The Localization of Trypsin in Cultured Mammalian Cells

1973 ◽  
Vol 12 (3) ◽  
pp. 887-902
Author(s):  
GISELE M. HODGES ◽  
D. C. LIVINGSTON ◽  
L. M. FRANKS
Keyword(s):  
Mammalian Cells ◽  
Cultured Cells ◽  
Intracellular Localization ◽  
Mouse Kidney ◽  
Kidney Cells ◽  
Intracellular Uptake ◽  
Primary Event ◽  
Temperature Dependent ◽  
Surface Localization ◽  
Cell Surface Localization

The localization of trypsin in HeLa and CBM17 baby mouse kidney cells was studied using fluorescent and electron-microscope autoradiographical techniques. Intracellular uptake of trypsin, as well as cell surface localization, was demonstrated by the use of direct FITC- and tritium acetylated-labelled trypsin and immunoreactive procedures. Intracellular penetration of the enzyme was temperature dependent and while evident at 37 and 25°C was negligible at 4°C. Higher proteolytic activity could be demonstrated in the supernatants from disrupted trypsin-treated cells than in supernatants from disrupted PBS-treated cultures. Treatment of trypsin with serum, whilst depressing enzyme protease activity, did not modify intracellular uptake of the enzyme and intracellular localization of trypsin persisted in cultured cells for up to about 48 h. The results, while not accounting for the primary event in cell alteration following exposure to trypsin, clearly suggest that consideration must be given to the fact that intracellular penetration of the enzyme may affect certain intrinsic processes of the cell.

Expression of oncomodulin does not lead to the transformation or immortalization of mammalian cells in vitro

1989 ◽  
Vol 94 (3) ◽  
pp. 517-525
Author(s):  
A.M. Mes-Masson ◽  
S. Masson ◽  
D. Banville ◽  
L. Chalifour
Keyword(s):  
Mammalian Cells ◽  
Mouse Kidney ◽  
Kidney Cells ◽  
T Antigens ◽  
Primary Mouse ◽  
Specific Antisera ◽  
Growth Properties ◽  
Metallothionein Promoter ◽  
Specific Mrna

A recombinant plasmid (pMTONCO) containing the coding sequences for rat oncomodulin under the direction of the metallothionein promoter was constructed. pMTONCO was co-transfected with the pSV2-NEO plasmid into primary mouse kidney cells or Rat-1 cells using the calcium phosphate technique and stable transformants were isolated after selection with G418. Transcription from the metallothionein promoter was inducible with heavy metals and produced an oncomodulin-specific mRNA. The presence of oncomodulin protein in stable cell lines was verified by immunoprecipitation with specific antisera. While a plasmid encoding the polyomavirus T-antigens was able to prolong the life-span of primary mouse kidney cells in culture, no equivalent activity was noted when the pMTONCO plasmid was used to transfect primary cells. When expressed in Rat-1 cells, oncomodulin did not affect the growth properties of these cells, nor did it predispose cells to higher frequencies of oncogenic transformation to a viral oncogene. We conclude that oncomodulin is neither an immortalizing nor transforming agent in vitro.

scholarly journals Native adiponectin in serum binds to mammalian cells expressing T-cadherin, but not AdipoRs or calreticulin

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Shunbun Kita ◽  
Shiro Fukuda ◽  
Norikazu Maeda ◽  
Iichiro Shimomura
Keyword(s):  
Cell Surface ◽  
Human Serum ◽  
Cho Cells ◽  
Mammalian Cells ◽  
Hek293 Cells ◽  
C2c12 Myotubes ◽  
Binding Partner ◽  
Surface Localization ◽  
Significant Attenuation ◽  
Cell Surface Localization

Adiponectin is an adipocyte-derived atypically abundant circulating factor that protects various organs and tissues through its receptors, AdipoRs, calreticulin, and T-cadherin. To identify the major binding partner of circulating native adiponectin, we expressed these receptors on the surface of HEK293 cells. Adiponectin, either that in mouse or human serum, purified from serum, or produced by mammalian cells, bound to cells expressing T-cadherin, but not to those expressing AdipoR1 or calreticulin. The stable introduction of T-cadherin and AdipoR1 into CHO cells resulted in the cell surface localization of these receptors. Native adiponectin in serum bound to cells expressing T-cadherin, not to those expressing AdipoR1. The knockdown of T-cadherin, but not AdipoRs resulted in the significant attenuation of native adiponectin binding to C2C12 myotubes. Therefore, native adiponectin binding depended on the amount of T-cadherin expressed in HEK293 cells, CHO cells, and C2C12 myotubes. Collectively, our mammalian cell-based studies suggest that T-cadherin is the major binding partner of native adiponectin in serum.

scholarly journals Intracellular Localization ofStaphylococcus aureuswithin Primary Cultured Mouse Kidney Cells

1992 ◽  
Vol 36 (5) ◽  
pp. 431-443 ◽  
Author(s):  
Miyo Murai ◽  
Akemi Usui ◽  
Keiko Seki ◽  
Junji Sakurada ◽  
Shogo Masuda
Keyword(s):  
Intracellular Localization ◽  
Mouse Kidney ◽  
Kidney Cells

Effect of sodium butyrate on induction of cellular and viral DNA syntheses in polyoma virus-infected mouse kidney cells.

1981 ◽  
Vol 38 (3) ◽  
pp. 973-981 ◽  
Author(s):  
E Wawra ◽  
E Pöckl ◽  
E Müllner ◽  
E Wintersberger
Keyword(s):  
Infected Mouse ◽  
Sodium Butyrate ◽  
Mouse Kidney ◽  
Polyoma Virus ◽  
Kidney Cells ◽  
Viral Dna

scholarly journals Itaconate Alters Succinate and Coenzyme A Metabolism via Inhibition of Mitochondrial Complex II and Methylmalonyl-CoA Mutase

Metabolites ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 117
Author(s):  
Thekla Cordes ◽  
Christian M. Metallo
Keyword(s):  
Amino Acid ◽  
Metabolic Pathways ◽  
Mammalian Cells ◽  
Coenzyme A ◽  
Cultured Cells ◽  
Tca Cycle ◽  
Regulatory Function ◽  
Reversible Inhibitor ◽  
Complex Ii ◽  
Branched Chain

Itaconate is a small molecule metabolite that is endogenously produced by cis-aconitate decarboxylase-1 (ACOD1) in mammalian cells and influences numerous cellular processes. The metabolic consequences of itaconate in cells are diverse and contribute to its regulatory function. Here, we have applied isotope tracing and mass spectrometry approaches to explore how itaconate impacts various metabolic pathways in cultured cells. Itaconate is a competitive and reversible inhibitor of Complex II/succinate dehydrogenase (SDH) that alters tricarboxylic acid (TCA) cycle metabolism leading to succinate accumulation. Upon activation with coenzyme A (CoA), itaconyl-CoA inhibits adenosylcobalamin-mediated methylmalonyl-CoA (MUT) activity and, thus, indirectly impacts branched-chain amino acid (BCAA) metabolism and fatty acid diversity. Itaconate, therefore, alters the balance of CoA species in mitochondria through its impacts on TCA, amino acid, vitamin B12, and CoA metabolism. Our results highlight the diverse metabolic pathways regulated by itaconate and provide a roadmap to link these metabolites to potential downstream biological functions.

scholarly journals Temperature-dependent secretion of Zika virus envelope and non-structural protein 1 in mammalian cells for clinical applications

2021 ◽  
pp. 114175
Author(s):  
Young Chan Kim ◽  
Joanne E. Nettleship ◽  
Nallely García-Larragoiti ◽  
Mar Maria Antonieta ◽  
Ariadna Lorena Mondragón-García ◽  
...  
Keyword(s):  
Zika Virus ◽  
Mammalian Cells ◽  
Clinical Applications ◽  
Structural Protein ◽  
Temperature Dependent ◽  
Virus Envelope

scholarly journals Replication Past the -Radiation-Induced Guanine-Thymine Cross-Link G[8,5-Me]T by Human and Yeast DNA Polymerase

2010 ◽  
Vol 2010 ◽  
pp. 1-10 ◽  
Author(s):  
Paromita Raychaudhury ◽  
Ashis K. Basu
Keyword(s):  
Dna Polymerase ◽  
Mammalian Cells ◽  
Translesion Synthesis ◽  
Kidney Cells ◽  
Equal Frequency ◽  
Cross Link ◽  
Dominant Mutation ◽  
Human Embryonic Kidney Cells ◽  
Radiation Induced

-Radiation-induced intrastrand guanine-thymine cross-link, G[8,5-Me]T, hinders replicationin vitroand is mutagenic in mammalian cells. Herein we reportin vitrotranslesion synthesis of G[8,5-Me]T by human and yeast DNA polymerase (hPol and yPol ). dAMP misincorporation opposite the cross-linked G by yPol was preferred over correct incorporation of dCMP, but further extension was 100-fold less efficient for :A compared to :C. For hPol , both incorporation and extension were more efficient with the correct nucleotides. To evaluate translesion synthesis in the presence of all four dNTPs, we have developed a plasmid-based DNA sequencing assay, which showed that yPol was more error-prone. Mutational frequencies of yPol and hPol were 36% and 14%, respectively. Targeted was the dominant mutation by both DNA polymerases. But yPol induced targeted in 23% frequency relative to 4% by hPol . For yPol , targeted and constituted 83% of the mutations. By contrast, with hPol , semi-targeted mutations (7.2%), that is, mutations at bases near the lesion, occurred at equal frequency as the targeted mutations (6.9%). The kind of mutations detected with hPol showed significant similarities with the mutational spectrum of G[8,5-Me]T in human embryonic kidney cells.

Influenza Virus: Association of Mouse-Lung Virulence with Plaque Formation in Mouse Kidney Cells

Intervirology ◽  
1976 ◽  
Vol 7 (4-5) ◽  
pp. 201-210 ◽  
Author(s):  
Bertram Flehmig ◽  
Angelika Vallbracht ◽  
Hans-Joachim Gerth
Keyword(s):  
Influenza Virus ◽  
Plaque Formation ◽  
Mouse Kidney ◽  
Mouse Lung ◽  
Kidney Cells

scholarly journals A NIMA-related Kinase, Fa2p, Localizes to a Novel Site in the Proximal Cilia of Chlamydomonas and Mouse Kidney Cells

2004 ◽  
Vol 15 (11) ◽  
pp. 5172-5186 ◽  
Author(s):  
Moe R. Mahjoub ◽  
M. Qasim Rasi ◽  
Lynne M. Quarmby
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Cell Types ◽  
Mouse Kidney ◽  
Kidney Cells ◽  
Polycystic Kidney ◽  
Cycle Progression ◽  
Cystic Kidneys ◽  
Ciliary Function ◽  
The Relationship

Polycystic kidney disease and related syndromes involve dysregulation of cell proliferation in conjunction with ciliary defects. The relationship between cilia and cell cycle is enigmatic, but it may involve regulation by the NIMA-family of kinases (Neks). We previously showed that the Nek Fa2p is important for ciliary function and cell cycle in Chlamydomonas. We now show that Fa2p localizes to an important regulatory site at the proximal end of cilia in both Chlamydomonas and a mouse kidney cell line. Fa2p also is associated with the proximal end of centrioles. Its localization is dynamic during the cell cycle, following a similar pattern in both cell types. The cell cycle function of Fa2p is kinase independent, whereas its ciliary function is kinase dependent. Mice with mutations in Nek1 or Nek8 have cystic kidneys; therefore, our discovery that a member of this phylogenetic group of Nek proteins is localized to the same sites in Chlamydomonas and kidney epithelial cells suggests that Neks play conserved roles in the coordination of cilia and cell cycle progression.

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