bims-tunefa Biomed News
on Tumor necrosis factor superfamily and post-translational modifications
Issue of 2020‒08‒23
sixteen papers selected by
John Silke
Walter and Eliza Hall Institute of Medical Research


  1. Viruses. 2020 Aug 17. pii: E901. [Epub ahead of print]12(8):
    Morrish E, Mackiewicz L, Silke N, Pellegrini M, Silke J, Brumatti G, Ebert G.
      Chronic hepatitis B virus (HBV) infection remains a global health threat and affects hundreds of millions worldwide. Small molecule compounds that mimic natural antagonists of inhibitor of apoptosis (IAP) proteins, known as Smac-mimetics (second mitochondria-derived activator of caspases-mimetics), can promote the death of HBV-replicating liver cells and promote clearance of infection in preclinical models of HBV infection. The Smac-mimetic birinapant is a substrate of the multidrug resistance protein 1 (MDR1) efflux pump, and therefore inhibitors of MDR1 increase intracellular concentration of birinapant in MDR1 expressing cells. Liver cells are known to express MDR1 and other drug pump proteins. In this study, we investigated whether combining the clinical drugs, birinapant and the MDR1 inhibitor zosuquidar, increases the efficacy of birinapant in killing HBV expressing liver cells. We showed that this combination treatment is well tolerated and, compared to birinapant single agent, was more efficient at inducing death of HBV-positive liver cells and improving HBV-DNA and HBV surface antigen (HBsAg) control kinetics in an immunocompetent mouse model of HBV infection. Thus, this study identifies a novel and safe combinatorial treatment strategy to potentiate substantial reduction of HBV replication using an IAP antagonist.
    Keywords:  HBV; IAP; IAP antagonist; MDR1 inhibitor; Smac-mimetics; TNF; cell death; combination therapy; inhibitor of apoptosis proteins
    DOI:  https://doi.org/10.3390/v12080901
  2. Cell Death Dis. 2020 Aug 13. 11(8): 680
    Miles MA, Hawkins CJ.
      Most anticancer drugs provoke apoptotic signaling by damaging DNA or other means. Genotoxic therapies may enhance a patient's risk of developing "therapy-related cancers" due to the accumulation of oncogenic mutations that may occur in noncancerous cells. Mutations can also form upon apoptotic signaling due to sublethal caspase activity, implying that apoptosis activating drugs may also be oncogenic. Necroptosis is a different way of killing cancer cells: this version of caspase-independent cell death is characterized by receptor-interacting protein kinase-3 (RIPK3) and mixed lineage kinase-like domain protein (MLKL) activation, leading to cell membrane rupture and controlled cell lysis. The mutagenic potential of sublethal necroptotic signaling has not yet been directly investigated. Smac mimetics drugs, which activate apoptotic or necroptotic cell death, do not induce mutations but the mechanistic basis for this lack of mutagenic activity has not been determined. In this study, we compared the mutagenic potential of these two cell death pathways by engineering cells to activate either apoptotic or necroptotic signaling by exposing them to Smac mimetics with or without TNFα, and/or enforcing or preventing expression of apoptotic or necroptotic regulators. We discovered that sublethal concentrations of Smac mimetics in contexts that activated apoptotic signaling provoked DNA damage and mutations in surviving cells. Mutagenesis was dependent on executioner caspase activation of the nuclease CAD. In contrast, RIPK3- and MLKL-dependent necroptotic signaling following Smac mimetic treatment was not mutagenic. Likewise, DNA damage was not provoked in cells expressing a lethal constitutively active MLKL mutant. These data reveal that cells surviving sublethal necroptotic signaling do not sustain genomic damage and provide hope for a reduced risk of therapy-related malignancies in patients treated with necroptosis-inducing drugs.
    DOI:  https://doi.org/10.1038/s41419-020-02879-y
  3. Mol Cell. 2020 Aug 17. pii: S1097-2765(20)30545-1. [Epub ahead of print]
    Huang D, Camacho CV, Setlem R, Ryu KW, Parameswaran B, Gupta RK, Kraus WL.
      Although ADP-ribosylation of histones by PARP-1 has been linked to genotoxic stress responses, its role in physiological processes and gene expression has remained elusive. We found that NAD+-dependent ADP-ribosylation of histone H2B-Glu35 by small nucleolar RNA (snoRNA)-activated PARP-1 inhibits AMP kinase-mediated phosphorylation of adjacent H2B-Ser36, which is required for the proadipogenic gene expression program. The activity of PARP-1 on H2B requires NMNAT-1, a nuclear NAD+ synthase, which directs PARP-1 catalytic activity to Glu and Asp residues. ADP-ribosylation of Glu35 and the subsequent reduction of H2B-Ser36 phosphorylation inhibits the differentiation of adipocyte precursors in cultured cells. Parp1 knockout in preadipocytes in a mouse lineage-tracing genetic model increases adipogenesis, leading to obesity. Collectively, our results demonstrate a functional interplay between H2B-Glu35 ADP-ribosylation and H2B-Ser36 phosphorylation that controls adipogenesis.
    Keywords:  ADP-ribosylation; PARP-1; adipogenesis; differentiation; histones; phosphorylation; proliferation; proteomics; snoRNA
    DOI:  https://doi.org/10.1016/j.molcel.2020.08.002
  4. Nat Struct Mol Biol. 2020 Aug 17.
    Luo EC, Nathanson JL, Tan FE, Schwartz JL, Schmok JC, Shankar A, Markmiller S, Yee BA, Sathe S, Pratt GA, Scaletta DB, Ha Y, Hill DE, Aigner S, Yeo GW.
      The molecular functions of the majority of RNA-binding proteins (RBPs) remain unclear, highlighting a major bottleneck to a full understanding of gene expression regulation. Here, we develop a plasmid resource of 690 human RBPs that we subject to luciferase-based 3'-untranslated-region tethered function assays to pinpoint RBPs that regulate RNA stability or translation. Enhanced UV-cross-linking and immunoprecipitation of these RBPs identifies thousands of endogenous mRNA targets that respond to changes in RBP level, recapitulating effects observed in tethered function assays. Among these RBPs, the ubiquitin-associated protein 2-like (UBAP2L) protein interacts with RNA via its RGG domain and cross-links to mRNA and rRNA. Fusion of UBAP2L to RNA-targeting CRISPR-Cas9 demonstrates programmable translational enhancement. Polysome profiling indicates that UBAP2L promotes translation of target mRNAs, particularly global regulators of translation. Our tethering survey allows rapid assignment of the molecular activity of proteins, such as UBAP2L, to specific steps of mRNA metabolism.
    DOI:  https://doi.org/10.1038/s41594-020-0477-6
  5. FASEB J. 2020 Aug 20.
    Olivares-González L, Velasco S, Millán JM, Rodrigo R.
      Retinitis pigmentosa (RP) is a group of inherited retinal dystrophies characterized by the progressive and irreversible loss of vision. We previously found that intraperitoneal administration of Adalimumab, a monoclonal anti-TNFα antibody, slowed down retinal degeneration in the murine model of RP, the rd10 mice. The aims of this study were to improve its neuroprotective effect and to deepen understanding of the molecular mechanisms involved in this effect. We analyzed (i) the in vitro effect of Adalimumab on the TNFα-mediated cell death in retinal cells; (ii) the effect of a single intravitreal injection of Adalimumab on retinal degeneration in rd10 mice at postnatal day (P) 23. In vitro studies showed that TNFα induced caspase and poly ADP ribose polymerase (PARP) activation, downregulation of (kinase receptor-interacting protein 1) RIPK1 and upregulation of RIPK3 in retinal cells. Adalimumab reduced cell death probably through the inhibition of caspase 3 activation. In vivo studies suggested that PARP and NLRP3 inflammasome are mainly activated and to a lesser extent caspase-dependent mechanisms in rd10 retinas at P23. Necroptosis seems to be inhibited by the downregulation of RIPK1. Adalimumab prevented from retinal degeneration without affecting caspase -dependent mechanisms but decreasing PARP activation, microglia activation as well as NLRP3 inflammasome.
    Keywords:  NLRP3 inflammasome; PARP; TNFα; photoreceptor degeneration
    DOI:  https://doi.org/10.1096/fj.202000044RR
  6. Cell Chem Biol. 2020 Aug 14. pii: S2451-9456(20)30298-1. [Epub ahead of print]
    Chakrabarty S, Verhelst SHL.
      Caspases control regulated cell death (apoptosis), a process that is crucial in the development of multicellular organisms as well as in various diseases. In order to spatiotemporally study apoptosis, we here develop photoactivatable caspase inhibitors. These are based on cysteine-reactive acyloxymethyl ketone electrophiles connected to a peptide targeting caspases. Importantly, the aspartate crucial for recognition by caspases is caged with a photoprotecting group. Ester photocages were found to be labile, and it was critical to have a nitroindoline cage, which forms a stable amide bond with the aspartate side chain. The nitroindoline-protected inhibitors lead to an efficient turn-on of inhibitory activity after irradiation with light. They are applicable in live cells, where they prevent anti-FAS-induced apoptosis only upon irradiation. Overall, these reagents will allow a better understanding of the spatial and temporal dimensions of apoptosis in complex, dynamic systems.
    Keywords:  apoptosis; caspase inhibitors; caspases; photocages; photopharmacology; photoprotecting groups; protease inhibitors
    DOI:  https://doi.org/10.1016/j.chembiol.2020.08.001
  7. Cell Death Differ. 2020 Aug 18.
    Lim Y, De Bellis D, Sandow JJ, Capalbo L, D'Avino PP, Murphy JM, Webb AI, Dorstyn L, Kumar S.
      Mitotic catastrophe (MC) is an important oncosuppressive mechanism that serves to eliminate cells that become polyploid or aneuploid due to aberrant mitosis. Previous studies have demonstrated that the activation and catalytic function of caspase-2 are key steps in MC to trigger apoptosis and/or cell cycle arrest of mitotically defective cells. However, the molecular mechanisms that regulate caspase-2 activation and its function are unclear. Here, we identify six new phosphorylation sites in caspase-2 and show that a key mitotic kinase, Aurora B kinase (AURKB), phosphorylates caspase-2 at the highly conserved residue S384. We demonstrate that phosphorylation at S384 blocks caspase-2 catalytic activity and apoptosis function in response to mitotic insults, without affecting caspase-2 dimerisation. Moreover, molecular modelling suggests that phosphorylation at S384 may affect substrate binding by caspase-2. We propose that caspase-2 S384 phosphorylation by AURKB is a key mechanism that controls caspase-2 activation during mitosis.
    DOI:  https://doi.org/10.1038/s41418-020-00604-y
  8. Mol Pharm. 2020 Aug 17.
    Corti A, Gasparri AM, Sacchi A, Colombo B, Monieri M, Rrapaj E, Ferreri AJM, Curnis F.
      The therapeutic index of cytokines in cancer therapy can be increased by targeting strategies based on protein engineering with peptides containing the CNGRC (NGR) motif, a ligand that recognize CD13-positive tumor vessels. We show here that the targeting domain of recombinant CNGRC-cytokine fusion proteins, such as NGR-TNF (a CNGRC-tumor necrosis factor-α (TNF) conjugate used in clinical studies) and NGR-EMAP-II, undergo various post-translational modification and degradation reactions that lead to the formation of markedly heterogeneous products. These modifications include N-terminal cysteine acetylation or formation of various asparagine degradation products, the latter owing to intramolecular interactions of the cysteine α-amino group with asparagine and/or its succinimide derivative. Blocking the cysteine α-amino group with a serine (SCNGRC) reduced both post-translational and degradation reactions. Furthermore, the serine residue reduced asparagine deamidation rate to isoaspartate (another degradation product) and improved the affinity of NGR for CD13. Accordingly, genetic engineering of NGR-TNF with the N-terminal serine produced a more stable and homogeneous drug (called S-NGR-TNF) with improved anti-tumor activity in tumor-bearing mice, either when used alone or in combination with chemotherapy. In conclusion, the targeting domain of NGR-cytokine conjugates can undergo various untoward modification and degradation reactions, which can be markedly reduced by fusing a serine to the N-terminus. The SCNGRC peptide may represent a ligand for cytokine delivery to tumors more robust than conventional CNGRC. The S-NGR-TNF conjugate (more stable, homogeneous and active than NGR-TNF) could be rapidly developed for clinical trial.
    DOI:  https://doi.org/10.1021/acs.molpharmaceut.0c00579
  9. Immunity. 2020 Aug 18. pii: S1074-7613(20)30330-7. [Epub ahead of print]53(2): 243-245
    Chapman NM, Gottschalk S, Chi H.
      Chimeric antigen receptor (CAR) T cells are potent drivers of antitumor immunity, but promoting durable CAR T cell responses remains challenging. In this issue of Immunity, Li et al. (2020) show that blockade of CAR ubiquitination induces CAR recycling to the cell surface, leading to increased CAR T cell cytotoxicity and longevity by amplifying 41BB-dependent signaling and mitochondrial metabolism.
    DOI:  https://doi.org/10.1016/j.immuni.2020.07.023
  10. Open Biol. 2020 Aug;10(8): 200130
    Su TT.
      This Open Question article highlights current advances in the study of non-apoptotic roles of apoptotic proteins. Apoptosis is a highly regulated and energy-requiring process in which cells actively kill themselves. Apoptosis helps remove extra cells to sculpt organs during embryo development and culls damaged cells throughout the body. Apoptosis relies on evolutionarily conserved proteins that include a family of proteases called caspases. Caspases activity has long been considered a hallmark of apoptosis. Yet an emerging body of literature indicates that caspase activity is required for a number of non-lethal processes that range from sculpting cells, removing protein aggregates, changing cell identity during differentiation or de-differentiation, and rebuilding tissues. Failure in each of these processes is associated with human disease. This article is not meant to be an exhaustive review but an introduction to the subject for an educated public, with caspases as a gateway example. I propose that it is time to explore non-apoptotic roles of caspases and other apoptotic proteins, in order to better understand their non-apoptosis function and to leverage new knowledge into new therapies.
    Keywords:  apoptosis; caspase; plasticity
    DOI:  https://doi.org/10.1098/rsob.200130
  11. J Biol Chem. 2020 Aug 14. pii: jbc.RA120.013873. [Epub ahead of print]
    Boehm AN, Bialas J, Catone N, Sacristán-Reviriego A, van der Spuy J, Groettrup M, Aichem A.
      The retina-specific chaperone AIPL1 is essential for the correct assembly of phosphodiesterase 6 (PDE6), which is a pivotal effector enzyme for phototransduction and vision because it hydrolyzes cGMP. AIPL1 interacts with the cytokine-inducible ubiquitin-like modifier FAT10 that gets covalently conjugated to hundreds of proteins and targets its conjugation substrates for proteasomal degradation, but whether FAT10 affects PDE6 function or turnover is unknown. Here, we show that FAT10 mRNA is expressed in human retina and identify rod PDE6 as a retina-specific substrate of FAT10 conjugation. We found that AIPL1 stabilizes the FAT10 monomer as well as the PDE6-FAT10 conjugate. Additionally, we elucidated the functional consequences of PDE6 FAT10ylation. On the one hand, we demonstrate that FAT10 targets PDE6 for proteasomal degradation by formation of a covalent isopeptide linkage. On the other hand, FAT10 inhibits PDE6 cGMP hydrolyzing activity by non-covalently interacting with the PDE6 GAFa and catalytic domains. Therefore, FAT10 may contribute to loss of PDE6 and, as a consequence, degeneration of retinal cells in eye diseases linked to inflammation and inherited blindness causing mutations in AIPL1.
    Keywords:  AIPL1; FAT10; PDE6; cell culture; photoreceptor; proteasome; protein processing; protein stability; retinal degeneration; ubiquitylation (ubiquitination)
    DOI:  https://doi.org/10.1074/jbc.RA120.013873
  12. Cancer Discov. 2020 Aug 14.
      Debio 1143 plus chemoradiotherapy improved head and neck squamous cell carcinoma disease control.
    DOI:  https://doi.org/10.1158/2159-8290.CD-RW2020-116
  13. Immunity. 2020 Aug 18. pii: S1074-7613(20)30323-X. [Epub ahead of print]53(2): 235-237
    Yang TB, Kim BS.
      In this issue of Immunity, Xu et al. reveal that dermal dendritic cells produce interleukin-31, which acts on neurons to promote wound itch. Their findings link itch associated with deeper wounds-wounds that extend beyond the epithelium-to the cells and cytokines that mediate wound healing.
    DOI:  https://doi.org/10.1016/j.immuni.2020.07.016
  14. Cell Death Differ. 2020 Aug 17.
    Hattori T, Takahashi Y, Chen L, Tang Z, Wills CA, Liang X, Wang HG.
      Autophagosomal membranes can serve as activation platforms for intracellular death-inducing signaling complexes (iDISCs) to initiate Caspase-8-dependent apoptosis. In this study, we explore the impact of ESCRT-III-dependent phagophore closure on iDISC assemblies and cell death in osteosarcoma and neuroblastoma cells. Inhibition of phagophore closure by conditional depletion of CHMP2A, an ESCRT-III component, stabilizes iDISCs on immature autophagosomal membranes and induces Caspase-8-dependent cell death. Importantly, suppression of the iDISC formation via deletion of ATG7, an E1 enzyme for ubiquitin-like autophagy-related proteins, blocks Caspase-8 activation and cell death following CHMP2A depletion. Although DR5 expression and TRAIL-induced apoptosis are enhanced in CHMP2A-depleted cells, the canonical extrinsic pathway of apoptosis is not responsible for the initiation of cell death by CHMP2A depletion. Furthermore, the loss of CHMP2A impairs neuroblastoma tumor growth associated with decreased autophagy and increased apoptosis in vivo. Together, these findings indicate that inhibition of the ESCRT-III-dependent autophagosome sealing process triggers noncanonical Caspase-8 activation and apoptosis, which may open new avenues for therapeutic targeting of autophagy in cancer.
    DOI:  https://doi.org/10.1038/s41418-020-00610-0
  15. Cancer Discov. 2020 Aug 14.
      Lysosome-targeting chimeras (LYTAC) directed extracellular and membrane proteins to lysosomes.
    DOI:  https://doi.org/10.1158/2159-8290.CD-RW2020-119
  16. Trends Endocrinol Metab. 2020 Aug 14. pii: S1043-2760(20)30139-9. [Epub ahead of print]
    Wang X, Wang Y, Antony V, Sun H, Liang G.
      Metabolic diseases pose a tremendous health threat in both developed and developing countries. The pathophysiology of metabolic diseases is complex but has been shown to be closely associated with sterile inflammation, which is initiated by various danger molecules derived from metabolic overload, such as oxidized low-density lipoproteins (OxLDLs), free fatty acids (FFAs), glucose, advanced glycation end products (AGEs), and cholesterol. These danger signals are sensed by pattern recognition receptors (PRRs) to activate proinflammatory signaling pathways and promote the release of proinflammatory mediators, leading to chronic low-grade inflammation. Although these harmful metabolic stimuli are generally regarded as damage-associated molecular patterns (DAMPs), a more specific definition and accurate classification for these DAMPs is still missing. In this opinion, we classify the harmful metabolic stimuli that can incite inflammatory responses and tissue damage via instigating PRRs as metabolism-associated molecular patterns (MAMPs), and we summarize their roles in metaflammation-mediated metabolic diseases.
    Keywords:  innate immunity; metabolic disease; metabolism-associated molecular patterns; metaflammation; pattern recognition receptors
    DOI:  https://doi.org/10.1016/j.tem.2020.07.001