Characterization of the Yeast Transcriptome

Velculescu, Victor E.; Zhang, Lin; Zhou, Wei; Vogelstein, Jacob V.; Basrai, Munira A.; Bassett, Douglas E.; Hieter, Phil; Vogelstein, Bert; Kinzler, Kenneth W.

doi:10.1016/s0092-8674(00)81845-0

Cited by 979 publications

(804 citation statements)

References 31 publications

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“…Indeed, recent SAGE analysis of *60 000 transcripts in the yeast Saccharomyces cerevisiae resulted in the identi®cation of nearly all of the anticipated 6000 yeast genes (Velculescu et al, 1997). Estimates for expressed genes range from 10 000 to 50 000 unique mRNAs in a given cell type (Bains, 1996) Ginsberg et al, 1991;Michalovitz et al, 1990) were maintained in DMEM containing 10% fetal bovine serum in 5% CO 2 at either 328C or 388C.…”

Section: Phe132mentioning

confidence: 99%

SAGE transcript profiles for p53-dependent growth regulation

et al. 1997

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Serial analysis of gene expression (SAGE) allows for a quantitative, representative, and comprehensive pro®le of gene expression. We have utilized SAGE technology to contrast the di erential gene expression pro®le in rat embryo ®broblast cells producing temperature-sensitive p53 tumor suppressor protein at permissive or nonpermissive temperatures. Analysis of *15 000 genes revealed that the expression of 14 genes (P50.001, 50.03% abundance) was dependent on functional p53 protein, whereas the expression of three genes was signi®cantly higher in cells producing non-functional p53 protein. Those genes whose expression was increased by functional p53 include RAS, U6 snRNA, cyclin G, EGR-1, and several novel genes. The expression of actin, tubulin, and HSP70 genes was elevated at the nonpermissive temperature for p53 function. Interestingly, the expression of several genes was dependent on a nontemperature-sensitive mutant p53 suggesting altered transcription pro®les dependent on speci®c p53 mutant proteins. These results demonstrate the utility of SAGE for rapidly and reproducibly evaluating global transcriptional responses within di erent cell populations.

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Section: Phe132mentioning

confidence: 99%

SAGE transcript profiles for p53-dependent growth regulation

et al. 1997

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“…A new approach to studying cell function is to gain a comprehensive view of all expressed gene products and their interactions in a given cell [13]. The first method used to perform global expression analyses in mammals was SAGE (serial analysis of gene expression, [14,15]). …”

Section: Introductionmentioning

confidence: 99%

“…The major advantages of SAGE in comparison to microarray analyses are that absolute values (tag counts) are obtained which can be easily compared with other SAGE libraries, and that SAGE yields very extensive transcriptome information whereas microarray studies are restricted by the number of DNAs spotted onto the arrays. The SAGE method has been used extensively to characterize 3 differences in gene expression profiles of cancer versus normal cells [14,17], and has proven to be an appropriate tool for the analysis of complete transcriptomes [15,18,19]. Currently, no global gene expression data for cardiac fibroblasts is available.…”

Section: Introductionmentioning

confidence: 99%

Quantitative analysis of the cardiac fibroblast transcriptome—implications for NO/cGMP signaling

et al. 2004

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SummaryCardiac fibroblasts regulate tissue repair and remodeling in the heart. To quantify transcript levels in these cells we performed a comprehensive gene expression study using serial analysis of gene expression (SAGE). Among 110,169 sequenced tags we could identify 30,507 unique transcripts. A comparison of SAGE data from cardiac fibroblasts with data derived from total mouse heart revealed a number of fibroblast-specific genes. Cardiac fibroblasts expressed a specific collection of collagens, matrix proteins and metalloproteinases, growth factors, and components of signalling pathways. The NO/cGMP signalling pathway was represented by the mRNAs for α 1 and β 1 subunits of guanylyl cyclase, cGMP-dependent protein kinase type I (cGK I) and interestingly the G-kinase anchoring protein GKAP42. The expression of cGK I was verified by RT-PCR and Western Blot. To establish a functional role for cGK I in cardiac fibroblasts we studied its effect on cell proliferation. Selective activation of cGK I with a cGMP-analog inhibited the proliferation of serum-stimulated cardiac fibroblasts which express cGK I, but not higher passage fibroblasts which contain no detectable cGK I. Currently, our data suggest that cGK I mediates the inhibitory effects of the NO/cGMP pathway on cardiac fibroblast growth.Furthermore the SAGE library of transcripts expressed in cardiac fibroblasts provides a basis for future investigations into the pathological regulatory mechanisms underlying cardiac fibrosis.

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“…Today the field of proteomics covers essentially all of the above described efforts. To understand complex cellular dynamics, powerful methods for quantification of the messenger ribonucleic acid (mRNA) synthesis have been developed [1][2][3]. Although these methods are very sensitive, mRNA expression levels do not necessarily reflect changes in the concentration of specific functional proteins in cells [4].…”

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confidence: 99%

Trypsin catalyzed ¹⁶O-to-¹⁸O exchange for comparative proteomics: Tandem mass spectrometry comparison using MALDI-TOF, ESI-QTOF, and ESI-ion trap mass spectrometers

Heller¹,

Mattou²,

Menzel³

et al. 2003

J. Am. Soc. Mass Spectrom.

144

186

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Quantitative or comparative proteome analysis was initially performed with 2-dimensional gel electrophoresis with the inherent disadvantages of being biased towards certain proteins and being labor intensive. Alternative mass spectrometry-based approaches in conjunction with gel-free protein/peptide separation have been developed in recent years using various stable isotope labeling techniques. Common to all these techniques is the incorporation, biosynthetically or chemically, of a labeling moiety having either a natural isotope distribution of hydrogen, carbon, oxygen, or nitrogen (light form) or being enriched with heavy isotopes like deuterium, 13 C, 18 O, or 15 N, respectively. By mixing equal amounts of a control sample possessing for instance the light form of the label with a heavy-labeled case sample, differentially labeled peptides are detected by mass spectrometric methods and their intensities serve as a means for direct relative protein quantification. While each of the different labeling methods has its advantages and disadvantages, the endoprotease 16 O-to-18 O catalyzed oxygen exchange at the C-terminal carboxylic acid is extremely promising because of the specificity assured by the enzymatic reaction and the labeling of essentially every proteasederived peptide. We show here that this methodology is applicable to complex biological samples such as a subfraction of human plasma. Furthermore, despite the relatively small mass difference of 4 Da between the two labeled forms, corresponding to the exchange of two oxygen atoms by two 18 O isotopes, it is possible to quantify differentially labeled proteins on an ion trap mass spectrometer with a mass resolution of about 2000 in automated data dependent LC-MS/MS acquisition mode. Post column sample deposition on a MALDI target parallel to on-line ESI-MS/MS enables the analysis of the same compounds by means of ESIand MALDI-MS/MS. This has the potential to increase the confidence in the quantification results as well as to increase the sequence coverage of potentially interesting proteins by complementary peptide ionization techniques. Additionally the paired y-ion signals in tandem mass spectra of 16 O/ 18 O-labeled peptide pairs provide a means to confirm automatic protein identification results or even to assist de novo sequencing of yet unknown proteins. (J Am Soc Mass Spectrom 2003, 14, 704 -718)

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Characterization of the Yeast Transcriptome

Cited by 979 publications

References 31 publications

SAGE transcript profiles for p53-dependent growth regulation

SAGE transcript profiles for p53-dependent growth regulation

Quantitative analysis of the cardiac fibroblast transcriptome—implications for NO/cGMP signaling

Trypsin catalyzed ¹⁶O-to-¹⁸O exchange for comparative proteomics: Tandem mass spectrometry comparison using MALDI-TOF, ESI-QTOF, and ESI-ion trap mass spectrometers

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Characterization of the Yeast Transcriptome

Cited by 979 publications

References 31 publications

SAGE transcript profiles for p53-dependent growth regulation

SAGE transcript profiles for p53-dependent growth regulation

Quantitative analysis of the cardiac fibroblast transcriptome—implications for NO/cGMP signaling

Trypsin catalyzed 16O-to-18O exchange for comparative proteomics: Tandem mass spectrometry comparison using MALDI-TOF, ESI-QTOF, and ESI-ion trap mass spectrometers

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Trypsin catalyzed ¹⁶O-to-¹⁸O exchange for comparative proteomics: Tandem mass spectrometry comparison using MALDI-TOF, ESI-QTOF, and ESI-ion trap mass spectrometers