scholarly journals The synthesis of ninety proteins including actin throughout the HeLa cell cycle.

1978 ◽  
Vol 79 (3) ◽  
pp. 833-838 ◽  
Author(s):  
C Milcarek ◽  
K Zahn
Keyword(s):  
Cell Cycle ◽  
Total Synthesis ◽  
Hela Cell ◽  
Gel Electrophoresis ◽  
Similar Proportion ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Time Points ◽  
Cytoplasmic Proteins ◽  
Actin Protein

Abundant cytoplasmic proteins pulse-labeled with [35S]methionine at specific times throughout the HeLa cell cycle were analyzed with two-dimensional gel electrophoresis. More than 300 proteins could be resolved in this way. The frequency of appearance of label in the most abundant 90 proteins, ranging from 4% to less than 0.1% of the total methionine incorporated, was determined at six time points in the cell cycle. 84 of these proteins were made as a similar proportion of the total at all times during the cell cycle. A nonmuscle actin protein (spot 1) identified by molecular weight and isoelectric point represented 2-4% of the total methionine incorporated at all the time points. Only six proteins were found which varied by greater than fourfold during cell division, four appearing to represent a greater proportion of the total synthesis during the period at or immediately surrounding M (spots 31b, 44, 53, and 70d). Two appear to represent a smaller percentage of total synthesis during the early (spot 78) or the total (spot 74) G2 period.

Characterization ofChlamydia trachomatis L2-induced tyrosine-phosphorylated HeLa cell proteins by two-dimensional gel electrophoresis

1997 ◽  
Vol 18 (3-4) ◽  
pp. 563-567 ◽  
Author(s):  
Svend Birkelund ◽  
Luca Bini ◽  
Vitaliano Pallini ◽  
Maria Sanchez-Campillo ◽  
Sabrina Liberatori ◽  
...  
Keyword(s):  
Hela Cell ◽  
Gel Electrophoresis ◽  
Two Dimensional ◽  
Ofchlamydia Trachomatis ◽  
Two Dimensional Gel Electrophoresis

scholarly journals A search for differential polypeptide synthesis throughout the cell cycle of HeLa cells.

1980 ◽  
Vol 84 (3) ◽  
pp. 795-802 ◽  
Author(s):  
R Bravo ◽  
J E Celis
Keyword(s):  
Cell Cycle ◽  
Hela Cells ◽  
Gel Electrophoresis ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Nonhistone Proteins ◽  
Polypeptide Synthesis ◽  
Acidic Proteins ◽  
Filament Protein

The polypeptides synthesized during the cell cycle of HeLa cells were analyzed by means of two-dimensional gel electrophoresis followed by fluorography under conditions in which the position of 700 polypeptides (acidic and basic) could be reproducibly assessed. Mitotic cells obtained by mechanical detachment and synchronized cells in other stages of the cell cycle were labeled with [35S]methionine for 30-min pulses or for long terms starting at the beginning of each phase. Visual comparison of the polypeptide maps obtained in the different stages of the cell cycle showed that these were strikingly similar, and there was no indication that the synthesis of any of the detected polypeptides was confined to only one of the cell cycle phases. Quantitation of 99 abundant polypeptides (acidic and basic) in pulse-labeled and long-term labeled cells revealed that the relative amount (i.e., the rate of synthesis) of most polypeptides, including total actin, alpha-actinin, 6 abundant basic nonhistone proteins, and 13 major acidic proteins present in Triton cytoskeletons, remains constant throughout the cell cycle. Among the few variable polypeptides (markers), we have identified alpha- and beta-tubulin (increase in M), the subunit of the 100-A filament protein "fibroblast type" (decreases in M), and a 36,000 mol wt acidic cytoarchitectural protein that increases in S. A few other unidentified polypeptides have also been found to vary in M and in M and G2, but no marker was found in G1.

Synthesis of stress-induced protein in isolated and perfused rat hearts

1986 ◽  
Vol 64 (5) ◽  
pp. 418-426 ◽  
Author(s):  
R. William Currie
Keyword(s):  
Protein Synthesis ◽  
Heat Shock ◽  
Total Synthesis ◽  
Gel Electrophoresis ◽  
Liquid Scintillation ◽  
Scintillation Counting ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Rat Hearts ◽  
Perfused Rat Hearts

Isolated and perfused rat hearts were examined by two-dimensional gel electrophoresis and liquid scintillation counting for alterations in protein synthesis following incubation with L-[3H]leucine at 0.5–2.5, 2.5–4.5, or 4.5–6.5 h of perfusion. When 35-mL volumes of three different buffers were recycled for a 2-h period from 0.5 to 2.5 h, by fluorography little effect was seen on the normal patterns of protein synthesis and there was a moderate synthesis of a stress-induced protein (heat-shock protein) with a molecular mass of 71 × 103 daltons (SP71). However, hearts perfused with Krebs-improved Ringer 1 bicarbonate had the highest incorporation of L-[3H]leucine. When buffers were recycled for 30-min periods from 0.5 to 2.5 h, SP71 was synthesized in hearts perfused with Krebs–Henseleit original Ringer bicarbonate. Hearts perfused in a similar fashion with Krebs-improved Ringer 1 bicarbonate had the lowest incorporation of label into SP71 and in fact SP71 was undetectable on fluorograms. Overall protein synthesis was decreased and the ratio of SP71 to the total synthesis was increased at 4.5–6.5 h of perfusion when 35-mL volumes of Krebs-improved Ringer 1 bicarbonate was recycled for 2-h periods. A similar result was observed at 2.5–4.5 h of perfusion when this buffer was recycled for either the duration of the experiment or 30-min periods.

scholarly journals Biogenesis of phagolysosomes proceeds through a sequential series of interactions with the endocytic apparatus

1994 ◽  
Vol 124 (5) ◽  
pp. 677-688 ◽  
Author(s):  
M Desjardins ◽  
LA Huber ◽  
RG Parton ◽  
G Griffiths
Keyword(s):  
Gel Electrophoresis ◽  
Sucrose Gradient ◽  
Small Gtpases ◽  
Endocytic Pathway ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Time Points ◽  
Multiple Contacts ◽  
Biochemical Assays ◽  
Latex Beads

We have examined the modifications occurring during the transformation of phagosomes into phagolysosomes in J-774 macrophages. The use of low density latex beads as markers of phagosomes (latex bead compartments, LBC) allowed the isolation of these organelles by flotation on a simple sucrose gradient. Two-dimensional gel electrophoresis, immunocytochemistry, and biochemical assays have been used to characterize the composition of LBC at different time points after their formation, as well as their interactions with the organelles of the endocytic pathway. Our results show that LBC acquire and lose various markers during their transformation into phagolysosomes. Among these are members of the rab family of small GTPases as well as proteins of the lamp family. The transfer of the LBC of lamp 2, a membrane protein associated with late endocytic structures, was shown to be microtubule dependent. Video-microscopy showed that newly formed phagosomes were involved in rapid multiple contacts with late components of the endocytic pathway. Collectively, these observations suggest that phagolysosome formation is a highly dynamic process that involves the gradual and regulated acquisition of markers from endocytic organelles.

Up-dated catalogue of HeLa cell proteins: percentages and characteristics of the major cell polypeptides labeled with a mixture of 16 14C-labeled amino acids.

1982 ◽  
Vol 28 (4) ◽  
pp. 766-781 ◽  
Author(s):  
R Bravo ◽  
J E Celis
Keyword(s):  
Cell Cycle ◽  
Amino Acids ◽  
High Resolution ◽  
Gel Electrophoresis ◽  
Neoplastic Transformation ◽  
Cellular Distribution ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Gradient Electrophoresis ◽  
Protein Map

Abstract A total of 1357 polypeptides [946 acidic (isoelectric focusing) and 411 basic (nonequilibrium pH-gradient electrophoresis)] from human HeLa cells have been separated and catalogued with use of high-resolution two-dimensional gel electrophoresis. Of these polypeptides, 1266 were detected by labeling cells with [35S]methionine, while the rest were revealed by silver staining or by labeling with a mixture of 16 14C-labeled amino acids. For convenience, all these polypeptides have been numbered and are indicated in a large fold-out protein map. The percentages of some of the major 14C-labeled proteins have been determined, and for some we list a few characteristics such as: variation during the cell cycle; cellular distribution in cytoplasts and karyoplasts; presence in Triton- and salt-extracted cytoskeletons; and phosphorylation and sensitivity to neoplastic transformation.

Effect of asynchronous transfer on bovine embryonic development and relationship with early cycle uterine proteome profiles

2012 ◽  
Vol 24 (7) ◽  
pp. 962 ◽  
Author(s):  
A. M. Ledgard ◽  
M. C. Berg ◽  
W. H. McMillan ◽  
G. Smolenski ◽  
A. J. Peterson
Keyword(s):  
Gel Electrophoresis ◽  
Purine Nucleoside Phosphorylase ◽  
Critical Time ◽  
Protein Composition ◽  
Matrix Metalloproteinase 2 ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Time Points ◽  
Phosphoserine Aminotransferase

The uterus provides the nurturing environment that supports the growth of the early preimplantation bovine conceptus. To determine critical time points of uterine influence, in vitro-produced Day 7 blastocysts were transferred into synchronous (Day 7) uteri and asynchronous uteri (Days 5 or 9). Embryo growth was evaluated 7 and 15 days after transfer and compared with that of embryos generated by AI. Conceptuses recovered from asynchronous Day 9 transfers were fourfold larger than synchronous transfer or gestational Day 14 AI conceptuses; by 15 days after transfer, differences were less marked. Two-dimensional gel electrophoresis was used to compare the histotroph protein composition of uterine luminal flushings (ULF) on Days 5 and 9 after oestrous to determine any protein differences that would promote embryo growth. The ULF were collected by serially flushing the uteri of the same heifers and mature cows at different times of the cycle. Ten proteins that differed in abundance between Day 5 and 9 were identified by mass spectrometry. Three, namely phosphoserine aminotransferase 1, purine nucleoside phosphorylase and aldose reductase, were verified by western blot analysis as more abundant on Day 9 (P < 0.002). Myostatin was present in only in Day 9 ULF, whereas tissue inhibitor of matrix metalloproteinase 2 (TIMP2) and legumain were only detected in Day 14 ULF. Although mature cows had lower progesterone concentrations on Days 5 and 14 (P < 0.05) and tended to have less TIMP2 than heifer groups, no other protein differences were detected. Thus, the embryo growth-enhancing environment on Day 9 was associated with temporal changes in the expression of several proteins of the histotroph.

Mapping and identification of interferon gammaregulated HeLa cell proteins separated by immobilized pH gradient two-dimensional gel electrophoresis

2007 ◽  
pp. 404-413
Author(s):  
Allan Christian Shaw ◽  
Martin Rssel Larsen ◽  
Peter Roepstorff ◽  
Just Justesen ◽  
Gunna Christiansen ◽  
...  
Keyword(s):  
Hela Cell ◽  
Gel Electrophoresis ◽  
Ph Gradient ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis
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