Plasma Level of Pyrophosphate Is Low in Pseudoxanthoma Elasticum Owing to Mutations in the ABCC6 Gene, but It Does Not Correlate with ABCC6 Genotype.

Background: Pseudoxanthoma elasticum (PXE), a monogenic disorder resulting in calcification affecting the skin, eyes and peripheral arteries, is caused by mutations in the ABCC6 gene, and is associated with low plasma inorganic pyrophosphate (PP). It is unknown how ABCC6 genotype affects plasma PP.

Methods: We studied the association of ABCC6 genotype (192 patients with biallelic pathogenic ABCC6 mutations) and PP levels, and its association with the severity of arterial and ophthalmological phenotypes.

Cloning and characterization of a novel functional organic anion transporting polypeptide 3A1 isoform highly expressed in the human brain and testis.

Organic anion transporting polypeptide 3A1 (OATP3A1, encoded by the gene) is a prostaglandin, oligopeptide, and steroid/thyroid hormone transporter with wide tissue distribution, expressed, e.g., in the human brain and testis.

Oral supplementation of inorganic pyrophosphate in pseudoxanthoma elasticum.

Pseudoxanthoma elasticum (PXE; OMIM 264800) is a rare heritable multisystem disorder, characterized by ectopic mineralization affecting elastic fibres in the skin, eyes and the cardiovascular system. Skin findings often lead to early diagnosis of PXE, but currently, no specific treatment exists to counteract the progression of symptoms. PXE belongs to a group of Mendelian calcification disorders linked to pyrophosphate metabolism, which also includes generalized arterial calcification of infancy (GACI) and arterial calcification due to CD73 deficiency (ACDC).

Creation of the first monoclonal antibody recognizing an extracellular epitope of hABCC6.

Mutations in the ABCC6 gene result in calcification diseases such as pseudoxanthoma elasticum or Generalized Arterial Calcification of Infancy. Generation of antibodies recognizing an extracellular (EC) epitope of ABCC6 has been hampered by the short EC segments of the protein. To overcome this limitation, we immunized bovine FcRn transgenic mice exhibiting an augmented humoral immune response with Human Embryonic Kidney 293 cells cells expressing human ABCC6 (hABCC6).

Pyrophosphate therapy prevents trauma-induced calcification in the mouse model of neurogenic heterotopic ossification.

Trauma-induced calcification is the pathological consequence of complex injuries which often affect the central nervous system and other parts of the body simultaneously. We demonstrated by an animal model recapitulating the calcification of the above condition that adrenaline transmits the stress signal of brain injury to the calcifying tissues. We have also found that although the level of plasma pyrophosphate, the endogenous inhibitor of calcification, was normal in calcifying animals, it could not counteract the acute calcification.

Dietary Pyrophosphate Modulates Calcification in a Mouse Model of Pseudoxanthoma Elasticum: Implication for Treatment of Patients.

Pseudoxanthoma elasticum is a heritable disease caused by ABCC6 deficiency. Patients develop ectopic calcification in skin, eyes, and vascular tissues. ABCC6, primarily found in liver and kidneys, mediates the cellular efflux of ATP, which is rapidly converted into inorganic pyrophosphate (PPi), a potent inhibitor of calcification.

Oral administration of pyrophosphate inhibits connective tissue calcification.

Various disorders including pseudoxanthoma elasticum (PXE) and generalized arterial calcification of infancy (GACI), which are caused by inactivating mutations in and , respectively, present with extensive tissue calcification due to reduced plasma pyrophosphate (PPi). However, it has always been assumed that the bioavailability of orally administered PPi is negligible. Here, we demonstrate increased PPi concentration in the circulation of humans after oral PPi administration.

Accumulation of the PX domain mutant Frank-ter Haar syndrome protein Tks4 in aggresomes.

Background: Cells deploy quality control mechanisms to remove damaged or misfolded proteins. Recently, we have reported that a mutation (R43W) in the Frank-ter Haar syndrome protein Tks4 resulted in aberrant intracellular localization.

Results: Here we demonstrate that the accumulation of Tks4(R43W) depends on the intact microtubule network.

Tubulin acetylation promoting potency and absorption efficacy of deacetylase inhibitors.

Background And Purpose: Histone deacetylase 6 (HDAC6) and silent information regulator 2 (SIRT2) control the dynamics of the microtubule network via their deacetylase activities. Tubulin polymerization promoting protein (TPPP/p25) enhances microtubule acetylation by its direct binding to HDAC6. Our objective was to characterize the multiple interactions of the deacetylases and to establish the inhibitory potency and the pharmacokinetic features of the deacetylase inhibitors, trichostatin A (TSA) and AGK2.

Identification of motives mediating alternative functions of the neomorphic moonlighting TPPP/p25.

The disordered Tubulin Polymerization Promoting Protein (TPPP/p25), a prototype of neomorphic moonlighting proteins, displays physiological and pathological functions by interacting with distinct partners. Here the role of the disordered N- and C-termini straddling a middle flexible segment in the distinct functions of TPPP/p25 was established, and the binding motives responsible for its heteroassociations with tubulin and α-synuclein, its physiological and pathological interacting partner, respectively, were identified. We showed that the truncation of the disordered termini altered the folding state of the middle segment and has functional consequences concerning its physiological function.

Moonlighting microtubule-associated proteins: regulatory functions by day and pathological functions at night.

The sensing, integrating, and coordinating features of the eukaryotic cells are achieved by the complex ultrastructural arrays and multifarious functions of the cytoskeletal network. Cytoskeleton comprises fibrous protein networks of microtubules, actin, and intermediate filaments. These filamentous polymer structures are highly dynamic and undergo constant and rapid reorganization during cellular processes.

Interactions of pathological hallmark proteins: tubulin polymerization promoting protein/p25, beta-amyloid, and alpha-synuclein.

The disordered tubulin polymerization promoting protein (TPPP/p25) was found to be co-enriched in neuronal and glial inclusions with α-synuclein in Parkinson disease and multiple system atrophy, respectively; however, co-occurrence of α-synuclein with β-amyloid (Aβ) in human brain inclusions has been recently reported, suggesting the existence of mixed type pathologies that could result in obstacles in the correct diagnosis and treatment. Here we identified TPPP/p25 as an interacting partner of the soluble Aβ oligomers as major risk factors for Alzheimer disease using ProtoArray human protein microarray. The interactions of oligomeric Aβ with proteins involved in the etiology of neurological disorders were characterized by ELISA, surface plasmon resonance, pelleting experiments, and tubulin polymerization assay.

Synthesis and in vitro antitumor effect of vinblastine derivative-oligoarginine conjugates.

Vinblastine is a widely used anticancer drug with undesired side effects. Its conjugation with carrier molecules could be an efficient strategy to reduce these side effects. Besides this, the conjugate could exhibit increased efficiency against resistant cells, e.

TPPP/p25 promotes tubulin acetylation by inhibiting histone deacetylase 6.

TPPP/p25 (tubulin polymerization-promoting protein/p25) is an unstructured protein that induces microtubule polymerization in vitro and is aligned along the microtubule network in transfected mammalian cells. In normal human brain, TPPP/p25 is expressed predominantly in oligodendrocytes, where its expression is proved to be crucial for their differentiation process. Here we demonstrated that the expression of TPPP/p25 in HeLa cells, in doxycycline-inducible CHO10 cells, and in the oligodendrocyte CG-4 cells promoted the acetylation of alpha-tubulin at residue Lys-40, whereas its down-regulation by specific small interfering RNA in CG-4 cells or by the withdrawal of doxycycline from CHO10 cells decreased the acetylation level of alpha-tubulin.

Tubulin polymerization-promoting protein (TPPP/p25) is critical for oligodendrocyte differentiation.

TPPP/p25, a recently identified tubulin polymerization-promoting protein (TPPP), is expressed mainly in myelinating oligodendrocytes of the CNS. Here, we show that TPPP/p25 is strongly upregulated during the differentiation of primary oligodendrocyte cells as well as the CG-4 cell line. The microRNA expression profile of CG-4 cells before and after induction of differentiation was established and revealed differential regulation of a limited subset of microRNAs.

Unique pharmacology of KAR-2, a potential anti-cancer agent: absorption modelling and selective mitotic spindle targeting.

Bis-indols are a large group of the anti-cancer agents, which effectively arrest the uncontrolled division of the cancerous cells. Their use in clinical chemotherapy is still limited because of: (i) the non-specific targeting of the mitotic cells; (ii) low bioavailability of the drugs. KAR-2 has been identified as a tubulin binding agent which displays significantly lower cytotoxicity but favourable anti-cancer potency than its mother molecule, vinblastine.

Progress in the development of early diagnosis and a drug with unique pharmacology to improve cancer therapy.

Cancer continues to be one of the major health and socio-economic problems worldwide, despite considerable efforts to improve its early diagnosis and treatment. The identification of new constituents as biomarkers for early diagnosis of neoplastic cells and the discovery of new type of drugs with their mechanistic actions are crucial to improve cancer therapy. New drugs have entered the market, thanks to industrial and legislative efforts ensuring continuity of pharmaceutical development.

Phosphorylation blocks the activity of tubulin polymerization-promoting protein (TPPP): identification of sites targeted by different kinases.

Tubulin polymerization-promoting protein (TPPP), an unfolded brain-specific protein interacts with the tubulin/microtubule system in vitro and in vivo, and is enriched in human pathological brain inclusions. Here we show that TPPP induces tubulin self-assembly into intact frequently bundled microtubules, and that the phosphorylation of specific sites distinctly affects the function of TPPP. In vitro phosphorylation of wild type and the truncated form (Delta3-43TPPP) of human recombinant TPPP was performed by kinases involved in brain-specific processes.

Unconventional translation initiation of human trypsinogen 4 at a CUG codon with an N-terminal leucine. A possible means to regulate gene expression.

Chromosomal rearrangements apparently account for the presence of a primate-specific gene (protease serine 3) in chromosome 9. This gene encodes, as the result of alternative splicing, both mesotrypsinogen and trypsinogen 4. Whereas mesotrypsinogen is known to be a pancreatic protease, neither the chemical nature nor biological function of trypsinogen 4 has been explored previously.

Tubulin polymerization promoting proteins (TPPPs): members of a new family with distinct structures and functions.

TPPP/p25 is a brain-specific protein, which induces tubulin polymerization and microtubule (MT) bundling and is enriched in Lewy bodies characteristic of Parkinson's disease [Tirián et al. (2003) Proc. Natl.

Interaction of TPPP/p25 protein with glyceraldehyde-3-phosphate dehydrogenase and their co-localization in Lewy bodies.

TPPP/p25, a flexible unstructured protein, binds to tubulin and induces aberrant microtubule assemblies. We identified hereby glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as a new interacting partner of TPPP/p25. The immunoprecipitation and affinity chromatographic experiments with bovine brain cell-free extract revealed that the interaction was salt and NAD(+) sensitive while ELISA showed resistant and firm association of the two isolated proteins.

Dynamic targeting of microtubules by TPPP/p25 affects cell survival.

Recently we identified TPPP/p25 (tubulin polymerization promoting protein/p25) as a brain-specific unstructured protein that induced aberrant microtubule assemblies and ultrastructure in vitro and as a new marker for Parkinson's disease and other synucleopathies. In this paper the structural and functional consequences of TPPP/p25 are characterized to elucidate the relationship between the in vitro and the pathological phenomena. We show that at low expression levels EGFP-TPPP/p25 specifically colocalizes with the microtubule network of HeLa and NRK cells.