Accelerated poly(A) loss and mRNA stabilization are independent effects of protein synthesis inhibition on α-tubulin mRNA in<i>Chlamydomonas</i>

Baker, Ellen J.; Liggit, Peggy

doi:10.1093/nar/21.9.2237

Cited by 12 publications

(13 citation statements)

References 48 publications

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“…Figure 2B, D, and F shows mRNA levels in nondeflagellated cells treated with Act-D. Figure 2B and D shows that the constitutively expressed ␣1-tubulin mRNA and the tubHApG mRNA both disappear relatively slowly in the presence of Act-D. Although this experiment was not carried out long enough to obtain an accurate half-life, an approximate half-life of 70 to 75 min was calculated, consistent with previous determinations for ␣-tubulin mRNA (2,5). We know that Act-D was effectively inhibiting transcription in these studies, because rehybridization of the blots with a probe for a short-lived mRNA showed that it disappeared rapidly (data not shown).…”

Section: Resultssupporting

confidence: 85%

“…The blot is hybridized with an antisense HA probe that recognizes the fragment derived from the intact tubHApG mRNA but not the decay intermediate. This blot shows that the poly(A) lengths exhibited by the tubHApG mRNA are typical of those exhibited by the endogenous tubulin mRNAs (4,5). The induced tubulin mRNAs are synthesized in a transcriptional burst that peaks within 15 min and is essentially over by 30 min after deflagellation (2, 3).…”

Section: Resultsmentioning

confidence: 99%

“…We know that tubulin mRNAs induced in the presence of CX are predominantly cytoplasmic because (i) they are polysomal, by the criterion of sedimentation in polysomal regions of sucrose gradients in an EDTA-releasable form, and (ii) they are subject to poly(A) shortening within minutes after their synthesis, a process only known to occur in the cytoplasm (4,5). Thus, it is most likely that CX is blocking a cytoplasmic process.…”

Section: Discussionmentioning

confidence: 99%

“…Methylene blue staining of the blots was routinely performed to evaluate the relative RNA loading and integrity. The filters were hybridized with 32 P-labeled riboprobes specific for the rbcS2 (ribulose bisphosphate carboxylase small-subunit 2) mRNA and ␣1-tubulin 3Ј untranslated region (UTR) as previously described (5). Oligonucleotide probes were 5Ј-end labeled with 32 P by the method of Sambrook et al (30).…”

Section: Methodsmentioning

confidence: 99%

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Deadenylation-Dependent and -Independent Decay Pathways for α1-Tubulin mRNA in Chlamydomonas reinhardtii

Gera

Baker

1998

Molecular and Cellular Biology

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Section: Resultssupporting

confidence: 85%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Deadenylation-Dependent and -Independent Decay Pathways for α1-Tubulin mRNA in Chlamydomonas reinhardtii

Gera

Baker

1998

Molecular and Cellular Biology

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“…In plant material not treated with CHX, results from the RNA gel blot/RT-PCR consistently concurred with the results obtained from the microarray analyses ( Figure 3). Some genes had enhanced expression in the presence of CHX compared with those not CHX treated ( Figure 3); this is consistent with prior observations of enhanced mRNA stability upon CHX treatment (Baker and Liggit, 1993;Goda et al, 2002). The relative stabilization of some transcripts upon CHX treatment indirectly suggests that posttranscriptional modifications may be occurring.…”

Section: Transcriptional Upregulation By Glucose Largely Requires De supporting

confidence: 89%

Global Transcription Profiling Reveals Multiple Sugar Signal Transduction Mechanisms in Arabidopsis[W]

et al. 2004

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Complex and interconnected signaling networks allow organisms to control cell division, growth, differentiation, or programmed cell death in response to metabolic and environmental cues. In plants, it is known that sugar and nitrogen are critical nutrient signals; however, our understanding of the molecular mechanisms underlying nutrient signal transduction is very limited. To begin unraveling complex sugar signaling networks in plants, DNA microarray analysis was used to determine the effects of glucose and inorganic nitrogen source on gene expression on a global scale in Arabidopsis thaliana. In whole seedling tissue, glucose is a more potent signal in regulating transcription than inorganic nitrogen. In fact, other than genes associated with nitrate assimilation, glucose had a greater effect in regulating nitrogen metabolic genes than nitrogen itself. Glucose also regulated a broader range of genes, including genes associated with carbohydrate metabolism, signal transduction, and metabolite transport. In addition, a large number of stress responsive genes were also induced by glucose, indicating a role of sugar in environmental responses. Cluster analysis revealed significant interaction between glucose and nitrogen in regulating gene expression because glucose can modulate the effects of nitrogen and vise versa. Intriguingly, cycloheximide treatment appeared to disrupt glucose induction more than glucose repression, suggesting that de novo protein synthesis is an intermediary event required before most glucose induction can occur. Cross talk between sugar and ethylene signaling may take place on the transcriptional level because several ethylene biosynthetic and signal transduction genes are repressed by glucose, and the repression is largely unaffected by cycloheximide. Collectively, our global expression data strongly support the idea that glucose and inorganic nitrogen act as both metabolites and signaling molecules.

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Tubulin mRNA instability and stabilization by protein synthesis inhibitors are reproducible in nontranslating extracts from Chlamydomonas

Baker

1993

Dev. Genet.

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In Chlamydomonas reinhardtii, flagellar amputation stimulates an induction in the synthesis of flagellar proteins which allows the cells to rapidly regenerate their flagella. The induction involves the coordinate accumulation and rapid degradation of a large number mRNAs, including those encoding the tubulins. The post-induction degradation of induced tubulin mRNAs has been shown to differ from the constitutive turnover pathway in two ways: (1) the rate of degradation is accelerated, and (2) degradation is prevented by inhibition of protein synthesis. In this report, it is shown that the post-induction degradation of all deflagellation-induced mRNAs examined is prevented by cycloheximide (CX), suggesting they all may be degraded via the same pathway. A cell-free decay system has been developed to investigate the degradation pathway. At least two characteristics of tubulin mRNA degradation are reproducible in these extracts: (1) endogenous alpha-tubulin mRNA is less stable than constitutive mRNAs in the same extract and (2) alpha-tubulin mRNA in extracts prepared from CX-treated cells (CX extracts) is significantly more stable than it is in extracts from untreated cells (control extracts). This indicates that the mechanism by which CX blocks rapid degradation of tubulin mRNA in vivo is not simply by preventing its translation and suggests the involvement of an altered trans-factor. The difference in tubulin mRNA stability in the two extracts is maintained when the extracts are prepared under conditions that dissociate ribosomes from mRNPs, indicating intact polysome structure is not necessary. Tubulin mRNA-containing polysomes isolated from control and CX extracts are equally stable when assayed alone. However, the polysomes from control extracts are more sensitive to exogenous RNAse treatment than are those from CX extracts, indicating a structural difference. There are no detectable differences in soluble factors that influence tubulin mRNA degradation rate between control and CX extracts; addition of excess soluble factors to either control or CX extracts does not alter the tubulin mRNA degradation in the extract, nor does a simple one-to-one combination of the two extracts result in stabilization or destabilization of the whole population of tubulin mRNAs in the mixture. The deflagellation-induced mRNAs, as a group, are shown to be particularly susceptible to a nuclease activity in extracts, inhibitable by vanadyl ribonucleoside complexes, which does not appear to attack constitutive mRNAs. It is proposed that a structural difference in the tubulin mRNPs produced in the presence and absence of CX underlies their differences in stabilities, and that a common nuclease targets the induced flagellar protein mRNAs.

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Accelerated poly(A) loss and mRNA stabilization are independent effects of protein synthesis inhibition on α-tubulin mRNA inChlamydomonas

Cited by 12 publications

References 48 publications

Deadenylation-Dependent and -Independent Decay Pathways for α1-Tubulin mRNA in Chlamydomonas reinhardtii

Deadenylation-Dependent and -Independent Decay Pathways for α1-Tubulin mRNA in Chlamydomonas reinhardtii

Global Transcription Profiling Reveals Multiple Sugar Signal Transduction Mechanisms in Arabidopsis[W]

Tubulin mRNA instability and stabilization by protein synthesis inhibitors are reproducible in nontranslating extracts from Chlamydomonas

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