Myeloid translocation gene-16 co-repressor promotes degradation of hypoxia-inducible factor 1.

The myeloid translocation gene 16 (MTG16) co-repressor down regulates expression of multiple glycolytic genes, which are targets of the hypoxia-inducible factor 1 (HIF1) heterodimer transcription factor that is composed of oxygen-regulated labile HIF1α and stable HIF1β subunits. For this reason, we investigated whether MTG16 might regulate HIF1 negatively contributing to inhibition of glycolysis and stimulation of mitochondrial respiration. A doxycycline Tet-On system was used to control levels of MTG16 in B-lymphoblastic Raji cells.

The transcriptional co-repressor myeloid translocation gene 16 inhibits glycolysis and stimulates mitochondrial respiration.

The myeloid translocation gene 16 product MTG16 is found in multiple transcription factor-containing complexes as a regulator of gene expression implicated in development and tumorigenesis. A stable Tet-On system for doxycycline-dependent expression of MTG16 was established in B-lymphoblastoid Raji cells to unravel its molecular functions in transformed cells. A noticeable finding was that expression of certain genes involved in tumor cell metabolism including 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 and 4 (PFKFB3 and PFKFB4), and pyruvate dehydrogenase kinase isoenzyme 1 (PDK1) was rapidly diminished when MTG16 was expressed.

The leukemia associated nuclear corepressor ETO homologue genes MTG16 and MTGR1 are regulated differently in hematopoietic cells.

Background: MTG16, MTGR1 and ETO are nuclear transcriptional corepressors of the human ETO protein family. MTG16 is implicated in hematopoietic development and in controlling erythropoiesis/megakaryopoiesis. Furthermore, ETO homologue genes are 3'participants in leukemia fusions generated by chromosomal translocations responsible of hematopoietic dysregulation.

Neutrophil elastase and proteinase 3 trafficking routes in myelomonocytic cells.

Neutrophil elastase (NE) and proteinase 3 (PR3) differ in intracellular localization, which may reflect different trafficking mechanisms of the precursor forms when synthesized at immature stages of neutrophils. To shed further light on these mechanisms, we compared the trafficking of precursor NE (proNE) and precursor PR3 (proPR3). Like proNE [1], proPR3 interacted with CD63 upon heterologous co-expression in COS cells but endogenous interaction was not detected although cell surface proNE/proPR3/CD63 were co-endocytosed in myelomonocytic cells.

The leukemia associated ETO nuclear repressor gene is regulated by the GATA-1 transcription factor in erythroid/megakaryocytic cells.

Background: The Eight-Twenty-One (ETO) nuclear co-repressor gene belongs to the ETO homologue family also containing Myeloid Translocation Gene on chromosome 16 (MTG16) and myeloid translocation Gene-Related protein 1 (MTGR1). By chromosomal translocations ETO and MTG16 become parts of fusion proteins characteristic of morphological variants of acute myeloid leukemia. Normal functions of ETO homologues have as yet not been examined.

Secretory lysosome targeting and induced secretion of human soluble TNF-alpha receptor in murine hematopoietic cells in vivo as a principle for immunoregulation in inflammation and malignancy.

Objective: Systemic administration of immunotherapeutics often gives rise to severe side effects. A local deposition, using secretory lysosomes of hematopoietic cells as vehicles for delivery, can overcome this problem. In the present study, the validity of this concept was investigated using retroviral transduction of the human soluble tumor necrosis factor-alpha receptor 1 (hsTNFR1) into murine bone marrow cells, followed by transfer of the genetically modified cells into irradiated mice.

The tetraspanin CD63 is involved in granule targeting of neutrophil elastase.

Targeting mechanisms of neutrophil elastase (NE) and other luminal proteins stored in myeloperoxidase (MPO)-positive secretory lysosomes/primary granules of neutrophils are unknown. These granules contain an integral membrane protein, CD63, with an adaptor protein-3-dependent granule delivery system. Therefore, we hypothesized that CD63 cooperates in granule delivery of the precursor of NE (proNE).

The human SIN3B corepressor forms a nucleolar complex with leukemia-associated ETO homologues.

Background: SIN3 (SWI-Independent) is part of a transcriptional deacetylase complex, which generally mediates the formation of repressive chromatin. The purpose of this work was to study possible interactions between corepressors human SIN3B (hSIN3B) and the ETO homologues - ETO (eight twenty-one), MTG16 (myeloid-transforming gene 16) and MTGR1 (MTG-related protein 1). In addition, the subnuclear localization of the hSIN3B and the ETO homologues was also examined.

Neutrophil elastase sorting involves plasma membrane trafficking requiring the C-terminal propeptide.

The primary granules/secretory lysosomes of neutrophils store mature neutrophil elastase (NE) as a luminal protein after proteolytic removal of N-terminal and C-terminal pro-peptides from a proform of NE. The N-terminal pro-peptide prevents premature activation that might be toxic to the cell, but the C-terminal pro-peptide has no defined function. In this study, we investigated the role of the C-terminal pro-peptide in trafficking of NE by expressing, in rat basophilic leukemia (RBL) cells, both wild-type NE and the mutant NE/Delta248-267, which lacks the C-terminal pro-peptide.

Granule targeting of soluble tumor necrosis factor (TNF) receptor expressed during granulopoietic maturation in murine bone marrow cells.

In this experiment, we explored the potential of secretory lysosomes of hematopoietic cells to act as vehicles for immunomodulatory protein delivery at an inflammation site. We investigated whether exogenous soluble TNF-receptor 1 (sTNFR1) could be expressed in primary hematopoietic progenitor cells and become targeted for storage and secretion during granulopoietic differentiation. An sTNFR1 construct with a transmembrane domain (tm) and a cytosol sorting signal (Y) taken from CD63, was retrovirally transduced to lineage-negative murine hematopoietic bone marrow stem/progenitor cells.

Biosynthesis, processing, and sorting of human myeloperoxidase.

Exclusively synthesized by normal neutrophil and monocyte precursor cells, myeloperoxidase (MPO) functions not only in host defense by mediating efficient microbial killing but also can contribute to progressive tissue damage in chronic inflammatory states such as atherosclerosis. The biosynthetic precursor, apoproMPO, is processed slowly in the ER, undergoing cotranslational N-glycosylation, transient interactions with the molecular chaperones calreticulin and calnexin, and heme incorporation to generate enzymatically active proMPO that is competent for export into the Golgi. After exiting the Golgi the propeptide is removed prior to final proteolytic processing in azurophil granules, resulting in formation of a symmetric MPO homodimer linked by a disulfide bond.

Reactivation of latent Epstein Barr virus infection induces remission of splenic lymphoma with villous lymphocytes.

Splenic lymphoma with villous lymphocytes (SLVL) is a rare, slowly progressing, lymphoproliferative disease of B-cell origin. Treatment aimed at cure or remission is limited, and spontaneous remission has not been described. We report on a patient who after 4 years with untreated SLVL developed symptoms of mononucleosis and was found to have reactivation of latent Epstein-Barr virus (EBV) infection.

The Leukemia-associated ETO homologues are differently expressed during hematopoietic differentiation.

The Eight twenty-one (ETO) homologues are nuclear repressor proteins including ETO, myeloid-transforming gene-related protein 1 (MTGR1), and myeloid-transforming gene chromosome 16 (MTG16). ETO and MTG16 are both part of fusion proteins resulting from chromosomal translocations associated with acute myeloid leukemia. Expression of these chimeras results in a differentiation block that contributes to the onset of leukemia.

Hematopoietic secretory granules as vehicles for the local delivery of cytokines and soluble cytokine receptors at sites of inflammation.

Cytokines play an important role in the regulation of homeostasis and inflammation. A de-regulated cytokine function can subsequently promote chronic inflammation. This is supported by clinical evidence showing the beneficial effect of inhibiting TNF-alpha through injection of antibodies and soluble receptor in disorders such as rheumatoid arthritis and Crohn's disease.

Biosynthesis and sorting of myeloperoxidase in hematopoietic cells.

The neutrophil granulocytes have a critical role in innate immunity through killing of phagocytized microorganisms, in which myeloperoxidase (MPO) participates. MPO is stored in cytoplasmic azurophil lysosome-like granules together with other antibiotic proteins and digestive enzymes. During passage in the secretory pathway pro-MPO is folded, subjected to oligosaccharide modification, and retrieval from constitutive secretion to become targeted to azurophil granules for final processing and storage.

Sorting soluble tumor necrosis factor (TNF) receptor for storage and regulated secretion in hematopoietic cells.

Hematopoietic cells contain secretory lysosomes that degranulate at sites of inflammation. We envisage that secretory granules can act as vehicles for targeting inflammatory sites, including malignancies, and thereafter, locally release therapeutically active agents to these sites. Exogenous proteins, such as the soluble tumor necrosis factor receptor 1 (sTNFR1), have been shown previously to be targeted to secretory lysosomes [1].

Sorting of Von Willebrand factor to lysosome-related granules of haematopoietic cells.

The aim of this work was to investigate sorting mechanisms of von Willebrand factor (VWF) when expressed in haematopoietic cells. The processing and sorting of both the wild-type VWF and a multimerization defective propeptide-mutant (VWF(m)) were investigated after expression in the 32D cell line. Normal proteolytic processing was observed for both proteins, however the processing of VWF(m) was much slower and a large portion was unprocessed.

Targeting proteins to secretory lysosomes of natural killer cells as a principle for immunoregulation.

Secretory lysosomes of natural killer (NK) cells combine storage, regulated secretion and lysosomal activity. We asked whether one could target exogenous proteins to the secretory lysosomes of NK-cells for final delivery into a tumor site upon degranulation. cDNAs for both soluble and transmembrane (tm) proteins were expressed in the human YT-Indy NK-cell line.

Artificially controlled aggregation of proteins and targeting in hematopoietic cells.

The targeting mechanisms for granule proteins in hematopoietic cells are largely unknown. Aggregation is believed to be important for protein sorting-for-entry and sorting-by-retention in endocrine and neuroendocrine cells. We asked whether artificially induced multimerization/aggregation of chimeric proteins could affect their sorting in hematopoietic cells.

Sorting of soluble TNF-receptor for granule storage in hematopoietic cells as a principle for targeting of selected proteins to inflamed sites.

Hematopoietic cells have secretory lysosomes that degranulate at the inflammatory site upon stimulation. We asked whether one could target exogenous proteins with a therapeutic potential to secretory lysosomes in hematopoietic cells. For this purpose, we expressed a soluble tumor necrosis factor (TNF) receptor form (sTNFR1) in hematopoietic cell lines.

Sorting of neutrophil-specific granule protein human cathelicidin, hCAP-18, when constitutively expressed in myeloid cells.

Neutrophil granulocytes carry storage organelles, e.g., azurophil and specific granules.