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Regulation of cytoplasmic mRNA prevalence in sea urchin embryos: Rates of appearance and turnover for specific sequences

Cabrera, Carlos V. and Lee, James J. and Ellison, Jay W. and Britten, Roy J. and Davidson, Eric H. (1984) Regulation of cytoplasmic mRNA prevalence in sea urchin embryos: Rates of appearance and turnover for specific sequences. Journal of Molecular Biology, 174 (1). pp. 85-111. ISSN 0022-2836. https://resolver.caltech.edu/CaltechAUTHORS:20160513-090917087

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Abstract

Complementary DNA clones representing cytoplasmic poly(A) RNAs of sea urchin embryos were hybridized with metabolically labeled cytoplasmic RNA preparations and the rates of appearance and of decay for each transcript species were determined at the blastula-gastrula stage of development. The prevalence of the transcripts chosen for this study ranged, on average, from about one molecule per cell to a few hundred molecules per cell. The embryos were labeled continuously for 18 hours with [^3H]guanosine, beginning at 24 hours post-fertilization. The amount of cytoplasmic [^3H]poly(A) RNA that hybridized to each cloned sequence was determined and the specific activity of the [^3H]GTP pool was measured in the same embryos. Rate constants for the entry of each transcript species into the cytoplasm, and for its decay were extracted from these data. The embryo transcript species identified by the cloned probes displayed a range of stabilities. Half-lives of only a few hours were measured both for a very rare sequence and for a moderately prevalent sequence. Other newly synthesized transcripts, including sequences that first appear during embryonic development, as well as sequences also represented in maternal RNA, are far more stable. We conclude that cytoplasmic RNA turnover rate is a major variable in the determination of the cytoplasmic level of expression of embryo genes. The entry rates of the transcripts into the cytoplasm also varied, from a few molecules per embryo per minute to several hundred, depending on the sequence. By comparing the mass of transcripts of a given sequence in the embryo to the mass of transcripts of that sequence accumulating as a result of new synthesis, the point at which embryo transcription accounts for the major fraction of the cytoplasmic molecules could be estimated. This calculation showed that for some sequences maternal transcripts persist well beyond gastrulation, while other embryo poly(A) RNA species are largely the product of transcription in the embryo nuclei from the blastula stage onwards. There is no single stage at which all maternal transcripts are suddenly replaced by newly synthesized embryo transcripts. Primary transcription rates were measured for two sequences by determining accumulation of label in these RNA species soon after addition of [^3H]guanosine to the cultures. Comparing these rates to the cytoplasmic entry rates, we did not detect a significantly greater nuclear transcription of the sequence homologous to the cloned probe.


Item Type:Article
Related URLs:
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http://dx.doi.org/10.1016/0022-2836(84)90366-8DOIArticle
http://www.sciencedirect.com/science/article/pii/0022283684903668PublisherArticle
Additional Information:© 1983 Elsevier Ltd. Received 23 August 1983, and in revised form 30 November 1983. We thank Dr D. Stafford for providing us with clone pLvl334 isolated from a genomic library of Lytechinus pictus. Dr Costas Flytzanis and Mr Edward Nolan provided invaluable assistance in providing some of the prevalence data in Table 3. We are much indebted to Mr James G. Moore of Microcomputer Graphics, Inc., who designed, constructed and perfected the MESSAGE program. We are grateful to Drs N. Davidson, B. Wold and E. Rothenberg of this Institute for reviewing drafts of this manuscript. The research was supported by NICHHD grant HD-05753. C.V.C. was supported by a NIH Fogarty international fellowship (TWO2996); J.J.L. was supported by a NIH training grant (GM-07616), and J.W.E. was supported by a NSF predoctoral fellowship.
Funders:
Funding AgencyGrant Number
NIHHD-05753
NIHTWO2996
NIH Predoctoral FellowshipGM-07616
NSF Predoctoral FellowshipUNSPECIFIED
Issue or Number:1
Record Number:CaltechAUTHORS:20160513-090917087
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20160513-090917087
Official Citation:Carlos V. Cabrera, James J. Lee, Jay W. Ellison, Roy J. Britten, Eric H. Davidson, Regulation of cytoplasmic mRNA prevalence in sea urchin embryos, Journal of Molecular Biology, Volume 174, Issue 1, 1984, Pages 85-111, ISSN 0022-2836, http://dx.doi.org/10.1016/0022-2836(84)90366-8. (http://www.sciencedirect.com/science/article/pii/0022283684903668)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:67064
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:13 May 2016 18:53
Last Modified:03 Oct 2019 10:02

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