bluuebunny/crossref_metadata_2025 huggingface dataset:
Retention of the Matricellular Protein SPARC in the Endoplasmic Reticulum of Chondrocytes from Patients with Pseudoachondroplasia
Authors: Hecht Jacqueline T. , Sage E. Helene
Published: November 2005

Pseudoachondroplasia (PSACH) is an autosomal dominant disease characterized by dwarfism, morphological irregularities of long bones and hips, and early-onset osteoarthritis. This disease has been attributed to mutations in a structural protein of the cartilage extracellular matrix (ECM), cartilage oligomeric matrix protein (COMP), which result in its selective retention in the chondrocyte rough endoplasmic reticulum (ER). Accumulation of excessive amounts of mutated COMP might reflect a defect in protein trafficking by PSACH chondrocytes. Here we identify the matricellular protein SPARC as a component of this trafficking deficit. SPARC was localized to the hypertrophic chondrocytes in the normal human tibial growth plate and in cultured control cartilage nodules. In contrast, concentrated intracellular depots of SPARC were identified in nodules cultured from three PSACH patients with mutations in COMP. The accumulated SPARC was coincident with COMP and with protein disulfide isomerase, a resident chaperone of the rough ER, whereas SPARC and COMP were not coincident in the ECM of control or PSACH nodules. SPARC-null mice develop severe osteopenia and degenerative intervertebral disc disease, and exhibit attenuation of collagenous ECM. The retention of SPARC in the ER of chondrocytes producing mutant COMP indicates a new intracellular function for SPARC in the trafficking/secretion of cartilage ECM.
Chiral bias of amyloid fibrils revealed by the twisted conformation of Thioflavin T: An induced circular dichroism/DFT study
Authors: Dzwolak Wojciech , Pecul Magdalena
Published: November 2005

Since it was implicated in a number of neurodegenerative conditions, such as Alzheimer disease, formation of β‐sheet‐rich protein fibrils (amyloids) has been drawing a lot of attention. One of elusive aspects of amyloidogenesis concerns the mechanisms of specific binding of molecules such as Congo red, or Thioflavin T by amyloid fibrils. A comprehensive understanding of these docking interactions is needed, however, for the sake of furthering biochemical studies and developing molecular, pharmacological strategies preventing proliferation of amyloids in vivo. Through the application of circular dichroism, here we show that upon binding to insulin fibrils, a twisted conformation is enforced in molecules of Thioflavin T, manifested in a strong negative Cotton effect around 450 nm, which is supported by density functional theory–based calculations. This finding may lead to circular dichroism of Thioflavin T becoming a new diagnostic technique for protein fibrils, complementary to fluorescence spectroscopy.
Malonyl‐CoA decarboxylase is present in the cytosolic, mitochondrial and peroxisomal compartments of rat hepatocytes
Authors: Joly Erik , Bendayan Moise , Roduit Raphaël , Saha Asish K. , Ruderman Neil B. , Prentki Marc
Published: November 2005

A role for cytosolic malonyl‐CoA decarboxylase (MCD) as a regulator of fatty acid oxidation has been postulated. However, there is no direct evidence that MCD is present in the cytosol. To address this issue, we performed cell fractionation and electron microscopic colloidal gold studies of rat liver to determine the location and activity of MCD. By both methods, substantial amounts of MCD protein and activity were found in the cytosol, mitochondria and peroxisomes, the latter with the highest specific activity. MCD species with different electrophoretic mobility were observed in the three fractions. The data demonstrate that active MCD is present in the cytosol, mitochondria and peroxisomes of rat liver, consistent with the view that MCD participates in the regulation of cytosolic malonyl‐CoA levels and of hepatic fatty acid oxidation.
Escherichia coli verotoxin 1 mediates apoptosis in human HCT116 colon cancer cells by inducing overexpression of the GADD family of genes and S phase arrest
Authors: Bhattacharjee Rabindra N. , Park Kwon-Sam , Uematsu Satoshi , Okada Kazuhisa , Hoshino Katsuaki , Takeda Kiyoshi , Takeuchi Osamu , Akira Shizuo , Iida Tetsuya , Honda Takeshi
Published: November 2005

The Escherichia coli verotoxin 1 (VT1) inhibits protein synthesis, cell proliferation, and damages endothelial cell in the hemolytic uremic syndrome. VT1 can specifically bind and act on endothelial cells as well as on many tumor cells because these cells express its high affinity receptor, globotriaosylceramide. This indicates that VT1 may have both antiangiogenic and antineoplastic activities. We investigated this potential of VT1 by incubating several colon cancer cell lines with VT1 for different time periods and found that HCT116 cells were especially sensitive to VT1. A combination of morphological studies, flow cytometry, DNA laddering and annexin V staining confirmed that VT1 irreversibly arrests these cells in S phase within 24 h and prolonged incubation triggers DNA fragmentation. Concomitant to the activation of the S phase checkpoint, increased levels of mRNA and proteins of growth arrest and DNA damage‐inducible gene family that include GADD34, GADD45α, and GADD45β was observed. Interestingly, no significant changes in expression of key cell cycle related proteins such as cdk2, cdk4, p21, p27, and p53 was found during the S phase arrest and apoptosis. We therefore suggest that GADD proteins might play an important role in VT1 induced S phase arrest and programmed cell death in HCT116 cells.
Phosphorylation of Thr695 and Thr850 on the myosin phosphatase target subunit: Inhibitory effects and occurrence in A7r5 cells
Authors: Murányi Andrea , Derkach Dmitry , Erdődi Ferenc , Kiss Andrea , Ito Masaaki , Hartshorne David J.
Published: November 2005

Major sites for Rho‐kinase on the myosin phosphatase target subunit (MYPT1) are Thr695 and Thr850. Phosphorylation of Thr695 inhibits phosphatase activity but the role of phosphorylation at Thr850 is not clear and is evaluated here. Phosphorylation of both Thr695 and Thr850 by Rho‐kinase inhibited activity of the type 1 phosphatase catalytic subunit. Rates of phosphorylation of the two sites were similar and efficacy of inhibition following phosphorylation was equivalent for each site. Phosphorylation of each site on MYPT1 was detected in A7r5 cells, but Thr850 was preferred by Rho‐kinase and Thr695 was phosphorylated by an unidentified kinase(s).
Combining nucleoside analogues to achieve recognition of oligopurine tracts by triplex‐forming oligonucleotides at physiological pH
Authors: Rusling David A. , Le Strat Loic , Powers Vicki E.C. , Broughton-Head Victoria J. , Booth James , Lack Oliver , Brown Tom , Fox Keith R.
Published: November 2005

We have used DNase I footprinting to examine DNA triple helix formation at a 12 base pair oligopurine · oligopyrimidine sequence, using oligonucleotides that contain combinations of 2′‐aminoethoxy‐5‐(3‐aminoprop‐1‐ynyl)uridine (bis‐amino‐U, BAU) and 3‐methyl‐2‐aminopyridine (MeP) in place of T and C, respectively. This combination acts cooperatively to enable high affinity triple helix formation at physiological pH. The affinity depends on the number of substitutions and their arrangement; oligonucleotides in which these analogues are evenly distributed throughout the third strand bind much better than those in which they are clustered together.
Characterization of centrosomal association of nucleophosmin/B23 linked to Crm1 activity
Authors: Shinmura Kazuya , Tarapore Pheruza , Tokuyama Yukari , George Kyle R. , Fukasawa Kenji
Published: November 2005

Nucleophosmin (NPM)/B23 is a multifunctional protein, involving in a wide variety of basic cellular processes, including ribosome assembly, DNA duplication, nucleocytoplasmic trafficking, and centrosome duplication. It has previously been shown that NPM/B23 localizes to centrosomes, and dissociate from centrosomes upon phosphorylation by Cdk2/cyclin E. However, detail characterization of centrosomal association of NPM/B23 has been hampered by the lack of appropriate antibodies that efficiently detects centrosomally localized NPM/B23, as well as by apparent loss of natural behavior of NPM/B23 when tagged with fluorescent proteins. Here, by the use of newly generated anti‐NPM/B23 antibody, we conducted a careful analysis of centrosomal localization of NPM/B23. We found that NPM/B23 localizes between the paired centrioles of unduplicated centrosomes, suggesting the role of NPM/B23 in the centriole pairing. Upon initiation of centrosome duplication, some NPM/B23 proteins remain at mother centrioles of the parental centriole pairs. We further found that inhibition of Crm1 nuclear export receptor results in both accumulation of cyclin E at centrosomes and efficient dissociation of NPM/B23 from centrosomes.
TAK1‐mediated transcriptional activation of CD28‐responsive element and AP‐1‐binding site within the IL‐2 promoter in Jurkat T cells
Authors: Sakurai Hiroaki , Singhirunnusorn Pattama , Shimotabira Emi , Chino Atsushi , Suzuki Shunsuke , Koizumi Keiichi , Saiki Ikuo
Published: November 2005

We focused on the functional involvement of transforming growth factor‐β‐activated kinase 1 (TAK1) in transcriptional regulation of interleukin‐2 (IL‐2) in T cells. Costimulation of Jurkat cells with 12‐O‐tetradecanoylphorbol‐13‐acetate and A23187 leads to a rapid phosphorylation of TAK1 and TAK1‐binding protein 1 (TAB1), critical for TAK1 activation. A specific inhibitor of TAK1 blocked production of IL‐2. In addition, overexpression of TAK1 and TAB1 induced secretion of IL‐2. CD28‐responsive element/activator protein‐1‐binding site (RE/AP) within the IL‐2 promoter was a functional target for TAK1. The RE/AP‐driven transcription was regulated by TAK1‐mediated activation of the c‐Jun NH2‐terminal kinase, p38 and IκB kinase. These results indicate that TAK1 plays a critical role in T cell activation by controlling production of IL‐2.
Postnatal development of cholinergic neurons in the mesopontine tegmentum revealed by histochemistry
Authors: Ninomiya Yaéko , Kayama Yukihiko , Koyama Yoshimasa
Published: November 2005

Cholinergic neurons in the laterodorsal tegmental nucleus (LDT) and pedunculopontine tegmental nucleus (PPT) play a role in the regulation of several kinds of behavior. Some of them, such as locomotion, motor inhibition or sleep, show dramatic changes at a certain period of postnatal development. To understand the neural substrate for the development of these physiological functions, we studied the development of cholinergic neurons in the LDT and PPT of postnatal and adult rats using histochemical staining of NADPH‐diaphorase (NADPH‐d) and immunohistochemical staining of choline acetyltransferase (ChAT) and the vesicular acetylcholine transporter (VAChT). At postnatal day 1 (P1), ChAT‐ and VAChT‐stained cells localized more dorsally than those of NADPH‐d‐stained cells, and at P7 their distributions became similar to those of NADPH‐d‐stained cells. The number of NADPH‐d‐stained cells increased rapidly after birth, reaching the adult level by P7. In contrast, the number of ChAT‐ and VAChT‐stained cells and the intensity of their staining decreased from P1 to P3 and then increased through P21. The volume of the LDT increased during the second postnatal week. These findings indicate that cholinergic neurons in the LDT develop their cholinergic properties during the second postnatal week and mature functionally thereafter. We discuss these results in light of the several physiological functions regulated by the cholinergic neurons in the mesopontine tegmentum.