scholarly journals Use of yeast artificial chromosomes (YASs) for studying control of gene expression: Correct regulation of the genes of a human  -globin locus YAC following transfer to mouse erythroleukemia cell lines

1994 ◽  
Vol 91 (5) ◽  
pp. 1979-1979 ◽  
Keyword(s):  
Gene Expression ◽  
Cell Lines ◽  
Control Of Gene Expression ◽  
Artificial Chromosomes ◽  
Erythroleukemia Cell ◽  
Yeast Artificial Chromosomes ◽  
Mouse Erythroleukemia

scholarly journals Use of yeast artificial chromosomes (YACs) for studying control of gene expression: correct regulation of the genes of a human beta-globin locus YAC following transfer to mouse erythroleukemia cell lines

1993 ◽  
Vol 90 (23) ◽  
pp. 11207-11211 ◽  
Author(s):  
K R Peterson ◽  
G Zitnik ◽  
C Huxley ◽  
C H Lowrey ◽  
A Gnirke ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Lines ◽  
Yeast Artificial Chromosome ◽  
Globin Gene ◽  
Artificial Chromosome ◽  
Globin Genes ◽  
Beta Globin ◽  
Control Of Gene Expression ◽  
Artificial Chromosomes ◽  
Mouse Erythroleukemia

We demonstrate that transfer of a yeast artificial chromosome (YAC) containing 230 kb of the human beta-globin locus into mouse erythroleukemia cells by fusion results in correct developmental regulation of the human beta-like globin genes. Additionally, we show that early after hybrid formation, human embryonic epsilon- and fetal gamma-globin genes are coexpressed with the adult beta gene but that after 10-20 weeks in culture, globin gene expression switches to predominantly adult. Thus, in contrast to shorter gene constructs, the globin genes of the beta-globin locus YAC are regulated like the chromosomal globin genes. These results indicate that transfer of YACs into established cell lines can be used for the analysis of the developmental control of multigenic and developmentally regulated human loci.

scholarly journals Stable episomal maintenance of yeast artificial chromosomes in human cells.

1996 ◽  
Vol 16 (9) ◽  
pp. 5117-5126 ◽  
Author(s):  
K Simpson ◽  
A McGuigan ◽  
C Huxley
Keyword(s):  
Cell Lines ◽  
Viral Genome ◽  
Close Association ◽  
Human Cells ◽  
Yeast Cells ◽  
Control Of Gene Expression ◽  
Artificial Chromosomes ◽  
Barr Virus ◽  
Episomal Maintenance ◽  
Yeast Artificial Chromosomes

Plasmids carrying the Epstein-Barr virus origin of plasmid replication (oriP) have been shown to replicate autonomously in latently infected human cells (J. Yates, N. Warren, D. Reisman, and B. Sugden, Proc. Natl. Acad. Sci. USA 81:3806-3810, 1984). We demonstrate that addition of this domain is sufficient for stable episomal maintenance of yeast artificial chromosomes (YACs), up to at least 660 kb, in human cells expressing the viral protein EBNA-1. To better approximate the latent viral genome, YACs were circularized before addition of the oriP domain by homologous recombination in yeast cells. The resulting OriPYACs were maintained as extrachromosomal molecules over long periods in selection; a 90-kb OriPYAC was unrearranged in all cell lines analyzed, whereas the intact form of a 660-kb molecule was present in two of three cell lines. The molecules were also relatively stable in the absence of selection. This finding indicates that the oriP-EBNA-1 interaction is sufficient to stabilize episomal molecules of at least 660 kb and that such elements do not undergo rearrangements over time. Fluorescence in situ hybridization analysis demonstrated a close association of OriPYACs, some of which were visible as pairs, with host cell chromosomes, suggesting that the episomes replicate once per cell cycle and that stability is achieved by attachment to host chromosomes, as suggested for the viral genome. The wide availability of YAC libraries, the ease of manipulation of cloned sequences in yeast cells, and the episomal stability make OriPYACs ideal for studying gene function and control of gene expression.

scholarly journals Genetic control of gene expression in whole blood and lymphoblastoid cell lines is largely independent

2011 ◽  
Vol 22 (3) ◽  
pp. 456-466 ◽  
Author(s):  
J. E. Powell ◽  
A. K. Henders ◽  
A. F. McRae ◽  
M. J. Wright ◽  
N. G. Martin ◽  
...  
Keyword(s):  
Gene Expression ◽  
Genetic Control ◽  
Cell Lines ◽  
Whole Blood ◽  
Lymphoblastoid Cell Lines ◽  
Control Of Gene Expression

scholarly journals Regulatory factors specific for adult and embryonic globin genes may govern their expression in erythroleukemia cells

Blood ◽  
1985 ◽  
Vol 65 (3) ◽  
pp. 705-712 ◽  
Author(s):  
NP Anagnou ◽  
TY Yuan ◽  
E Lim ◽  
J Helder ◽  
S Wieder ◽  
...  
Keyword(s):  
Gene Expression ◽  
Human Chromosome ◽  
Globin Gene ◽  
Globin Genes ◽  
Chromosome 16 ◽  
Regulatory Factors ◽  
Erythroleukemia Cells ◽  
Erythroleukemia Cell ◽  
Globin Gene Expression ◽  
Mouse Erythroleukemia

Abstract In order to test if trans-acting regulatory factors specific for globin genes of the adult and embryonic stages of development exist in erythroid cells, transcriptionally active embryonic and adult globin genes on the same chromosome were transferred by cell fusion from the human leukemia cell K562 into phenotypically adult mouse erythroleukemia cells. Restriction-fragment-length polymorphisms of the K562 zeta (embryonic) globin genes were used to establish that all three copies of human chromosome 16 present in the K562 cell showed the same pattern of human globin gene expression after transfer to the mouse erythroleukemia cell. Adult (alpha) but not embryonic (zeta) human globin mRNA was detected in all nine of the independently derived mouse erythroleukemia hybrid cells, each of which contained human chromosome 16. Restriction endonuclease studies of the K562 alpha- and zeta-globin genes after transfer into the mouse erythroleukemia cell showed no evidence of rearrangements or deletions that could explain this loss of zeta-globin gene expression. These data suggest that regulation of globin gene expression in these erythroleukemia cells involves trans-acting regulatory factors specific for the adult and embryonic stages of development.

Cell-specific posttranscriptional regulation of CFTR gene expression via influence of MAPK cascades on 3′UTR part of transcripts

2005 ◽  
Vol 289 (5) ◽  
pp. C1240-C1250 ◽  
Author(s):  
Maryvonne Baudouin-Legros ◽  
Alexandre Hinzpeter ◽  
Amandine Jaulmes ◽  
Franck Brouillard ◽  
Bruno Costes ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Lines ◽  
Posttranscriptional Regulation ◽  
P38 Map Kinase ◽  
Cftr Gene ◽  
Control Of Gene Expression ◽  
Cytosolic Proteins ◽  
Tnf Α ◽  
Ht 29

Expression of the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) gene, which contains the mutations responsible for CF, is regulated by cytokines (TNF-α and IL-1β) in a cell-specific manner. TNF-α decreases CFTR mRNA in human colon cell lines (HT-29), but not in pulmonary cell lines (Calu-3), and IL-1β increases it only in Calu-3 cells. We looked for the cytokine-induced posttranscriptional regulation of CFTR gene expression and studied the modulation of CFTR mRNA stability linked to its 3′ untranslated sequence (3′UTR) in HT-29 and Calu-3 cells. The stability of CFTR mRNA was analyzed by Northern blot after in vitro incubation of total RNAs from CFTR-expressing cells with cytosolic proteins extracted from control or cytokine-treated HT-29 and Calu-3 cells. CFTR mRNA was degraded only by extracts of TNF-α-treated HT-29 cells and not by cytosolic proteins from untreated or IL-1β-treated HT-29 cells. In contrast, extracts of untreated Calu-3 cells enhanced CFTR mRNA degradation, and IL-1β treatment inhibited this; TNF-α had no significant effect. The 3′UTR part of CFTR mRNA was found to be required for this posttranscriptional regulation. The 5′ part of the 3′UTR (the 217 first bases), which contains two AUUUA sequences, was implicated in CFTR mRNA destabilization and the following 136 bases, containing several C-repeats in U-rich environment, in its protection. The proteins, which reacted with the U- and C-repeats of CFTR mRNA 3′UTR, were mainly controlled by stimulation of the p42/p44 and p38 MAP kinase cascades with interaction between these pathways. This posttranscriptional control of gene expression is a common feature of CFTR and many proteins of inflammation.

scholarly journals SUMO Paralog Specific Functions Revealed through Systematic Analysis of Human Knockout Cell Lines and Gene Expression Data

2021 ◽  
pp. mbc.E21-01-0031
Author(s):  
Danielle Bouchard ◽  
Wei Wang ◽  
Wei-Chih Yang ◽  
Shuying He ◽  
Anthony Garcia ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Lines ◽  
Genotoxic Stress ◽  
Nuclear Body ◽  
Human Tissues ◽  
Cellular Functions ◽  
Systematic Analysis ◽  
Control Of Gene Expression ◽  
Knock Out ◽  
Sequence Homologies

The small ubiquitin-related modifiers (SUMOs) regulate nearly every aspect of cellular function, from gene expression in the nucleus to ion transport at the plasma membrane. In humans, the SUMO pathway has five SUMO paralogs with sequence homologies that range from 45% to 97%. SUMO1 and SUMO2 are the most distantly related paralogs, and also the best studied. To what extent SUMO1, SUMO2 and the other paralogs impart unique and non-redundant effects on cellular functions, however, has not been systematically examined and is therefore not fully understood. For instance, knockout studies in mice have revealed conflicting requirements for the paralogs during development and studies in cell culture have relied largely on transient paralog overexpression or knockdown. To address the existing gap in understanding, we first analyzed SUMO paralog gene expression levels in normal human tissues and found unique patterns of SUMO1-3 expression across 30 tissue types, suggesting paralog-specific functions in adult human tissues. To systematically identify and characterize unique and non-redundant functions of the SUMO paralogs in human cells, we next used CRISPR-Cas9 to knock out SUMO1 and SUMO2 expression in osteosarcoma (U2OS) cells. Analysis of these knockout cell lines revealed essential functions for SUMO1 and SUMO2 in regulating cellular morphology, PML nuclear body structure, responses to proteotoxic and genotoxic stress, and control of gene expression. Collectively, our findings reveal non-redundant regulatory roles for SUMO1 and SUMO2 in controlling essential cellular processes and provide a basis for more precise SUMO-targeting therapies.

scholarly journals Effects of the maleless mutation on X and autosomal gene expression in Drosophila melanogaster.

Genetics ◽  
1994 ◽  
Vol 136 (3) ◽  
pp. 913-926
Author(s):  
J C Hiebert ◽  
J A Birchler
Keyword(s):  
Gene Expression ◽  
Regulatory Genes ◽  
Autosomal Gene ◽  
Mrna Levels ◽  
Chromosome 2 ◽  
Artificial Chromosomes ◽  
Yeast Artificial Chromosomes ◽  
Mutational Effect ◽  
Lethal Genes ◽  
Do So

Abstract The mutational effect of the maleless (mle) gene in Drosophila has been reexamined. Earlier work had suggested that mle along with other male-lethal genes was responsible for hypertranscription of the X chromosome in males to bring about dosage compensation. Prompted by studies on dosage sensitive regulatory genes, we tested for effects of mlets on the phenotypes of 16 X or autosomal mutations in adult escapers of lethality. In third instar larvae, prior to the major lethal phase of mle, we examined activities of 6 X or autosomally encoded enzymes, steady state mRNA levels of 15 X-linked or autosomal genes and transcripts from two large genomic segments derived from either the X or from chromosome 2 and present in yeast artificial chromosomes. In contrast to the previously hypothesized role, we detected pronounced effects of mle on the expression of both X-linked and autosomal loci such that a large proportion of the tested genes were increased in expression, while only two X-linked loci were reduced. The most prevalent consequence was an increase of autosomal gene expression, which can explain previously observed reduced X:autosome transcription ratios. These observations suggest that if mle plays a role in the discrimination of the X and the autosomes, it may do so by modification of the effects of dosage sensitive regulatory genes.

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