental data and modeling demonstrate that microtubule nucleation factors regulate phragmoplast architecture and axial asymmetry directly by generating new microtubules and indirectly by modulating the abundance of free tubulin. 

Organelles function as hubs of cellular metabolism and elements of cellular architecture. In addition to 3 spatial dimensions that describe the morphology and localization of each organelle, the time dimension describes complexity of the organelle life cycle, comprising formation, maturation, functioning, decay, and degradation. Thus, structurally identical organelles could be biochemically different. All organelles present in a biological system at a given moment of time constitute the organellome. The homeostasis of the organellome is maintained by complex feedback and feedforward interactions between cellular chemical reactions and by the energy demands. Synchronized changes of organelle structure, activity, and abundance in response to environmental cues generate the fourth dimension of plant polarity. Temporal variability of the organellome highlights the importance of organellomic parameters for understanding plant phenotypic plasticity and environmental resiliency. Organellomics involves experimental approaches for characterizing structural diversity and quantifying the abundance of organelles in individual cells, tissues, or organs. Expanding the arsenal of appropriate organellomics tools and determining parameters of the organellome complexity would complement existing -omics approaches in comprehending the phenomenon of plant polarity. To highlight the importance of the fourth dimension, this review provides examples of organellome plasticity during different developmental or environmental situations.

 

Abstract TPX2 proteins were first identified in vertebrates as a key mitotic spindle assembly factor. Subsequent studies demonstrated that TPX2 is an intricate protein, with functionally and structurally distinct domains and motifs including Aurora kinase-binding, importin-binding, central microtubule-binding, and C-terminal TPX2 conserved domain, among others. The first plant TPX2-like protein, WAVE-DAMPENED2, was identified in Arabidopsis as a dominant mutation responsible for reducing the waviness of roots grown on slanted agar plates. Each plant genome encodes at least one ‘canonical’ protein with all TPX2 domains and a family of proteins (20 in Arabidopsis) that diversified to contain only some of the domains. Although all plant TPX2-family proteins to date bind microtubules, they function in distinct processes such as cell division, regulation of hypocotyl cell elongation by hormones and light signals, vascular development, or abiotic stress tolerance. Consequently, their expression patterns, regulation, and functions have diverged considerably. Here we summarize the current body of knowledge surrounding plant TPX2-family proteins.