TFAP2 transcription factors are regulators of lipid droplet biogenesis

How trafficking pathways and organelle abundance adapt in response to metabolic and physiological changes is still mysterious, although a few transcriptional regulators of organellar biogenesis have been identified in recent years. We previously found that the Wnt signaling directly controls lipid droplet formation, linking the cell storage capacity to the established functions of Wnt in development and differentiation. In the present paper, we report that Wnt-induced lipid droplet biogenesis does not depend on the canonical TCF/LEF transcription factors. Instead, we find that TFAP2 family members mediate the pro-lipid droplet signal induced by Wnt3a, leading to the notion that the TFAP2 transcription factor may function as a ‘master’ regulator of lipid droplet biogenesis

DN-p73 is activated after DNA damage in a p53-dependent manner to regulate p53-induced cell cycle arrest

p53 and p73 genes are both activated in response to DNA damage to induce either cell cycle arrest or apoptosis, depending on the strength and the quality of the damaging stimulus. p53/p73 transcriptional activity must be tightly regulated to ensure that the appropriate biological response is achieved and to allow the cell to re-enter into the cell cycle after the damage has been repaired. In addition to multiple transcriptionally active (TA) isoforms, dominant negative (DN) variants, that lack the amino-terminal transactivation domain and function as trans-repressors of p53, p63 and p73, are expressed from a second internal promoter (P2-p73Pr). Here we show that, in response to a non apoptotic DNA damage induced by low doses of doxorubicin, p53 binds in vivo, as detected by a p53-specific chromatin immunoprecipitation assay, and activates the P2-p73 promoter. DN-p73alpha protein accumulates under the same conditions and exogenously expressed DN-p73alpha is able to counteract the p53-induced activation of the P2-p73Pr. These results suggest that DN-p73 may contribute to the autoregulatory loops responsible for the termination of p53/p73 responses in cells that do not undergo apoptosis. Accordingly, the activation of the P2-p73Pr is markedly enhanced in both p73-/- murine fibroblasts and in human cells in which p73 transcripts are selectively knocked-out by p73-specific small interfering RNAs

Antibody against apolipoprotein-A1, non-alcoholic fatty liver disease and cardiovascular risk:a translational study

Background: Non-alcoholic fatty liver disease (NAFLD) is a common liver disease increasing cardiovascular disease (CVD) morbidity and mortality. Autoantibodies against apolipoprotein A-1 (AAA-1) are a possible novel CVD risk factor promoting inflammation and disrupting cellular lipid homeostasis, two prominent pathogenic features of NAFLD. We explored the role of AAA-1 in NAFLD and their association with CVD risk. Methods: HepaRG cells and liver sections from ApoE−/− mice exposed to AAA-1 were used for lipid quantification and conditional protein expression. Randomly selected sera from 312 subjects of the Prevention of Renal and Vascular End-stage Disease (PREVEND) general population cohort were used to measure AAA-1. A Fatty Liver Index (FLI) ≥ 60 and a 10-year Framingham Risk Score (FRS) ≥ 20% were used as proxy of NAFLD and high CVD risk, respectively. Results: In-vitro and mouse models showed that AAA-1 increased triglyceride synthesis leading to steatosis, and promoted inflammation and hepatocyte injury. In the 112 PREVEND participants with FLI ≥ 60, AAA-1 were associated with higher FRS, alkaline phosphatase levels, lower HDL cholesterol and tended to display higher FLI values. Univariate linear and logistic regression analyses (LRA) confirmed significant associations between AAA-1, FLI and FRS ≥ 20%, while in adjusted LRA, FLI was the sole independent predictor of FRS ≥ 20% (OR: 1.05, 95%CI 1.01–1.09, P = 0.003). AAA-1 was not an independent FLI predictor. Conclusions: AAA-1 induce a NAFLD-compatible phenotype in vitro and in mice. Intricate associations exist between AAA-1, CVD risk and FLI in the general population. Further work is required to refine the role of AAA-1 in NAFLD and to determine if the AAA-1 association with CVD is affected by hepatic steatosis.</p

Toward a Traceless Tag for the Thiol‐Mediated Uptake of Proteins

The emergence of thiol‐mediated uptake (TMU) as a powerful strategy to penetrate cells calls for the development of practical small‐molecule TMU tools for traceless delivery. Toward this goal, esters are explored here as bioreversible linkers between dynamic covalent cascade exchangers accounting for TMU and the protein of interest (POI). The method relies on α ‐aryl‐ α ‐diazoamides that react with carboxylic acids of the POI to form esters that can be enzymatically hydrolyzed inside cells to release the native POI. A two‐step protocol is established for bioreversible conjugation of TMU tags to the POI. Despite the small number of tags attached to POIs to prevent isoelectric precipitation, POIs with traceless TMU tags are shown to efficiently enter cells not only in 2D culture but also in 3D spheroids mimicking deep tissue, confirming a key advantage of TMU. Uptake inhibition by various thiol‐reactive agents confirms the participation of cell‐surface thiols in cell penetration, i. e., the occurrence of TMU.</p

hSirT1-Dependent Regulation of the PCAF-E2F1-p73 Apoptotic Pathway in Response to DNA Damage▿

The NAD+-dependent histone deacetylase hSirT1 regulates cell survival and stress responses by inhibiting p53-, NF-κB-, and E2F1-dependent transcription. Here we show that the hSirT1/PCAF interaction controls the E2F1/p73 apoptotic pathway. hSirT1 represses E2F1-dependent P1p73 promoter activity in untreated cells and inhibits its activation in response to DNA damage. hSirT1, PCAF, and E2F1 are corecruited in vivo on theP1p73 promoter. hSirT1 deacetylates PCAF in vitro and modulates PCAF acetylation in vivo. In cells exposed to apoptotic DNA damage, nuclear NAD+ levels decrease and inactivate hSirT1 without altering the hSirT1 interaction with PCAF and hSirT1 binding to the P1p73 promoter. The reactivation of hSirT1 by pyruvate that increases the [NAD+]/[NADH] ratio completely abolished the DNA damage-induced activation of TAp73 expression, thus linking the modulation of chromatin-bound hSirT1 deacetylase activity by the intracellular redox state with P1p73 promoter activity. The release of PCAF from hSirT1 repression favors the assembly of transcriptionally active PCAF/E2F1 complexes onto the P1p73 promoter and p53-independent apoptosis. Our results identify hSirT1 and PCAF as potential targets to modulate tumor cell survival and chemoresistance irrespective of p53 status

Cyclodextrin triggers MCOLN1-dependent endo-lysosome secretion in Niemann-Pick type C cells[S]

In specialized cell types, lysosome-related organelles support regulated secretory pathways, whereas in nonspecialized cells, lysosomes can undergo fusion with the plasma membrane in response to a transient rise in cytosolic calcium. Recent evidence also indicates that lysosome secretion can be controlled transcriptionally and promote clearance in lysosome storage diseases. In addition, evidence is also accumulating that low concentrations of cyclodextrins reduce the cholesterol-storage phenotype in cells and animals with the cholesterol storage disease Niemann-Pick type C, via an unknown mechanism. Here, we report that cyclodextrin triggers the secretion of the endo/lysosomal content in nonspecialized cells and that this mechanism is responsible for the decreased cholesterol overload in Niemann-Pick type C cells. We also find that the secretion of the endo/lysosome content occurs via a mechanism dependent on the endosomal calcium channel mucolipin-1, as well as FYCO1, the AP1 adaptor, and its partner Gadkin. We conclude that endo-lysosomes in nonspecialized cells can acquire secretory functions elicited by cyclodextrin and that this pathway is responsible for the decrease in cholesterol storage in Niemann-Pick C cells

Antibody against apolipoprotein-A1, non-alcoholic fatty liver disease and cardiovascular risk: a translational study

Background: Non-alcoholic fatty liver disease (NAFLD) is a common liver disease increasing cardiovascular disease (CVD) morbidity and mortality. Autoantibodies against apolipoprotein A-1 (AAA-1) are a possible novel CVD risk factor promoting inflammation and disrupting cellular lipid homeostasis, two prominent pathogenic features of NAFLD. We explored the role of AAA-1 in NAFLD and their association with CVD risk. Methods: HepaRG cells and liver sections from ApoE-/- mice exposed to AAA-1 were used for lipid quantification and conditional protein expression. Randomly selected sera from 312 subjects of the Prevention of Renal and Vascular End-stage Disease (PREVEND) general population cohort were used to measure AAA-1. A Fatty Liver Index (FLI) ≥ 60 and a 10-year Framingham Risk Score (FRS) ≥ 20% were used as proxy of NAFLD and high CVD risk, respectively. Results: In-vitro and mouse models showed that AAA-1 increased triglyceride synthesis leading to steatosis, and promoted inflammation and hepatocyte injury. In the 112 PREVEND participants with FLI ≥ 60, AAA-1 were associated with higher FRS, alkaline phosphatase levels, lower HDL cholesterol and tended to display higher FLI values. Univariate linear and logistic regression analyses (LRA) confirmed significant associations between AAA-1, FLI and FRS ≥ 20%, while in adjusted LRA, FLI was the sole independent predictor of FRS ≥ 20% (OR: 1.05, 95%CI 1.01-1.09, P = 0.003). AAA-1 was not an independent FLI predictor. Conclusions: AAA-1 induce a NAFLD-compatible phenotype in vitro and in mice. Intricate associations exist between AAA-1, CVD risk and FLI in the general population. Further work is required to refine the role of AAA-1 in NAFLD and to determine if the AAA-1 association with CVD is affected by hepatic steatosis.</p

Toward a Traceless Tag for the Thiol‐Mediated Uptake of Proteins

Abstract The emergence of thiol‐mediated uptake (TMU) as a powerful strategy to penetrate cells calls for the development of practical small‐molecule TMU tools for traceless delivery. Toward this goal, esters are explored here as bioreversible linkers between dynamic covalent cascade exchangers accounting for TMU and the protein of interest (POI). The method relies on α‐aryl‐α‐diazoamides that react with carboxylic acids of the POI to form esters that can be enzymatically hydrolyzed inside cells to release the native POI. A two‐step protocol is established for bioreversible conjugation of TMU tags to the POI. Despite the small number of tags attached to POIs to prevent isoelectric precipitation, POIs with traceless TMU tags are shown to efficiently enter cells not only in 2D culture but also in 3D spheroids mimicking deep tissue, confirming a key advantage of TMU. Uptake inhibition by various thiol‐reactive agents confirms the participation of cell‐surface thiols in cell penetration, i. e., the occurrence of TMU