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. 1991 Feb;11(2):813–821. doi: 10.1128/mcb.11.2.813

Yeast CBP1 mRNA 3' end formation is regulated during the induction of mitochondrial function.

S A Mayer 1, C L Dieckmann 1
PMCID: PMC359733  PMID: 1990285

Abstract

Alternative mRNA processing is one mechanism for generating two or more polypeptides from a single gene. While many mammalian genes contain multiple mRNA 3' cleavage and polyadenylation signals that change the coding sequence of the mature mRNA when used at different developmental stages or in different tissues, only one yeast gene has been identified with this capacity. The Saccharomyces cerevisiae nuclear gene CPB1 encodes a mitochondrial protein that is required for cytochrome b mRNA stability. This 66-kDa protein is encoded by a 2.2-kb mRNA transcribed from CPB1. Previously we showed that a second 1.2-kb transcript is initiated at the CBP1 promoter but has a 3' end near the middle of the coding sequence. Furthermore, it was shown that the ratio of the steady-state level of 2.2-kb CBP1 message to 1.2-kb message decreases 10-fold during the induction of mitochondrial function, while the combined levels of both messages remain constant. Having proposed that regulation of 3' end formation dictates the amount of each CBP1 transcript, we now show that a 146-bp fragment from the middle of CBP1 is sufficient to direct carbon source-regulated production of two transcripts when inserted into the yeast URA3 gene. This fragment contains seven polyadenylation sites for the wild-type 1.2-kb mRNA, as mapped by sequence analysis of CBP1 cDNA clones. Deletion mutations upstream of the polyadenylation sites abolished formation of the 1.2-kb transcript, whereas deletion of three of the sites only led to a reduction in abundance of the 1.2-kb mRNA. Our results indicate that regulation of the abundance of both CBP1 transcripts is controlled by elements in a short segment of the gene that directs 3' end formation of the 1.2-kb transcript, a unique case in yeast cells.

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Selected References

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  1. Alt F. W., Bothwell A. L., Knapp M., Siden E., Mather E., Koshland M., Baltimore D. Synthesis of secreted and membrane-bound immunoglobulin mu heavy chains is directed by mRNAs that differ at their 3' ends. Cell. 1980 Jun;20(2):293–301. doi: 10.1016/0092-8674(80)90615-7. [DOI] [PubMed] [Google Scholar]
  2. Alwine J. C., Kemp D. J., Stark G. R. Method for detection of specific RNAs in agarose gels by transfer to diazobenzyloxymethyl-paper and hybridization with DNA probes. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5350–5354. doi: 10.1073/pnas.74.12.5350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Amara S. G., Evans R. M., Rosenfeld M. G. Calcitonin/calcitonin gene-related peptide transcription unit: tissue-specific expression involves selective use of alternative polyadenylation sites. Mol Cell Biol. 1984 Oct;4(10):2151–2160. doi: 10.1128/mcb.4.10.2151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Birnstiel M. L., Busslinger M., Strub K. Transcription termination and 3' processing: the end is in site! Cell. 1985 Jun;41(2):349–359. doi: 10.1016/s0092-8674(85)80007-6. [DOI] [PubMed] [Google Scholar]
  5. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  6. Butler J. S., Platt T. RNA processing generates the mature 3' end of yeast CYC1 messenger RNA in vitro. Science. 1988 Dec 2;242(4883):1270–1274. doi: 10.1126/science.2848317. [DOI] [PubMed] [Google Scholar]
  7. Butler J. S., Sadhale P. P., Platt T. RNA processing in vitro produces mature 3' ends of a variety of Saccharomyces cerevisiae mRNAs. Mol Cell Biol. 1990 Jun;10(6):2599–2605. doi: 10.1128/mcb.10.6.2599. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Carswell S., Alwine J. C. Efficiency of utilization of the simian virus 40 late polyadenylation site: effects of upstream sequences. Mol Cell Biol. 1989 Oct;9(10):4248–4258. doi: 10.1128/mcb.9.10.4248. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. DeZazzo J. D., Imperiale M. J. Sequences upstream of AAUAAA influence poly(A) site selection in a complex transcription unit. Mol Cell Biol. 1989 Nov;9(11):4951–4961. doi: 10.1128/mcb.9.11.4951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dieckmann C. L., Gandy B. Preferential recombination between GC clusters in yeast mitochondrial DNA. EMBO J. 1987 Dec 20;6(13):4197–4203. doi: 10.1002/j.1460-2075.1987.tb02767.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Dieckmann C. L., Homison G., Tzagoloff A. Assembly of the mitochondrial membrane system. Nucleotide sequence of a yeast nuclear gene (CBP1) involved in 5' end processing of cytochrome b pre-mRNA. J Biol Chem. 1984 Apr 25;259(8):4732–4738. [PubMed] [Google Scholar]
  12. Dieckmann C. L., Koerner T. J., Tzagoloff A. Assembly of the mitochondrial membrane system. CBP1, a yeast nuclear gene involved in 5' end processing of cytochrome b pre-mRNA. J Biol Chem. 1984 Apr 25;259(8):4722–4731. [PubMed] [Google Scholar]
  13. Dieckmann C. L., Mittelmeier T. M. Nuclearly-encoded CBP1 interacts with the 5' end of mitochondrial cytochrome b pre-mRNA. Curr Genet. 1987;12(6):391–397. doi: 10.1007/BF00434815. [DOI] [PubMed] [Google Scholar]
  14. Dieckmann C. L., Pape L. K., Tzagoloff A. Identification and cloning of a yeast nuclear gene (CBP1) involved in expression of mitochondrial cytochrome b. Proc Natl Acad Sci U S A. 1982 Mar;79(6):1805–1809. doi: 10.1073/pnas.79.6.1805. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Dorsman J. C., van Heeswijk W. C., Grivell L. A. Identification of two factors which bind to the upstream sequences of a number of nuclear genes coding for mitochondrial proteins and to genetic elements important for cell division in yeast. Nucleic Acids Res. 1988 Aug 11;16(15):7287–7301. doi: 10.1093/nar/16.15.7287. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Early P., Rogers J., Davis M., Calame K., Bond M., Wall R., Hood L. Two mRNAs can be produced from a single immunoglobulin mu gene by alternative RNA processing pathways. Cell. 1980 Jun;20(2):313–319. doi: 10.1016/0092-8674(80)90617-0. [DOI] [PubMed] [Google Scholar]
  17. Falck-Pedersen E., Logan J. Regulation of poly(A) site selection in adenovirus. J Virol. 1989 Feb;63(2):532–541. doi: 10.1128/jvi.63.2.532-541.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Fitzgerald M., Shenk T. The sequence 5'-AAUAAA-3'forms parts of the recognition site for polyadenylation of late SV40 mRNAs. Cell. 1981 Apr;24(1):251–260. doi: 10.1016/0092-8674(81)90521-3. [DOI] [PubMed] [Google Scholar]
  19. Frohman M. A., Dush M. K., Martin G. R. Rapid production of full-length cDNAs from rare transcripts: amplification using a single gene-specific oligonucleotide primer. Proc Natl Acad Sci U S A. 1988 Dec;85(23):8998–9002. doi: 10.1073/pnas.85.23.8998. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Gampel A., Nishikimi M., Tzagoloff A. CBP2 protein promotes in vitro excision of a yeast mitochondrial group I intron. Mol Cell Biol. 1989 Dec;9(12):5424–5433. doi: 10.1128/mcb.9.12.5424. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Gil A., Proudfoot N. J. A sequence downstream of AAUAAA is required for rabbit beta-globin mRNA 3'-end formation. 1984 Nov 29-Dec 5Nature. 312(5993):473–474. doi: 10.1038/312473a0. [DOI] [PubMed] [Google Scholar]
  22. Gil A., Proudfoot N. J. Position-dependent sequence elements downstream of AAUAAA are required for efficient rabbit beta-globin mRNA 3' end formation. Cell. 1987 May 8;49(3):399–406. doi: 10.1016/0092-8674(87)90292-3. [DOI] [PubMed] [Google Scholar]
  23. Grivell L. A. Nucleo-mitochondrial interactions in yeast mitochondrial biogenesis. Eur J Biochem. 1989 Jul 1;182(3):477–493. doi: 10.1111/j.1432-1033.1989.tb14854.x. [DOI] [PubMed] [Google Scholar]
  24. Helfman D. M., Cheley S., Kuismanen E., Finn L. A., Yamawaki-Kataoka Y. Nonmuscle and muscle tropomyosin isoforms are expressed from a single gene by alternative RNA splicing and polyadenylation. Mol Cell Biol. 1986 Nov;6(11):3582–3595. doi: 10.1128/mcb.6.11.3582. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Hill J. E., Myers A. M., Koerner T. J., Tzagoloff A. Yeast/E. coli shuttle vectors with multiple unique restriction sites. Yeast. 1986 Sep;2(3):163–167. doi: 10.1002/yea.320020304. [DOI] [PubMed] [Google Scholar]
  26. Humphrey T., Proudfoot N. J. A beginning to the biochemistry of polyadenylation. Trends Genet. 1988 Sep;4(9):243–245. doi: 10.1016/0168-9525(88)90028-5. [DOI] [PubMed] [Google Scholar]
  27. Ito H., Fukuda Y., Murata K., Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Leff S. E., Rosenfeld M. G., Evans R. M. Complex transcriptional units: diversity in gene expression by alternative RNA processing. Annu Rev Biochem. 1986;55:1091–1117. doi: 10.1146/annurev.bi.55.070186.005303. [DOI] [PubMed] [Google Scholar]
  29. Liu Y. X., Dieckmann C. L. Overproduction of yeast viruslike particles by strains deficient in a mitochondrial nuclease. Mol Cell Biol. 1989 Aug;9(8):3323–3331. doi: 10.1128/mcb.9.8.3323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Mather E. L., Nelson K. J., Haimovich J., Perry R. P. Mode of regulation of immunoglobulin mu- and delta-chain expression varies during B-lymphocyte maturation. Cell. 1984 Feb;36(2):329–338. doi: 10.1016/0092-8674(84)90226-5. [DOI] [PubMed] [Google Scholar]
  31. Mayer S. A., Dieckmann C. L. The yeast CBP1 gene produces two differentially regulated transcripts by alternative 3'-end formation. Mol Cell Biol. 1989 Oct;9(10):4161–4169. doi: 10.1128/mcb.9.10.4161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. McDevitt M. A., Hart R. P., Wong W. W., Nevins J. R. Sequences capable of restoring poly(A) site function define two distinct downstream elements. EMBO J. 1986 Nov;5(11):2907–2913. doi: 10.1002/j.1460-2075.1986.tb04586.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. McDevitt M. A., Imperiale M. J., Ali H., Nevins J. R. Requirement of a downstream sequence for generation of a poly(A) addition site. Cell. 1984 Jul;37(3):993–999. doi: 10.1016/0092-8674(84)90433-1. [DOI] [PubMed] [Google Scholar]
  34. McGraw P., Tzagoloff A. Assembly of the mitochondrial membrane system. Characterization of a yeast nuclear gene involved in the processing of the cytochrome b pre-mRNA. J Biol Chem. 1983 Aug 10;258(15):9459–9468. [PubMed] [Google Scholar]
  35. McLauchlan J., Simpson S., Clements J. B. Herpes simplex virus induces a processing factor that stimulates poly(A) site usage. Cell. 1989 Dec 22;59(6):1093–1105. doi: 10.1016/0092-8674(89)90765-4. [DOI] [PubMed] [Google Scholar]
  36. Montell C., Fisher E. F., Caruthers M. H., Berk A. J. Inhibition of RNA cleavage but not polyadenylation by a point mutation in mRNA 3' consensus sequence AAUAAA. Nature. 1983 Oct 13;305(5935):600–605. doi: 10.1038/305600a0. [DOI] [PubMed] [Google Scholar]
  37. Nevins J. R. The pathway of eukaryotic mRNA formation. Annu Rev Biochem. 1983;52:441–466. doi: 10.1146/annurev.bi.52.070183.002301. [DOI] [PubMed] [Google Scholar]
  38. Nevins J. R., Wilson M. C. Regulation of adenovirus-2 gene expression at the level of transcriptional termination and RNA processing. Nature. 1981 Mar 12;290(5802):113–118. doi: 10.1038/290113a0. [DOI] [PubMed] [Google Scholar]
  39. Olesen J., Hahn S., Guarente L. Yeast HAP2 and HAP3 activators both bind to the CYC1 upstream activation site, UAS2, in an interdependent manner. Cell. 1987 Dec 24;51(6):953–961. doi: 10.1016/0092-8674(87)90582-4. [DOI] [PubMed] [Google Scholar]
  40. Osborne B. I., Guarente L. Mutational analysis of a yeast transcriptional terminator. Proc Natl Acad Sci U S A. 1989 Jun;86(11):4097–4101. doi: 10.1073/pnas.86.11.4097. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Osborne B. I., Guarente L. Transcription by RNA polymerase II induces changes of DNA topology in yeast. Genes Dev. 1988 Jun;2(6):766–772. doi: 10.1101/gad.2.6.766. [DOI] [PubMed] [Google Scholar]
  42. Proudfoot N. J. How RNA polymerase II terminates transcription in higher eukaryotes. Trends Biochem Sci. 1989 Mar;14(3):105–110. doi: 10.1016/0968-0004(89)90132-1. [DOI] [PubMed] [Google Scholar]
  43. Roberts J. W. Phage lambda and the regulation of transcription termination. Cell. 1988 Jan 15;52(1):5–6. doi: 10.1016/0092-8674(88)90523-5. [DOI] [PubMed] [Google Scholar]
  44. Rogers J., Early P., Carter C., Calame K., Bond M., Hood L., Wall R. Two mRNAs with different 3' ends encode membrane-bound and secreted forms of immunoglobulin mu chain. Cell. 1980 Jun;20(2):303–312. doi: 10.1016/0092-8674(80)90616-9. [DOI] [PubMed] [Google Scholar]
  45. Rose M., Winston F. Identification of a Ty insertion within the coding sequence of the S. cerevisiae URA3 gene. Mol Gen Genet. 1984;193(3):557–560. doi: 10.1007/BF00382100. [DOI] [PubMed] [Google Scholar]
  46. Rothstein R. J. One-step gene disruption in yeast. Methods Enzymol. 1983;101:202–211. doi: 10.1016/0076-6879(83)01015-0. [DOI] [PubMed] [Google Scholar]
  47. Russnak R., Ganem D. Sequences 5' to the polyadenylation signal mediate differential poly(A) site use in hepatitis B viruses. Genes Dev. 1990 May;4(5):764–776. doi: 10.1101/gad.4.5.764. [DOI] [PubMed] [Google Scholar]
  48. Russo P., Sherman F. Transcription terminates near the poly(A) site in the CYC1 gene of the yeast Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1989 Nov;86(21):8348–8352. doi: 10.1073/pnas.86.21.8348. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Rödel G., Fox T. D. The yeast nuclear gene CBS1 is required for translation of mitochondrial mRNAs bearing the cob 5' untranslated leader. Mol Gen Genet. 1987 Jan;206(1):45–50. doi: 10.1007/BF00326534. [DOI] [PubMed] [Google Scholar]
  50. Rödel G., Körte A., Kaudewitz F. Mitochondrial suppression of a yeast nuclear mutation which affects the translation of the mitochondrial apocytochrome b transcript. Curr Genet. 1985;9(8):641–648. doi: 10.1007/BF00449816. [DOI] [PubMed] [Google Scholar]
  51. Saez C. G., Myers J. C., Shows T. B., Leinwand L. A. Human nonmuscle myosin heavy chain mRNA: generation of diversity through alternative polyadenylylation. Proc Natl Acad Sci U S A. 1990 Feb;87(3):1164–1168. doi: 10.1073/pnas.87.3.1164. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Simonsen C. C., Levinson A. D. Analysis of processing and polyadenylation signals of the hepatitis B virus surface antigen gene by using simian virus 40-hepatitis B virus chimeric plasmids. Mol Cell Biol. 1983 Dec;3(12):2250–2258. doi: 10.1128/mcb.3.12.2250. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Smith C. W., Patton J. G., Nadal-Ginard B. Alternative splicing in the control of gene expression. Annu Rev Genet. 1989;23:527–577. doi: 10.1146/annurev.ge.23.120189.002523. [DOI] [PubMed] [Google Scholar]
  55. Snyder M., Sapolsky R. J., Davis R. W. Transcription interferes with elements important for chromosome maintenance in Saccharomyces cerevisiae. Mol Cell Biol. 1988 May;8(5):2184–2194. doi: 10.1128/mcb.8.5.2184. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Tzagoloff A., Myers A. M. Genetics of mitochondrial biogenesis. Annu Rev Biochem. 1986;55:249–285. doi: 10.1146/annurev.bi.55.070186.001341. [DOI] [PubMed] [Google Scholar]
  57. Van Loon A. P., Van Eijk E., Grivell L. A. Biosynthesis of the ubiquinol-cytochrome c reductase complex in yeast. Discoordinate synthesis of the 11-kd subunit in response to increased gene copy number. EMBO J. 1983;2(10):1765–1770. doi: 10.1002/j.1460-2075.1983.tb01655.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Vincent R. D., Hofmann T. J., Zassenhaus H. P. Sequence and expression of NUC1, the gene encoding the mitochondrial nuclease in Saccharomyces cerevisiae. Nucleic Acids Res. 1988 Apr 25;16(8):3297–3312. doi: 10.1093/nar/16.8.3297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Weber E. R., Dieckmann C. L. Identification of the CBP1 polypeptide in mitochondrial extracts from Saccharomyces cerevisiae. J Biol Chem. 1990 Jan 25;265(3):1594–1600. [PubMed] [Google Scholar]
  60. Wickens M., Stephenson P. Role of the conserved AAUAAA sequence: four AAUAAA point mutants prevent messenger RNA 3' end formation. Science. 1984 Nov 30;226(4678):1045–1051. doi: 10.1126/science.6208611. [DOI] [PubMed] [Google Scholar]
  61. Woychik R. P., Lyons R. H., Post L., Rottman F. M. Requirement for the 3' flanking region of the bovine growth hormone gene for accurate polyadenylylation. Proc Natl Acad Sci U S A. 1984 Jul;81(13):3944–3948. doi: 10.1073/pnas.81.13.3944. [DOI] [PMC free article] [PubMed] [Google Scholar]
  62. Wu M., Tzagoloff A. Identification and characterization of a new gene (CBP3) required for the expression of yeast coenzyme QH2-cytochrome c reductase. J Biol Chem. 1989 Jul 5;264(19):11122–11130. [PubMed] [Google Scholar]
  63. Zaret K. S., Sherman F. DNA sequence required for efficient transcription termination in yeast. Cell. 1982 Mar;28(3):563–573. doi: 10.1016/0092-8674(82)90211-2. [DOI] [PubMed] [Google Scholar]
  64. Zarkower D., Stephenson P., Sheets M., Wickens M. The AAUAAA sequence is required both for cleavage and for polyadenylation of simian virus 40 pre-mRNA in vitro. Mol Cell Biol. 1986 Jul;6(7):2317–2323. doi: 10.1128/mcb.6.7.2317. [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. Ziff E. B. Transcription and RNA processing by the DNA tumour viruses. Nature. 1980 Oct 9;287(5782):491–499. doi: 10.1038/287491a0. [DOI] [PubMed] [Google Scholar]
  66. van Loon A. P., de Groot R. J., van Eyk E., van der Horst G. T., Grivell L. A. Isolation and characterization of nuclear genes coding for subunits of the yeast ubiquinol-cytochrome c reductase complex. Gene. 1982 Dec;20(3):323–337. doi: 10.1016/0378-1119(82)90201-3. [DOI] [PubMed] [Google Scholar]

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