Immunoglobulin mRNA stability varies during B lymphocyte differentiation.

Jäck, Hans‐Martin; Wabl, Matthias

doi:10.1002/j.1460-2075.1988.tb02911.x

Cited by 75 publications

(31 citation statements)

References 36 publications

(17 reference statements)

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“…The temperature of the culture was abruptly shifted to 37°C by adding an equal volume of YPD medium that had been prewarmed to 49°C. Aliquots of the culture (100 ml) were removed at 0, 5,10,15,20,30,40,50, and 60 min after the temperature shift. Cells were rapidly harvested on a nitrocellulose filter (Whatman no.…”

Section: Methodsmentioning

confidence: 99%

“…mRNA decay rates determine how quickly each message can adapt to a new steady-state level after a change in transcription rate, and dynamic control of the decay of specific transcripts can have an important role in their regulation (1). The decay rates of specific mRNAs can vary by 100-fold or more (2,3) and are affected by a wide variety of stimuli and cellular signals, including specific hormones (2,4), iron (5,6), cell cycle progression (7,8), cell differentiation (9,10), and viral infection (11). Characterizing this stage in the natural history of each mRNA is an important step toward understanding the logic and molecular mechanisms underlying the regulation of the gene expression program of a genome.…”

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confidence: 99%

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Precision and functional specificity in mRNA decay

Wang

Liu

Storey

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

652

758

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Posttranscriptional processing of mRNA is an integral component of the gene expression program. By using DNA microarrays, we precisely measured the decay of each yeast mRNA, after thermal inactivation of a temperature-sensitive RNA polymerase II. The half-lives varied widely, ranging from ϳ3 min to more than 90 min. We found no simple correlation between mRNA half-lives and ORF size, codon bias, ribosome density, or abundance. However, the decay rates of mRNAs encoding groups of proteins that act together in stoichiometric complexes were generally closely matched, and other evidence pointed to a more general relationship between physiological function and mRNA turnover rates. The results provide strong evidence that precise control of the decay of each mRNA is a fundamental feature of the gene expression program in yeast.A though initiation of transcription is well studied and its importance in regulation is clear, we know much less about the specificity, precision and regulatory role of mRNA decay. The abundance of each mRNA in the cell is determined not only by the rate at which it is produced, but also by its rate of degradation. mRNA decay rates determine how quickly each message can adapt to a new steady-state level after a change in transcription rate, and dynamic control of the decay of specific transcripts can have an important role in their regulation (1). The decay rates of specific mRNAs can vary by 100-fold or more (2, 3) and are affected by a wide variety of stimuli and cellular signals, including specific hormones (2, 4), iron (5, 6), cell cycle progression (7,8), cell differentiation (9, 10), and viral infection (11). Characterizing this stage in the natural history of each mRNA is an important step toward understanding the logic and molecular mechanisms underlying the regulation of the gene expression program of a genome.With its ease of biochemical and genetic manipulation, yeast makes an excellent model for studying eukaryotic mRNA turnover. Simple and reliable procedures have been developed to measure the decay rates of individual mRNAs in yeast, including global or specific transcriptional shut-off assays (12)(13)(14) and in vivo kinetic labeling (15). However, the global profile of yeast mRNA turnover has not been systematically and quantitatively determined. In this report, we present the results of a genomewide determination of mRNA decay rates, coupling the global transcriptional shut-off assay with DNA microarray analysis. ura3-52 his4-939am rpb1-1), carrying a temperature-sensitive mutation in RNA polymerase II (13), was used in this study. Y262 was grown in 500 ml of yeast extract͞peptone͞dextrose (YPD) medium at 24°C to OD 600 ϳ0.5. The temperature of the culture was abruptly shifted to 37°C by adding an equal volume of YPD medium that had been prewarmed to 49°C. Aliquots of the culture (100 ml) were removed at 0,5,10,15,20,30,40, 50, and 60 min after the temperature shift. Cells were rapidly harvested on a nitrocellulose filter (Whatman no. 141109) followed by immediate freezing in l...

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Precision and functional specificity in mRNA decay

Wang

Liu

Storey

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

652

758

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“…Total RNA from HeLa-tTA or H2KD cells was prepared with the RNeasy mini kit (QIAGEN) and analyzed by Northern blotting as described previously (25). Briefly, RNA was separated on 1.2% denaturing agarose-formaldehyde gels and transferred onto GeneScreen Plus membrane (PerkinElmer, Zaventems, Belgium).…”

Section: Methodsmentioning

confidence: 99%

“…Briefly, RNA was separated on 1.2% denaturing agarose-formaldehyde gels and transferred onto GeneScreen Plus membrane (PerkinElmer, Zaventems, Belgium). Hybridizations with radioactively labeled DNA probes and subsequent washings were performed as previously described (25). Radioactive bands were detected by autoradiography and with a Fujifilm FLA 3000 phosphorimager.…”

Section: Methodsmentioning

confidence: 99%

hUPF2 Silencing Identifies Physiologic Substrates of Mammalian Nonsense-Mediated mRNA Decay

Wittmann

Hol

Jäck

2006

Molecular and Cellular Biology

216

213

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Nonsense-mediated mRNA decay (NMD) is a conserved eukaryotic surveillance pathway that selectively degrades aberrant mRNAs with premature termination codons (PTCs). Although a small number of cases exist in mammals, where NMD controls levels of physiologic PTC transcripts, it is still unclear whether the engagement of NMD in posttranscriptional control of gene expression is a more prevalent phenomenon. To identify physiologic NMD substrates and to study how NMD silencing affects the overall dynamics of a cell, we stably down-regulated hUPF2, the human homolog of the yeast NMD factor UPF2, by RNA interference. As expected, hUPF2-silenced HeLa cells were impaired in their ability to recognize ectopically expressed aberrant PTC transcripts. Surprisingly, hUPF2 silencing did not affect cell growth and viability but clearly diminished phosphorylation of hUPF1, suggesting a role of hUPF2 in modulating NMD activity through phosphorylation of hUPF1. Genome-wide DNA microarray expression profiling identified 37 novel up-regulated and 57 downregulated transcripts in hUPF2-silenced cells. About 60% of the up-regulated mRNAs carry typical NMD motifs. Hence, NMD is important not only for maintaining the transcriptome integrity by removing nonfunctional and aberrant PTC-bearing transcripts but also for posttranscriptional control of selected physiologic transcripts with NMD features.Eukaryotes have acquired numerous ways to control the integrity and quality of transcripts; one of them is the so-called nonsense-mediated mRNA decay (NMD). This posttranscriptional mRNA surveillance pathway recognizes and degrades aberrant (nonsense) transcripts with premature termination codons (PTCs), thereby preventing accumulation of truncated nonfunctional or potentially noxious polypeptides as well as dissipation of energy for translating aberrant mRNA. The physiological relevance of NMD is demonstrated in several pathological situations, where NMD-resistant nonsense mRNAs accumulate. For example, the presence of stable nonsense mRNAs encoding truncated ␤-globin or ␤-amyloid precursor protein correlates with the onset of ␤ 0 -thalassemia (52) or the formation of neuritic plaques in Alzheimer's patients (65), respectively.NMD was first observed in Saccharomyces cerevisiae (40) and Caenorhabditis elegans (23) and seems to operate in all eukaryotes, including mammals (13). Recently, homologs of the three yeast NMD factors Upf1p, Upf2p, and Upf3p (for Up frameshift protein) have been identified in humans. hUPF1/ rent1 (3,41,45,56,58), the human homolog of yeast UPF1/ regulator of nonsense transcripts, is an ATP-dependent RNA helicase (5), can be phosphorylated at serine residues in SQ motifs (55,70), and resides in the cytoplasm (3), although it can enter the nucleus (46). hUPF1 is supposed to trigger efficient degradation of nonsense transcripts once it has been recruited to the mRNA by hUPF2/rent2. hUPF2 can also be phosphorylated (12), has no known enzymatic activity, shows strong perinuclear staining, and interacts not only with hUPF1 (41) ...

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“…described by Jack and Wabl (30). DRB (100 ,uM from a 100 mM stock dissolved in dimethyl sulfoxide) was added to cells (5 x 105/ml) in tissue culture medium and incubated at 37°C for various times.…”

Section: Methodsmentioning

confidence: 99%

Lipopolysaccharide and dexamethasone induce mouse mammary tumor proviral gene expression and differentiation in B lymphocytes through distinct regulatory pathways.

King¹,

Corley²

1990

Mol. Cell. Biol.

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Endogenous mouse mammary tumor virus (MMTV) proviral transcripts are up regulated during the normal course of B-lymphocyte differentiation. We report here that the regulatory mechanisms which lead to increased levels of MMTV transcripts in differentiating, lipopolysaccharide (LPS)-stimulated normal B cells and in the inducible B-cell lymphoma line CH12 are at least partially distinct from those controlling increases in immunoglobulin and J-chain gene expression. In studies designed to characterize the stimulatory pathways leading to MMTV expression in CH12 cells, we found that stimulation with either LPS or dexamethasone (Dex), a transcriptional activator of MMTV genes, induced not only MMTV expression but also differentiation to antibody secretion. Only Dex-induced and not LPS-induced MMTV expression and differentiation were inhibited by the glucocorticoid antagonist RU486, demonstrating that Dex and LPS stimulate B cells by distinct molecular pathways. Therefore, in B cells, MMTV expression can be regulated via either the conventional hormone receptor-dependent pathway or a hormone receptor-independent pathway. Furthermore, these results suggest that steroid stimulation of B cells can lead to alterations in the expression of other steroid-responsive genes that can become involved in the process of B-cell differentiation.The stimulation of B lymphocytes results in the replication and differentiation of the responding subset into cells that secrete antibodies at a high rate. An understanding of the signal transduction pathways that are involved in the early stages of antigen-specific and mitogen-induced B-cell activation is emerging (reviewed in references 16 and 33), and the molecular events which regulate the increased steady-state expression of immunoglobulin and immunoglobulin-associated J-chain genes, known to be important to the differentiation process, are being characterized (9,11,37,46). Despite advances in these areas, however, the molecular events that are responsible for the differentiation of B cells into antibody-secreting cells remain ill defined. We have approached this problem by identifying and characterizing non-immunoglobulin gene products that are present in higher abundance in differentiating, antibody-secreting cells than in proliferating, non-antibody-secreting cells. We reasoned that an analysis of the molecular events which regulate the inducible expression of such gene products coupled with an understanding of their function should facilitate our understanding of the B-cell differentiation process.Using subtraction analysis, we recently identified a nonimmunoglobulin gene transcript that is expressed in higher abundance in lipopolysaccharide (LPS)-stimulated murine B cells than in unstimulated cells (54). Restriction mapping and sequence analysis of the gene product expressed in the inducible B-cell lymphoma line CH12 revealed that it was encoded by an endogenous mouse mammary tumor virus (MMTV) proviral locus, Mtv-9 (55). Although the cell surface expression of proteins which cross-re...

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Immunoglobulin mRNA stability varies during B lymphocyte differentiation.

Cited by 75 publications

References 36 publications

Precision and functional specificity in mRNA decay

Precision and functional specificity in mRNA decay

hUPF2 Silencing Identifies Physiologic Substrates of Mammalian Nonsense-Mediated mRNA Decay

Lipopolysaccharide and dexamethasone induce mouse mammary tumor proviral gene expression and differentiation in B lymphocytes through distinct regulatory pathways.

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