Posts Tagged: autophagy

New Insights Into the Role of mTOR Signaling in the Cardiovascular System

New Insights Into the Role of mTOR Signaling in the Cardiovascular System

Sebastiano Sciarretta, Maurizio Forte, Giacomo Frati, Junichi Sadoshima

Schematic overview of the upstream signaling modulators of mTORC1 (mTOR [mechanistic target of rapamycin] complex 1) and mTORC2. Signaling network regulating mTORC1 and mTORC2 activity. Arrows indicate the effects of different conditions or cellular events on mTOR activity. Red arrows represent maladaptive input signals, whereas green arrows represent adaptive input signals. Akt indicates protein kinase B; AMPK, adenosine monophosphate activated protein kinase; CASTOR 1/2, cellular arginine sensor for mTORC1 1/2; ERK1/2, extracellular signal–regulated kinase 1/2; GATOR 1/2, GAP activity toward Rags 1/2; GSK-3β, glycogen synthase kinase-3β; IKKβ, inhibitor of NF-κB kinase-β; PI3K, phosphoinositide 3 kinase; PRAS40, proline-rich Akt substrate 40; Rag, Ras-related GTPase; Raptor, regulatory-associated protein of mTOR; REDD1, regulated in development and DNA damage responses 1; Rheb, Ras homolog enriched in brain; and TSC1/2, tuberous sclerosis protein 1/2. [Powerpoint File]

Macromolecular Degradation Systems and Cardiovascular Aging

Macromolecular Degradation Systems and Cardiovascular Aging

Hiroyuki Nakayama, Kazuhiko Nishida, Kinya Otsu

Fundamental pathway of ubiquitin (Ub)/proteasome system. Ub conjugation is an ATP-dependent process and is mediated by a series of enzymes. UPS consists of the Ub-activating enzyme E1 that transfers Ub to the active site of the E2 Ub-conjugating enzymes. The E3 Ub-ligases ligate Ub to an internal lysine in the target protein. The ubiquitinated protein will be targeted to the 26S proteasome for degradation or spared form degradation by the action of cytosolic deubiquitinating enzymes. The 26S proteasome consists of the 20S core complex and of 1 or 2 19S regulatory complexes that bind one or both ends of the 20S. The 20S proteasome consists of 28 subunits divided into 4 rings including 2 flanking α and 2 central β rings and contains proteolytic activities. The 3 peptidase activities of the proteasome are localized in the β1, β2, and β5 subunits of β rings. 19S complex is comprised of 20 subunits and contains ATPase activity and function as a polyubiquitin receptor. The immunoproteasome is an alternative form of the standard 20S proteasome in which the proteolytically active subunits β1, β2, and β5 are replaced by alternative subunits β1i (proteasome subunit [PSMB] 9), β2i (PSMB10), and β5i (PSMB8), respectively. The immunoproteasome is induced by interferon (IFN)-γ or tumor necrosis factor (TNF) α and plays an important role in the degradation of oxidized proteins. All 3 immunoproteasome-specific subunits are also upregulated under acute exposure to oxidative stress in cultured cells. On the contrary, 20S proteasome degrades oxidized proteins without prior targeting by ubiquitination. Oxidized proteasome subunits or aggregated proteins induced by extensive oxidation inhibit proteasomal activity that may contribute to proteasomal functional insufficiency observed in advanced aging. ROS indicates reactive oxygen species. [Powerpoint File]

Macromolecular Degradation Systems and Cardiovascular Aging

Macromolecular Degradation Systems and Cardiovascular Aging

Hiroyuki Nakayama, Kazuhiko Nishida, Kinya Otsu

Degradation of DNA and inflammation. Degradation of superfluous DNA by three DNase (DNase I, II, and III) prevents sterile inflammation. The secreted extracellular DNase I degrades inflammogenic genomic DNA released by necrotic cells, what is called a damage-associated molecular patterns, to prevent inflammasome activation. The lysosomal DNase II degrades mitochondrial-DNA during autophagic process thereby hamper toll-like receptor 9–dependent immune responses. Intrinsic self-DNA is degraded by DNase III/3′ repair exonuclease 1 in cytosol, whereby hinder stimulator of interferon gene (STING)–dependent induction of cytokine production caused by cytosolic DNA. [Powerpoint File]

How Mitochondrial Dynamism Orchestrates Mitophagy

How Mitochondrial Dynamism Orchestrates Mitophagy

Orian S. Shirihai, Moshi Song, Gerald W. Dorn II

Molecular events leading to selective mitophagy of dysfunctional mitochondria. Left, Mitochondrial depolarization or other impairment interrupts normal proteolytic processing of PTEN-induced putative kinase 1 (PINK1) kinase. PINK1 accumulates on the outer mitochondrial membrane (OMM) and phosphorylates mitofusin 2 (Mfn2), promoting its recruitment of Parkin, and ubiquitin (Ub), enabling its utilization by Parkin. Polyubiquitinated OMM proteins bind p62 linked to autophagosomal LC3, thus targeting the mitochondrion for mitophagy. [Powerpoint File]

How Mitochondrial Dynamism Orchestrates Mitophagy

How Mitochondrial Dynamism Orchestrates Mitophagy

Orian S. Shirihai, Moshi Song, Gerald W. Dorn II

Mechanisms of mitochondrial quality improvement, from macro to micro. Top, PTEN-induced putative kinase 1 (PINK1)/Parkin-mediated mitophagy of an intact organelle, as with the depolarized daughter of an asymmetrical fission event (see Figure 3). Left, Bit-by-bit autophagy of localized mitochondrial damage, which is excised and engulfed by an autophagosome via Parkin activity. Bottom, PINK1/Parkin-mediated formation of mitochondria-derived vesicle, incorporating damaged components into a vesicle that transports them to lysosomes. Right, PINK1–Parkin-independent proteasomal removal of outer mitochondrial membrane proteins, and protease-dependent degradation of internal mitochondrial proteins, the most selective of the quality improvement processes. [Powerpoint File]

Proteotoxicity and Cardiac Dysfunction

Proteotoxicity and Cardiac Dysfunction

Patrick M. McLendon, Jeffrey Robbins

Schematic diagram of selected protection strategies or proteotoxic pathways in the cardiomyocyte. Strategy 1: Rescue. The cell’s chaperones, which can be constitutively expressed or inducible, act alone or in concert to maintain or restore the peptide’s functional conformation. Strategy 2: Degradation. The misfolded protein is recognized by the protein quality control machinery, ubiquitinated, and trafficked to the proteasome for degradation and amino acid recycling. Strategy 3: Sequestration. The misfolded protein forms larger aggregates, which are unable to be processed by the proteasome. These aggregates can be attached to dynein motors and transported on microtubules to a perinuclear location where the potentially proteotoxic peptides are sequestered. These aggregates can be cleared from the cytoplasm by autophagy (see text for details).37,38,39 Strategy 4: ERAD. The endoplasmic reticulum associated degradation (ERAD) pathway is complex and a simplified schematic diagram simply outlines the process. Mutated or terminally misfolded proteins are recognzied by ER-associated proteins that may have either enzymatic or chaperone activities. The protein is then shepherded to the retrotranslocation machinery whose exact identity(ies) remain obscure. Membrane-associated E3 ligases and associated proteins are able to mediate ubiquitination of the protein, which may partially drive its retrotranslocation. There are multiple checkpoints and components underlying these processes as well (see text for details). The ubiquitinated proteins are then targeted for proteasomal degradation. [Powerpoint File]

Proteotoxicity and Cardiac Dysfunction

Proteotoxicity and Cardiac Dysfunction

Patrick M. McLendon, Jeffrey Robbins

Selective cargo recognition and degradation by autophagy. Step 1, Ubiquitin binding is necessary for most modes of selective autophagy. Polyubiquitin conjugation onto a substrate protein is accomplished via an E3 ubiquitin ligase. Many of these proteins are ubiquitinated via a K63 linkage, which signals lysosomal degradation rather than the K48 linkage that is typical for proteasomal targeting. Step 2, Ubiquitinated proteins can be recognized by cargo receptors, such as p62, NBR1, or histone deacetylase (HDAC)6 for transport towards the lysosome or aggresome. p62 (icon) binds ubiquitinated proteins for interaction with LC3 in a growing autophagosome, thus selectively targeting the protein for autophagic degradation after fusion with lysosomes. In addition, HDAC6 can bind ubiquitinated proteins through its ubiquitin-binding domain (icon) and transport them in a retrograde fashion along microtubules by binding to the motor protein dynein through the dynein motor–binding domain (icon). These can be single misfolded species or soluble proto-aggregates. This event can happen many times, causing the proteins to coalesce and form aggresomes in the perinuclear region of the cell. Aggresomal proteins can also be degraded by autophagy, and these cargo transport proteins may be involved in this process as well, particularly HDAC6 with known roles in mediating the autophagosome–lysosome fusion event. [Powerpoint File]

Molecular Mechanisms of Mitochondrial Autophagy/Mitophagy in the Heart

Molecular Mechanisms of Mitochondrial Autophagy/Mitophagy in the Heart

Toshiro Saito, Junichi Sadoshima

The mechanism of mitochondrial autophagy mediated by PTEN-inducible putative kinase 1 (Pink1)–Parkinson juvenile disease protein 2 (Parkin) in mammalian cells. Mitochondrial proteases and peptidases continuously degrade Pink1 in intact mitochondria. However, Pink1 is not imported to the inner membrane and is not cleaved in depolarized mitochondria. Pink1 then accumulates at the outer membrane and recruits Parkin. Mfn2 phosphorylated by Pink1 acts as a receptor for Parkin on mitochondria. Parkin ubiquitinates multiple proteins of the outer membrane. These ubiquitinated proteins are recognized by p62, a ubiquitin- and microtubule-associated protein 1 light chain 3 (LC3)–binding adaptor protein, followed by mitochondrial autophagy. [Powerpoint File]

Autophagy in Vascular Disease

Autophagy in Vascular Disease

Guido R.Y. De Meyer, Mandy O.J. Grootaert, Cédéric F. Michiels, Ammar Kurdi, Dorien M. Schrijvers, Wim Martinet

Role of autophagy in advanced atherosclerosis. In advanced plaques, ceroid deposition, high amounts of nitric oxide (NO) via iNOS expression, and aging impair autophagy. Failure of the autophagic process further stimulates accumulation of damaged organelles, increases reactive oxygen species generation and enhances formation of ceroid-containing lysosomes. Impaired autophagy in endothelial cells (EC) is associated with endothelial dysfunction and apoptotic cell death, possibly leading to atherothrombosis. In macrophages (MΦ), impaired autophagy results in increased sensitivity to apoptotic stimuli and impaired efferocytosis, resulting in plaque instability. Smooth muscle cells (SMC) undergo senescence, which accelerates plaque progression. [Powerpoint File]

The Role of Autophagy in Vascular Biology

The Role of Autophagy in Vascular Biology

Samuel C. Nussenzweig, Subodh Verma, Toren Finkel

Stimulation of autophagy occurs through mechanistic target of rapamycin (MTOR)–dependent and MTOR-independent pathways within vascular cells. Complex I of MTOR (mTORC1) is a well characterized, negative regulator of autophagosome formation. The classical autophagic stimulus of nutrient withdrawal is thought to be largely transduced through this pathway, whereas the pharmacological agent, rapamycin, works as an inhibitor of MTOR, thereby relieving the tonic autophagic-inhibition of MTORC1. Other agents, such as endoplasmic reticulum (ER) stress, reactive oxygen species (ROS), and oxidized lipids seem to largely work through a mix of both MTOR-dependent and MTOR-independent pathways, although the precise mechanism of action for these stimuli is largely unknown (illustration credit: Ben Smith). [Powerpoint File]

The Role of Autophagy in Vascular Biology

The Role of Autophagy in Vascular Biology

Samuel C. Nussenzweig, Subodh Verma, Toren Finkel

Endothelial disruption of autophagic flux impairs Weibel–Palade Body (WPB) formation. Graphical illustration of the normal maturation and secretion of von Willebrand factor (vWF) within endothelial cells (top). After deletion of Atg7 or Atg5 within the endothelium (bottom), there is no change in the level of vWF, but more is found in the endoplasmic reticulum (ER) and Golgi and less within the mature WPB. Moreover, perhaps, because of impaired ER and Golgi homeostasis, the pH of the WPB is altered in the absence of autophagy, leading to a shorter and more rounded morphology. Electron micrographs of these different WPB morphologies are shown (Reprinted from Torisu et al64 with permission). [Powerpoint File]

The Role of Autophagy in Vascular Biology

The Role of Autophagy in Vascular Biology

Samuel C. Nussenzweig, Subodh Verma, Toren Finkel

The convergence of autophagic flux and vascular aging. A variety of intrinsic and extrinsic factors (age, glucose, lipids, etc.) can regulate vascular autophagy. The induction of autophagy may be important in minimizing the damage that these factors can induce. With increasing age, vascular autophagic flux seems to decline and this decline may contribute to the impairment in endothelial function (with decreased nitric oxide [NO] signaling) and to the increase in vascular stiffness. Efforts to maintain and augment vascular autophagy may prove to be therapeutically beneficial. eNOS indicates endothelial nitric oxide synthase; and ox-LDL, oxidized low-density lipoprotein. [Powerpoint File]

Therapeutic Targeting of Autophagy: Potential and Concerns in Treating Cardiovascular Disease

Therapeutic Targeting of Autophagy: Potential and Concerns in Treating Cardiovascular Disease

Amabel M. Orogo, Åsa B. Gustafsson

Mechanisms of autophagy. A, Chaperone-mediated autophagy; import of proteins into lysosomes. B, Microautophagy; direct internalization of cargo into lysosomes. C, Macroautophagy; autophagosome membrane as it engulfs and sequesters cargo. Syntaxin 17 (STX17) is recruited to mature autophagosome and interacts with cytosolic synaptosomal-associated protein 29 kDa (SNAP29) and lysosomal vesicle-associated membrane protein 8 (VAMP8) to mediate autophagosome–lysosome fusion. [Powerpoint File]

Therapeutic Targeting of Autophagy: Potential and Concerns in Treating Cardiovascular Disease

Therapeutic Targeting of Autophagy: Potential and Concerns in Treating Cardiovascular Disease

Amabel M. Orogo, Åsa B. Gustafsson

Transcriptional regulation of autophagy by forkhead box O (FOXO). AKT and AMP-activated protein kinase (AMPK) exert opposing effects on FOXO activity. When growth factors are present, AKT phosphorylation of FOXO leads to FOXO translocation from the nucleus to the cytosol, where FOXO is sequestered by 14-3-3 proteins. When energy levels are low, AMPK phosphorylates and activates FOXO. sirtuin1 (SIRT1) deacetylates FOXO to increase its transcriptional activity. FOXO proteins activate transcription of several autophagy genes. [Powerpoint File]

Therapeutic Targeting of Autophagy: Potential and Concerns in Treating Cardiovascular Disease

Therapeutic Targeting of Autophagy: Potential and Concerns in Treating Cardiovascular Disease

Amabel M. Orogo, Åsa B. Gustafsson

Alternative autophagy. The autophagosome membrane is derived from the trans-Golgi. This process occurs independently of ATG5 and ATG7. These RAB9-positive vesicles subsequently fuse with the lysosome. [Powerpoint File]