Lipid Droplet-Organelle Contact Sites as Hubs for Fatty Acid Metabolism, Trafficking, and Metabolic Channeling

Cells prepare for fluctuations in nutrient availability by storing energy in the form of neutral lipids in organelles called Lipid Droplets (LDs). Upon starvation, fatty acids (FAs) released from LDs are trafficked to different cellular compartments to be utilized for membrane biogenesis or as a source of energy. Despite the biochemical pathways being known in detail, the spatio-temporal regulation of FA synthesis, storage, release, and breakdown is not completely understood. Recent studies suggest that FA trafficking and metabolism are facilitated by inter-organelle contact sites that form between LDs and other cellular compartments such as the Endoplasmic Reticulum (ER), mitochondria, peroxisomes, and lysosomes. LD-LD contact sites are also sites where FAs are transferred in a directional manner to support LD growth and expansion. As the storage site of neutral lipids, LDs play a central role in FA homeostasis. In this mini review, we highlight the role of LD contact sites with other organelles in FA trafficking, channeling, and metabolism and discuss the implications for these pathways on cellular lipid and energy homeostasis.

Regulatory phosphorylation site tunes Phosphoglucomutase 1 as a metabolic valve to control mobilization of glycogen stores.

Regulation of glycogen metabolism is of vital importance in organisms of all three kingdoms of life. Although the pathways involved in glycogen synthesis and degradation are well known, many regulatory aspects around the metabolism of this polysaccharide remain undeciphered. Here, we used the unicellular cyanobacterium Synechocystis as a model to investigate how glycogen metabolism is regulated in dormant nitrogen-starved cells, which entirely rely on glycogen catabolism to restore growth. We found that the activity of the enzymes involved in glycogen synthesis and degradation is tightly controlled at different levels via post-translational modifications. Phosphorylation of phosphoglucomutase 1 (Pgm1) on a peripheral residue (Ser63) regulates Pgm1 activity and controls the mobilization of the glycogen stores. Inhibition of Pgm1 activity via phosphorylation on Ser63 appears essential for survival of Synechocystis in the dormant state. Remarkably, this regulatory mechanism seems to be conserved from bacteria to humans. Moreover, phosphorylation of Pgm1 influences the formation of a metabolon, which includes Pgm1, oxidative pentose phosphate cycle protein (OpcA) and glucose-6-phosphate dehydrogenase (G6PDH). Analysis of the steady-state levels of the metabolic products of glycogen degradation together with protein-protein interaction studies revealed that the activity of G6PDH and the formation of this metabolon are under additional redox control, likely to ensure metabolic channeling of glucose-6-phosphate to the required pathways for each developmental stage.

Indole-3-glycerolphosphate synthase, a branchpoint for the biosynthesis of tryptophan, indole, and benzoxazinoids in maize

The maize (Zea mays) genome encodes three indole-3-glycerolphosphate synthase enzymes (IGPS1, 2, and 3) catalyzing the conversion of 1-(2-carboxyphenylamino)-l-deoxyribulose-5-phosphate to indole-3-glycerolphosphate. Three further maize enzymes (BX1, benzoxazinoneless 1; TSA, tryptophan synthase α subunit; and IGL, indole glycerolphosphate lyase) convert indole-3-glycerolphosphate to indole, which is released as a volatile defense signaling compound and also serves as a precursor for the biosynthesis of tryptophan and defense-related benzoxazinoids. Phylogenetic analyses showed that IGPS2 is similar to enzymes found in both monocots and dicots, whereas maize IGPS1 and IGPS3 are in monocot-specific clades. Fusions of yellow fluorescent protein (YFP) with maize IGPS enzymes and indole-3-glycerolphosphate lyases were all localized in chloroplasts. In bimolecular fluorescence complementation assays, IGPS1 interacted strongly with BX1 and IGL, IGPS2 interacted primarily with TSA, and IGPS3 interacted equally with all three indole-3-glycerolphosphate lyases. Whereas IGPS1 and IGPS3 expression was induced by insect feeding, IGPS2 expression was not. Transposon insertions in IGPS1 and IGPS3 reduced the abundance of both benzoxazinoids and free indole. Spodoptera exigua (beet armyworm) larvae show improved growth on igps1 mutant maize plants. Together, these results suggest that IGPS1 and IGPS3 function mainly in the biosynthesis of defensive metabolites, whereas IGPS2 may be involved in the biosynthesis of tryptophan. This metabolic channeling is similar, though less exclusive than that proposed for the three maize indole-3-glycerolphosphate lyases.

Local PI(4,5)P2 pool dynamics detected by the coincidence biosensor tubbyCT maintain the integrity of ER-PM junctions during PLC signaling

ABSTRACTPhosphoinositides (PIs) are important signaling molecules and determinants of membrane identity in the eukaryotic plasma membrane, where they multi-task in divergent signaling pathways. Signaling pleiotropy likely depends on distinct PI pools in the same membrane, although the physical definition of such pools has remained ambiguous. PI(4,5)P2, specifically, is also the precursor for the second messengers in the Gq/PLC pathway, IP3 and DAG, and is broken down by PLCβ during signaling. Endoplasmic reticulum-plasma membrane contact sites (ER-PM junctions) have emerged as central hubs for lipid transport between both membranes, and specifically for PI homeostasis by supplying the PM with phosphatidylinositol.Here we show that the tubby protein, by virtue of its C-terminal tubby-domain, preferentially localizes to ER-PM junctions by binding to both PI(4,5)P2 and the ER-PM tether E-Syt3. Under conditions of vigorous PI(4,5)P2 consumption by PLCβ, additional recruitment of tubby revealed an increase of a local PI(4,5)P2 pool fed by local synthesis through PI kinases. Inhibition of this pool-filling process led to the release of the ER-PM tethers, E-Syts, from the membrane and hence to loss of integrity of the ER-PM contact sites.We conclude that spatiotemporal metabolic channeling of PI synthesis initiated by non-vesicular transport in the ER-PM junctions specifies a local pool of PI(4,5)P2 that is pivotal for the maintenance of homeostatic functions during global depletion of PI(4,5)P2. The findings further suggest that the tubby-like proteins (TULPs), so far known to impact on energy homeostasis and obesity through primary cilia signaling, have an additional function at ER-PM junctions.HIGHLIGHTSThe tubby domain preferentially assembles into ER-PM junctions due to coincidence detection of PI(4,5)P2 and E-Syt3Tubby recruitment reveals an increase of a local pool of PI(4,5)P2 in ER-PM junctions during PLCβ signalingJunctional PI(4,5)P2 dynamics require local synthesis of PI(4,5)P2Local PI(4,5)P2 supply is required for integrity of ER-PM junctions during PLCβ activity.

Transcriptional analysis of arogenate dehydratase genes identifies a link between phenylalanine biosynthesis and lignin biosynthesis

Biogenesis of the secondary cell wall in trees involves the massive biosynthesis of the phenylalanine-derived polymer lignin. Arogenate dehydratase (ADT) catalyzes the last, and rate-limiting, step of the main pathway for phenylalanine biosynthesis. In this study, we found that transcript levels for several members of the large ADT gene family, including ADT-A and ADT-D, were enhanced in compression wood of maritime pine, a xylem tissue enriched in lignin. Transcriptomic analysis of maritime pine silenced for PpMYB8 revealed that this gene plays a critical role in coordinating the deposition of lignin with the biosynthesis of phenylalanine. Specifically, it was found that ADT-A and ADT-D were strongly down-regulated in PpMYB8-silenced plants and that they were transcriptionally regulated through direct interaction of this transcription factor with regulatory elements present in their promoters. Another transcription factor, PpHY5, exhibited an expression profile opposite to that of PpMYB8 and also interacted with specific regulatory elements of ADT-A and ADT-D genes, suggesting that it is involved in transcriptional regulation of phenylalanine biosynthesis. Taken together, our results reveal that PpMYB8 and PpHY5 are involved in the control of phenylalanine formation and its metabolic channeling for lignin biosynthesis and deposition during wood formation in maritime pine.