Protocol for monitoring fatty acid trafficking from lipid droplets to mitochondria in cultured cells.

Intracellular trafficking of fatty acids (FAs) between organelles is critical for cells to adjust their metabolism in response to stimuli such as exercise, fasting, and cold exposure. Here, we describe a protocol to monitor trafficking of FAs from lipid droplets to mitochondria. We describe the labeling of organelles in cultured C2C12 myoblasts with transfection and dyes.

Contact-FP: A Dimerization-Dependent Fluorescent Protein Toolkit for Visualizing Membrane Contact Site Dynamics.

Membrane contact sites (MCSs) are sites of close apposition between two organelles used to exchange ions, lipids, and information. Cells respond to changing environmental or developmental conditions by modulating the number, extent, or duration of MCSs. Because of their small size and dynamic nature, tools to study the dynamics of MCSs in live cells have been limited.

PLIN5 interacts with FATP4 at membrane contact sites to promote lipid droplet-to-mitochondria fatty acid transport.

Cells adjust their metabolism by remodeling membrane contact sites that channel metabolites to different fates. Lipid droplet (LD)-mitochondria contacts change in response to fasting, cold exposure, and exercise. However, their function and mechanism of formation have remained controversial.

Quantitative proteomic profiling identifies global protein network dynamics in murine embryonic heart development.

Defining the mechanisms that govern heart development is essential for identifying the etiology of congenital heart disease. Here, quantitative proteomics was used to measure temporal changes in the proteome at critical stages of murine embryonic heart development. Global temporal profiles of the over 7,300 proteins uncovered signature cardiac protein interaction networks that linked protein dynamics with molecular pathways.

High Thoracic Tortuosity Is Associated With CT-Markers of Degeneration of the Descending Thoracic Aortic Wall.

Objective/introduction: Increased vascular tortuosity may be an independent marker of generalized aortic pathology. This study investigates the association between descending thoracic aortic tortuosity, aneurysm pathophysiology, and outcomes following EVAR in AAA patients.

Methods: Patients who underwent elective EVAR between 2004 and 2018 were reviewed.

The interdependent transport of yeast vacuole Ca and H and the role of phosphatidylinositol 3,5-bisphosphate.

Yeast vacuoles are acidified by the v-type H-ATPase (V-ATPase) that is comprised of the membrane embedded V complex and the soluble cytoplasmic V complex. The assembly of the V-V holoenzyme on the vacuole is stabilized in part through interactions between the V a-subunit ortholog Vph1 and the lipid phosphatidylinositol 3,5-bisphosphate (PI(3,5)P). PI(3,5)P also affects vacuolar Ca release through the channel Yvc1 and uptake through the Ca pump Pmc1.

Whole genome sequencing identifies loci specifically associated with thoracic aortic wall defects and abdominal aortic aneurysms in patients with European ancestry.

Objective: The objective of this study was to better understand the pathophysiology and underlying genetic mechanisms behind two abdominal aortic aneurysm (AAA) subtypes using computed tomographic imaging in combination with whole genome sequencing.

Methods: Patients with a known AAA and European ancestry were included in this investigation and underwent genetic and image analysis. Patients with AAAs and indications of descending thoracic aortic pathology (aortic dissection, penetrating aortic ulcers, intramural hematoma, atheromas, ulcerative plaque, and intramural ulceration, and intimal flaps/tears) were classified as having thoracic aortic disease, grouped together, and compared with patients with an AAA and a normal descending thoracic aorta.

Sustainable irrigation based on co-regulation of soil water supply and atmospheric evaporative demand.

Irrigation is an important adaptation to reduce crop yield loss due to water stress from both soil water deficit (low soil moisture) and atmospheric aridity (high vapor pressure deficit, VPD). Traditionally, irrigation has primarily focused on soil water deficit. Observational evidence demonstrates that stomatal conductance is co-regulated by soil moisture and VPD from water supply and demand aspects.

Phosphatidylinositol 3,5-bisphosphate regulates Ca transport during yeast vacuolar fusion through the Ca ATPase Pmc1.

The transport of Ca across membranes precedes the fusion and fission of various lipid bilayers. Yeast vacuoles under hyperosmotic stress become fragmented through fission events that requires the release of Ca stores through the TRP channel Yvc1. This requires the phosphorylation of phosphatidylinositol-3-phosphate (PI3P) by the PI3P-5-kinase Fab1 to produce transient PI(3,5)P pools.

Decreased Abdominal Aortic Aneurysm Size Following EVAR in Patients With CT Evidence of Subclinical Thoracic Aortic Dissection.

Background: Aneurysm sac regression following endovascular repair (EVAR) of an abdominal aortic aneurysm (AAA) is an established indicator of surgical success. However, even with a completely excluded aneurysm, the degree of aortic sac regression may vary. This study evaluates the relationship between aneurysm sac regression after EVAR and the presence of morphological features in the thoracic aorta that can be associated with a subclinical aortic dissection, termed dissection morphology in this study.

Copper blocks V-ATPase activity and SNARE complex formation to inhibit yeast vacuole fusion.

The accumulation of copper in organisms can lead to altered functions of various pathways and become cytotoxic through the generation of reactive oxygen species. In yeast, cytotoxic metals such as Hg , Cd and Cu are transported into the lumen of the vacuole through various pumps. Copper ions are initially transported into the cell by the copper transporter Ctr1 at the plasma membrane and sequestered by chaperones and other factors to prevent cellular damage by free cations.

Seasonal responses of photosynthetic parameters in maize and sunflower and their relationship with leaf functional traits.

Estimates of seasonal variation in photosynthetic capacity (P ) are critical for modeling the time course of carbon fluxes. Given the time-intensive nature of calculating P parameters via gas exchange, it is appealing to calculate parameter variation via changes in chlorophyll (Chl) and nitrogen (N) content by assuming that P scales with these variables. Although seasonal changes in P and the relationships between N and P have been evaluated in forest canopies, there is limited data on seasonal parameter values in crops, nor is it clear if seasonal changes in P can be estimated from leaf traits under the high N fertility of managed systems.

Phosphatidylinositol 3,5-bisphosphate regulates the transition between trans-SNARE complex formation and vacuole membrane fusion.

Phosphoinositides (PIs) regulate a myriad of cellular functions including membrane fusion, as exemplified by the yeast vacuole, which uses various PIs at different stages of fusion. In light of this, the effect of phosphatidylinositol 3,5-bisphosphate (PI(3,5)P) on vacuole fusion remains unknown. PI(3,5)P is made by the PI3P 5-kinase Fab1 and has been characterized as a regulator of vacuole fission during hyperosmotic shock, where it interacts with the TRP Ca channel Yvc1.

Real-Time Fluorescence Detection of Calcium Efflux During Vacuolar Membrane Fusion.

During in vitro homotypic yeast vacuole fusion Ca is transported into and out of the organelle lumen. In vitro, Ca is taken up from the medium by vacuoles upon the addition of ATP. During the docking stage of vacuole fusion Ca is effluxed from the lumen upon the formation of trans-SNARE complexes between vesicles.

Seasonal variability of the parameters of the Ball-Berry model of stomatal conductance in maize (Zea mays L.) and sunflower (Helianthus annuus L.) under well-watered and water-stressed conditions.

The Ball-Berry (BB) model of stomatal conductance (g ) is frequently coupled with a model of assimilation to estimate water and carbon exchanges in plant canopies. The empirical slope (m) and 'residual' g (g ) parameters of the BB model influence transpiration estimates, but the time-intensive nature of measurement limits species-specific data on seasonal and stress responses. We measured m and g seasonally and under different water availability for maize and sunflower.

Deleting the DAG kinase Dgk1 augments yeast vacuole fusion through increased Ypt7 activity and altered membrane fluidity.

Diacylglycerol (DAG) is a fusogenic lipid that can be produced through phospholipase C activity on phosphatidylinositol 4,5-bisphosphate [PI(4,5)P ], or through phosphatidic acid (PA) phosphatase activity. The fusion of Saccharomyces cerevisiae vacuoles requires DAG, PA and PI(4,5)P , and the production of these lipids is thought to provide temporally specific stoichiometries that are critical for each stage of fusion. Furthermore, DAG and PA can be interconverted by the DAG kinase Dgk1 and the PA phosphatase Pah1.

Estimating the sensitivity of stomatal conductance to photosynthesis: a review.

A common approach for estimating fluxes of CO and water in canopy models is to couple a model of photosynthesis (A ) to a semi-empirical model of stomatal conductance (g ) such as the widely validated and utilized Ball-Berry (BB) model. This coupling provides an effective way of predicting transpiration at multiple scales. However, the designated value of the slope parameter (m) in the BB model impacts transpiration estimates.

Phosphatidylinositol (3,4,5)-trisphosphate binds to sortilin and competes with neurotensin: Implications for very low density lipoprotein binding.

Sortilin is a multi-ligand sorting receptor that interacts with B100-containing VLDL and LDL as well as other ligands including neurotensin (NT). The current study investigates the hypothesis that phosphatidylinositol (3,4,5)-trisphosphate (PIP3) generated downstream of insulin action can directly bind to sortilin. NT binds to sortilin at a well characterized site via its carboxy terminus (C-term).

The Central Polybasic Region of the Soluble SNARE (Soluble N-Ethylmaleimide-sensitive Factor Attachment Protein Receptor) Vam7 Affects Binding to Phosphatidylinositol 3-Phosphate by the PX (Phox Homology) Domain.

The yeast vacuole requires four SNAREs to trigger membrane fusion including the soluble Qc-SNARE Vam7. The N-terminal PX domain of Vam7 binds to the lipid phosphatidylinositol 3-phosphate (PI3P) and the tethering complex HOPS (homotypic fusion and vacuole protein sorting complex), whereas the C-terminal SNARE motif forms SNARE complexes. Vam7 also contains an uncharacterized middle domain that is predicted to be a coiled-coil domain with multiple helices.

An update on the etiology of abdominal aortic aneurysms: implications for future diagnostic testing.

Abdominal aortic aneurysm (AAA) disease is multifactorial with both environmental and genetic risk factors. The current research in AAA revolves around genetic profiles and expression studies in both human and animal models. Variants in genes involved in extracellular matrix degradation, inflammation, the renin-angiotensin system, cell growth and proliferation and lipid metabolism have been associated with AAA using a variety of study designs.

An Evolving Understanding of the Genetic Causes of Abdominal Aortic Aneurysm Disease.

Aneurysms of the abdominal aorta (AAA) are relatively common - affecting as many as 8% of men and 1% of women over the age of 65. AAAs are characterized by a 50% increase in the diameter of the aneurysmal aorta compared with the normal vessel. Degeneration of structural components of the aortic wall is believed to be central in the pathogenesis of AAAs.

14-3-3 (Bmh) proteins regulate combinatorial transcription following RNA polymerase II recruitment by binding at Adr1-dependent promoters in Saccharomyces cerevisiae.

Adr1 and Cat8 are nutrient-regulated transcription factors in Saccharomyces cerevisiae that coactivate genes necessary for growth in the absence of a fermentable carbon source. Transcriptional activation by Adr1 is dependent on the AMP-activated protein kinase Snf1 and is inhibited by binding of Bmh, yeast 14-3-3 proteins, to the phosphorylated Adr1 regulatory domain. We show here that Bmh inhibits transcription by binding to Adr1 at promoters that contain a preinitiation complex, demonstrating that Bmh-mediated inhibition is not due to nuclear exclusion, inhibition of DNA binding, or RNA polymerase II (Pol II) recruitment.

Solution structure of the HIV-1 exon splicing silencer 3.

Alternative splicing of the human immunodeficiency virus type 1 (HIV-1) genomic RNA is necessary to produce the complete viral protein complement, and aberrations in the splicing pattern impair HIV-1 replication. Genome splicing in HIV-1 is tightly regulated by the dynamic assembly/disassembly of trans host factors with cis RNA control elements. The host protein, heterogeneous nuclear ribonucleoprotein (hnRNP) A1, regulates splicing at several highly conserved HIV-1 3' splice sites by binding 5'-UAG-3' elements embedded within regions containing RNA structure.

The genome of the sea urchin Strongylocentrotus purpuratus.

We report the sequence and analysis of the 814-megabase genome of the sea urchin Strongylocentrotus purpuratus, a model for developmental and systems biology. The sequencing strategy combined whole-genome shotgun and bacterial artificial chromosome (BAC) sequences. This use of BAC clones, aided by a pooling strategy, overcame difficulties associated with high heterozygosity of the genome.

The DNA sequence, annotation and analysis of human chromosome 3.

After the completion of a draft human genome sequence, the International Human Genome Sequencing Consortium has proceeded to finish and annotate each of the 24 chromosomes comprising the human genome. Here we describe the sequencing and analysis of human chromosome 3, one of the largest human chromosomes. Chromosome 3 comprises just four contigs, one of which currently represents the longest unbroken stretch of finished DNA sequence known so far.