bluuebunny/crossref_metadata_2025_split huggingface dataset:
Novel Regulation of the Stability of Transcriptional Cofactors Ski and SnoN by GPCR Signals and Actin‐Cytoskeleton Dynamics
Authors: Macias‐Silva Marina , Vazquez‐Victorio Genaro , Caligaris Cassandre 
Published: November 2023

TGF‐β signaling controls several cellular processes from growth arrest to apoptosis in epithelial cells. The transcriptional cofactors Ski and SnoN are main regulators of TGF‐β signaling by sequestering SMAD proteins or recruiting transcriptional repressor complexes at TGF‐β/SMADs target genes. TGF‐β signaling regulates the stability of Ski and SnoN proteins through the ubiquitin‐proteasome system. Even though the important role that Ski and SnoN proteins play in early development and homeostasis, cell cycle was the unique signal besides TGF‐β pathway that regulated their stability. Nevertheless, the mechanisms that regulate Ski and SnoN protein stability in diverse cellular processes remain unknown. In this study, we found out that the actin cytoskeleton dynamics is a novel regulator of Ski and SnoN protein stability in hepatocytes. Inhibitors of actin cytoskeleton dynamics and cell confluence status regulate Ski and SnoN protein levels. Actin polymerization is a signal for Ski protein degradation via proteasome. We also demonstrated that G protein‐coupled receptors (GPCR) signaling control Ski and SnoN protein levels through promoting and decreasing its stability by cAMP or Rho signals, respectively. In conclusion, we identified a new regulation for Ski and SnoN protein stability by changes in actin cytoskeleton rearrangements. These findings suggest that changes in Ski and SnoN protein stability could be controlled by this novel regulation in physiological and pathological events as a result of changes in actin cytoskeleton dynamics induced by cell contact, extracellular matrix remodeling and soluble ligands; thus, it will be important to evaluate TGF‐β/SMADs signaling in all these cellular contexts.Support or Funding InformationThis work is supported by grants from DGAPA/PAPIIT/UNAM (IN208115) and CONACYT (No. 240224).
Role of the Orphan Receptors RORα and REV‐Erb α/β in Liver Zonal Gene Regulation
Authors: Schroder Kristofer R , Clinkenbeard Erica , Spear Brett 
Published: November 2023

Numerous hepatic metabolic pathways, such as glycolysis and gluconeogenesis, cannot occur in the same cell. To overcome this, opposing pathways are compartmentalized within hepatocytes located within different regions of the liver, a phenomenon called metabolic zonation or zonal regulation. The expression of hepatic genes in different zonal regions is orchestrated by multiple transcription factors. Studies by Benhamouche et al. (Dev. Cell, 10:759) revealed that activation of the β‐catenin signaling pathway, through binding of β‐catenin to TCF4, is required for expression of genes that are active solely in hepatocytes surrounding the central vein (pericentral region). In hepatocytes around the portal triad (periportal region), APC inhibits β‐catenin and prevents expression of pericentral genes. A genome wide study of chromatin occupancy of β‐catenin, TCF4, and the liver transcription factor HNF4α revealed that TCF4 is an opportunistic transcription factor that shifts between interacting with HNF4α in periportal hepatocytes and β‐catenin in pericentral hepatocytes.Previous transgenic studies in our lab demonstrated that enhancer E3 of the activity of mouse alpha‐fetoprotein (AFP) was active in the adult liver, even though AFP expression is silenced at birth, Interestingly, E3 activity is highly restricted to a single layer of hepatocytes surrounding the central vein, thus providing a system to investigate the molecular basis of pericentral gene expression (Peyton et al, PNAS, 97:10890). Additional studies indicate that E3 activity in pericentral hepatocytes requires β‐catenin signaling, since E3 is no longer active in b‐catenin deficient hepatocytes. Additional in vitro studies indicate that a highly conserved TCF4/β‐catenin binding site at the 3′ end of E3 is required for responsiveness to b‐catenin (Clinkenbeard et al, Hepatology, 56:1892). We have also identified a site at the 5′ end of E3 that binds the related orphan nuclear receptors RORα, Rev‐Erbα, and Rev‐Erbβ. Additional data in cultured cells and mice suggest that RORα is a positive regulator of E3, whereas Rev‐Erbα/β repress E3 activity.Based on these data, we propose that differential binding of RORa and Rev‐Erbα/β contribute to zonal E3 activity in the adult liver. We proposed that RORa binding is required for E3 activity in pericentral hepatocytes. In contrast, the binding of Rev‐Erbα/β represses E3 activity in periportal regions. To test this, we are using ChIP to evaluate RORα and Rev‐Erbα/β binding to E3 in enriched pericentral and periportal hepatocytes. We will also knock down Rev‐Erbα/β levels in the adult liver and test whether this results in increased expression of E3 transgenes (and other pericentral genes) in periportal regions. In conclusion, we present data that indicates that RORα and Rev‐Erbα/β regulate the zonal expression of AFP through binding to E3.
Regulation of the mRNA reading frame
Authors: Dunham Christine M. , Sunita S. , Dunkle Jack , Maehigashi Tatsuya 
Published: November 2023

The ribosome is a massive, universally conserved macromolecular machine responsible for all cellular protein synthesis. The ribosome selects correct tRNAs to decode the mRNA, facilitates peptide bond formation, and then moves the tRNAs through its three functionally distinct tRNA binding sites in a dynamic and exquisitely orchestrated manner. Accurate maintenance of the universal mRNA three‐nucleotide code (or “reading frame”) is critical for correct protein expression but the molecular details of how this is controlled, or deviated from in a programmed manner, are not well understood. The absolute requirement for the precise correlation between the mRNA frame and the correct protein sequence expressed underlies an important fundamental but unanswered question in molecular biology: what regulates the mRNA reading frame? To address this question, we study two examples of defined biological mechanisms that subvert the three‐nucleotide mRNA reading frame and undergo high levels of frameshifting, i.e. movement to a noncanonical reading of the mRNA template. Here we describe two ribosomal frameshift events, a +1 (i.e. four rather than the normal three mRNA nucleotide) reading of the mRNA template by frameshift suppressor tRNAs and a ‐1 (i.e. two mRNA nucleotide) reading of the mRNA template induced by complex mRNA tertiary structures proximal to the surface of the ribosome. Our biochemical and structural results reveal that tRNA distortion and conformational changes of the small ribosomal subunit are induced by frameshift‐prone tRNAs and mRNAs. We propose nonproductive EF‐G interaction with the ribosome results in a pausing of EF‐G‐mediated translocation of the mRNA‐tRNA pair allowing for the shift in a new mRNA frame.Support or Funding InformationNIH R01GM093278; Pew Biomedical Trusts
A Conserved Family of Proteins Facilitates Nascent Lipid Droplet Budding from the ER
Authors: Choudhary Vineet , Ojha Namrata , Golden Andy , Prinz William A. 
Published: November 2023

Lipid droplets (LDs) are found in almost all cells and play central roles in energy and lipid metabolism; it is also becoming clear that LDs have additional functions, including important roles in protein degradation, ER stress, and viral replication. Unlike all other membrane‐bound organelles, LDs are surrounded by a phospholipid monolayer rather than a bilayer. How this unique structure is formed has remained an important unsolved mystery for many years. Quite a number of models of LD biogenesis have been proposed but the simplest and arguably most popular is one that has sometimes been termed the “lens” model. Neutral lipids, triacylglycerols (TAG) and steryl esters (SE) which form the core of all LDs, are synthesized in the ER and it is thought that as these lipids accumulate in the ER bilayer they form a lens (or blister) in the ER that can grow and bud. We visualized nascent LD formation in yeast using electron microscopy (EM)‐tomography and found that the lens model is correct. Lens of neutral lipid approximately 30–60 nm wide were found between the two leaflets of the ER bilayer. This is the first time the beginnings of LD biogenesis have been visualized and resolves a long‐standing question in LD biogenesis. We next asked whether proteins are necessary to facilitate nascent LD budding from the ER and focused on a conserved family of proteins, called fat storage‐inducing transmembrane (FIT) proteins. Yeast has two FITs (called Fit2a and Fit2b) and we found that in cells missing both most LDs are wrapped by the ER membrane and that this is caused by LD budding into the ER lumen rather than the cytosol. The wrapping membrane is not derived from the phagophore or related to autophagy since we found that wrapping still occurred in mutants lacking the FIT proteins and various proteins required for autophagy. The role of FIT proteins in LD budding is conserved; knock down of FIT2 in 3T3 fibroblasts and in C.elegans caused LDs to become wrapped in the ER. Interestingly deletion of the single FIT protein in C. elegans is lethal, suggesting that LD budding is an essential process in this organism. Our findings indicate that nascent LDs form lenses in the ER and that FIT proteins are necessary to promote proper budding of LDs from the ER. The mechanism of how FIT proteins control the directionality of LD budding remains to be investigated.Support or Funding InformationThis work was supported by the Intramural Research Program of the National Institute of Diabetes and Digestive and Kidney Diseases. Vineet Choudhary is supported by fellowship from Swiss National Science Foundation (Grant Nr; PA00P3_145358, P300P3_158454).
ATGL Is Not Required for Oleate Protection from Lipotoxicity in NIH3T3 Cells
Authors: Roelike Lauren , Plasek Leah , Mashek Douglas G. , Listenberger Laura L. 
Published: November 2023

The long chain, saturated fatty acid palmitate is toxic to cultured cells. However, co‐supplementation with monounsaturated oleate promotes storage of excess palmitate in lipid droplets and prevents cell death. We tested whether events downstream of lipid storage in lipid droplets contribute to oleate protection from lipotoxicity. Specifically, we asked whether protection from lipotoxicity requires adipose triglyceride lipase (ATGL) or sirtuin 1 (SIRT1) activity. We found that oleate continues to prevent palmitate induction of caspase 3/7 activity in NIH3T3 cells in the presence of ATGListatin, an inhibitor of ATGL. Moreover, oleate and palmitate co‐supplementation does not increase the activity of SIRT1, a transcriptional co‐activator downstream of ATGL, nor does it lead to ATGL‐dependent changes in expression of ACOT1, ACSL1, or CPT1α mRNA. Thus, our results fail to support a role for ATGL in oleate protection from lipotoxicity in NIH3T3 cells. Additional pathways through which oleate protects a cell from lipotoxicity should be explored.Support or Funding InformationThis research was funded by the Arnold and Mabel Beckman Foundation and St. Olaf College CURI.
Mitochondria‐Generated Acetyl‐CoA Regulates Gene Expression in Hepatic Lipid Synthesis
Authors: Patel Mulchand S , Mahmood Saleh , Birkaya Barbara , Rideout Todd C 
Published: November 2023

During the absorptive state the liver stores excess glucose as glycogen and synthesizes fatty acids for triglyceride synthesis for export as very low density lipoproteins. For de novo synthesis of fatty acids from glucose, the mitochondrial pyruvate dehydrogenase complex (PDC) plays a gatekeeper's role for generation of the carbon‐flux as acetyl‐CoA. We have shown previously that there was no incorporation of radioactive glucose‐carbon into newly synthesized fatty acids in livers from liver‐specific PDC‐deficient male mice (L‐PDHKO) due to the null mutation of the Pdha1 gene. In the present study, although the postnatal growth of L‐PDHKO mice was largely unaltered, they developed hyperinsulinemia with lower blood glucose levels in the fed state. Serum and liver lipid triglyceride and cholesterol levels remained unaltered in L‐PDHKO mice. To determine the effect of liver‐specific PDC deficiency on lipid and glucose metabolism, we investigated expression of several key genes as well as their upstream regulators in lipid and glucose metabolism. Interestingly, there was significant downregulation of expression of several key genes (such as ATP‐citrate lyase, acetyl‐CoA carboxylase) in the de novo fatty acid synthesis pathway and their upstream regulators (such as LXR, SREBP1, ChREBP). Downregulation of several genes in the metabolism of glucose (glucose‐6‐phosphate dehydrogenase, pyruvate kinase, phosphoenolpyruvate carboxykinase) and fatty acid oxidation (fatty acid transporter, carnitine‐palmitoyl‐CoA transferase‐1) were also observed in livers of L‐PDHKO mice. Interestingly, there was upregulation of lipogenic genes in epididymal adipose tissue from L‐PDHKO mice. Although, the total hepatic acetyl‐CoA content remained unaltered in these mice, modified acetylation profiles of several proteins in the nucleo‐cytoplasmic compartment (including increased level of acetylated histone3lysine9, H3K9) in livers of L‐PDHKO mice suggest an important role of mitochondria‐generated acetyl‐CoA from glucose‐derived pyruvate by PDC in gene expression in hepatic lipid metabolism. This finding has important implications for regulation of hepatic lipid biosynthesis in obesity, type 2 diabetes and non‐alcoholic fatty liver disease.Support or Funding Information(Supported in part by NIH Grant DK20478 and Department Fund, The Research Foundation of The State University of New York.)
A Novel Mouse Model with Targeted and Inducible Deletion of Intestinal CTP:Phosphocholine Cytidylyltransferase α
Authors: Kennelly John Paul , van der Veen Jelske , Nelson Randy , da Silva Robin , Kelly Karen , Leonard Kelly‐Ann , Wan Sereana , Jacobs René 
Published: November 2023

Phosphatidylcholine (PC) is the primary phospholipid in mammalian cell membranes, bile, lipid droplets and plasma lipoproteins, and also serves as a precursor for signalling molecules. PC is produced from choline by the CDP‐choline (Kennedy) pathway in all nucleated cells, and the enzyme CTP: phosphocholine Cytidylyltransferase (CT) regulates flux through the pathway. CTα is the predominant isoform in most major tissues, including the liver and intestine. CTα is an important regulator of very‐low density lipoprotein (VLDL) and high‐density lipoprotein (HDL) metabolism the liver. CT also functions to maintain the structural integrity of membranes and to incorporate fatty acids into lipid droplets to avoid cell toxicity, roles which may be particularly important in the intestine which sees large quantities of dietary lipid. However, our understanding of the role of CT and PC in intestinal lipid and lipoprotein metabolism is incomplete.To determine the functional importance of CTα in regulating intestinal lipid and lipoprotein metabolism in vivo, we have generated mice in which we can induce partial (CTα −/+) or complete (CTα −/−) deletion of CTα using a tamoxifen‐inducible Cre‐Lox system. Preliminary data suggest that CTα+/− mice have altered levels of PC in the intestine and impaired chylomicron secretion as compared to controls with normal intestinal CTα activity. Surprisingly, plasma triglyceride was also increased in CTα+/− mice. Future studies will investigate the compensatory mechanisms in the intestine in response to impaired PC supply.Support or Funding InformationNSERC and CIHR
Valproate increases ceramide levels and induces the unfolded protein response
Authors: Jadhav Shyamalagauri , Russo Sarah , Cowart Ashley , Greenberg Miriam 
Published: November 2023

Bipolar disorder (BD), which is characterized by depression and mania, affects about 1% of the world population. Current treatments are effective in only 40–60% of cases and cause severe side effects. Valproic acid (VPA), a branched short‐chain fatty acid, is one of the most widely used drugs for the treatment BD. Although many hypotheses have been postulated to explain the molecular mechanism of action of this drug in BD, the therapeutic mechanism is not understood. This knowledge gap has hampered the development of new drugs to treat this disorder. To identify candidate pathways affected by VPA, we performed a genome wide expression analysis in yeast cells grown in the presence and absence of VPA.In response to VPA, we observed upregulation of FEN1 and SUR4, encoding fatty acid elongases that catalyze the synthesis of very long chain fatty acids (C24 to C26) required for the synthesis of ceramide. Interestingly, fen1Δ and sur4Δ exhibit VPA sensitivity. VPA also increases levels of ceramides, especially those that contain C24 and C26 fatty acids. Ceramide acts as a signaling molecule that downregulates the expression of amino acid transporters and induces stress by reducing the intracellular levels of amino acids. Consistent with an increase in ceramide, VPA decreases the expression of amino acid transporters. In addition, VPA increases the expression of ER chaperone proteins (markers of UPR) as well as activates the unfolded protein response element (UPRE), suggesting that VPA induces the UPR pathway. In the presence of the ceramide synthase inhibitor fumonisin, the UPR pathway is not induced by VPA, suggesting that VPA induction of the UPR is mediated by ceramide. Studies show that BD patients exhibit a decreased UPR activation, decreased expression of ER chaperone proteins and a (‐116C‐‐>G) polymorphism in XBP1, a key transcription factor that upregulates the UPR. This study identifies the UPR pathway as a potential new target that could be important in the therapeutic action of VPA.
Ca2+‐dependent Pyk2 Activation Destabilizes Endothelial Adherens Junctions by Disrupting Interaction between VE‐PTP and VE‐Cadherin
Authors: Soni Dheeraj , DebRoy Auditi , Wang DongMei , Vogel Stephen M , Tiruppathi Chinnaswamy 
Published: November 2023

Expression of vascular endothelial cadherin (VE‐cad) and its interacting protein partner vascular endothelial protein tyrosine phosphatase (VE‐PTP) at endothelial Adherens Junctions (AJs) is vital for endothelial barrier integrity. Previously, we showed that stromal interacting molecule 1 (STIM1)‐activated Ca2+ entry disassembles endothelial AJs to increase vascular permeability. However, the signaling pathways activated secondary to STIM1‐activated Ca2+ entry (i.e., store‐operated Ca2+ entry [SOCE]) in endothelial cells, which mediate the disassembly of endothelial AJs, are not fully understood. Here we provide genetic evidence that SOCE signaling disrupts the association between VE‐cad and VE‐PTP at endothelial AJs by activation of the Ca2+‐dependent tyrosine kinase Pyk2. We generated endothelial cell‐restricted STIM1 knockout (Stim1ΔEC) mice by crossing loxP‐flanked STIM1 (Stim1fl/fl) mice with Cdh5‐Cre transgenic mice. Lung endothelial cells (LECs) from Stim1ΔEC mice showed abrogation of SOCE in response to protease‐activated receptor‐1 (PAR‐1) activation with either thrombin or PAR‐1 agonist peptide (TFLLRNPNDK‐NH2). Further, we observed that PAR‐1‐induced Pyk2 activation (measured by Pyk2 phosphorylation at Tyr‐402) and VE‐PTP inactivation (measured by VE‐PTP phosphorylation) were prevented in LECs of Stim1ΔEC mice. Consistent with these results, PAR‐1 induced VE‐cad phosphorylation was markedly suppressed in LECs from Stim1ΔEC mice. As expected, PAR‐1 activation failed to disassemble endothelial AJs in LECs of Stim1ΔEC mice. Moreover, pharmacological inhibition or silencing of Pyk2 in human lung vascular endothelial cells prevented endothelial AJ disassembly and inactivation of VE‐PTP in response to thrombin. These findings suggest that SOCE signaling regulates endothelial AJs through Pyk2‐mediated inactivation of VE‐PTP.Support or Funding InformationNIH/HLBI 1R01HL128359‐01
Physicochemical properties and plasma lipid profile of walnut (Tetracapidium conophorum) oil‐based diet in rats
Authors: Fadaka Adewale Oluwaseun , Ajayi Olubunmi 
Published: November 2023

ContextWalnuts are widely distributed all over the world, and they are common in Nigeria. On a global basis, walnuts rank second behind almonds in tree nut production. In 2010, global production of walnuts was 1,500,000 t. Walnuts are receiving increasing interest as a healthy foodstuff because their regular consumption has been reported to decrease the risk of coronary heart disease. Therefore, walnuts can be utilized as ingredients of many foodstuffs such as bakery products to enhance the nutrition value and sensory properties of the final product. The physicochemical properties of walnut oil (Tetracapidium conophorum) and the plasma lipid profile were investigated in Wistar rats.ObjectiveThis study determines the physicochemical properties, fatty acid composition and the effect of Tetracapidiun conophorum oil on the lipid profile of wister rats.Materials and methodsThe rats were assigned into three groups A, B, and C. Group A was the control fed with commercial oil based diet; Group B and C were fed with 5% and 10% walnut oil based diet, respectively. After six weeks of feeding trial, the lipid profile was evaluated.ResultThe physicochemical properties include the Refractive index (1.35), Acid value (1.01), free fatty acid (0.25), Saponification number (191.45) and Relative density with (0.64). The main fatty acids of walnut oil were oleic, linolenic, stearic and palmitic acids. Oleic acid content of the walnut was found to be 67.82 while that of linolenic is 17.06. The result shows that there is no significant difference between the test groups of triglycerides and LDL/HDL ratio but shows a reduction relative to control (P<0.05). Significant decrease was observed in the level of TC, HDL‐C and LDL‐C but there is no change in the test group of HDL‐C. The reduction observed in TC was concentration dependent.Discussion and ConclusionAs a result, the present study showed that walnut oil could be a potential source of valuable oil which might be used for edible and other industrial applications and may also be useful for the evaluation of dietary information.
Digging into rice diterpenoid biosynthesis
Authors: Peters Reuben 
Published: November 2023

Rice (Oryza sativa) is an important crop plant whose susceptibility to disease can critically impact food security. Among the means by which rice resists microbial pathogens is the production of diterpenoid phytoalexins, although the importance of such natural products biosynthesis is not fully understood. My group has taken a systematic approach towards elucidating both the underlying metabolic network and the physiological relevance of the resulting diterpenoids. Our work on diterpenoid metabolism has been based on the extensive sequence information available for rice (i.e., functional genomics), and includes the discovery of relevant biosynthetic gene clusters, as well as development of a synthetic biology approach in which we reconstitute rice metabolic pathways in E. coli, relying on the use of synthetic genes for functional incorporation of cytochrome P450 mono‐oxygenases. Based on our comprehensive map for the early steps mediated by diterpene synthase in rice metabolism, we also are now taking a reverse genetic approach towards elucidating the biological function of the resulting diterpenoid natural products, including the application of genome editing technology. Our most recent results will be discussed.Support or Funding InformationNIH (GM109773)USDA‐NIFA‐AFRI (2014‐67013‐21720)