1887

Abstract

Alternative processing of human cytomegalovirus (HCMV) UL37 pre-mRNA predominantly produces the unspliced UL37 exon 1 (UL37x1) RNA and multiple, lower abundance, alternatively spliced UL37 RNAs. The relative abundance of UL37x1 unspliced RNA is surprising because it requires the favoured use of a polyadenylation signal within UL37 intron 1, just upstream of the UL37 exon 2 (UL37x2) acceptor. Here, it was shown that a downstream element (DSE) in UL37x2 strongly enhanced processing at the UL37x1 polyadenylation site, but did not influence UL37x1–x2 splicing. There was a potential binding site (UCUU) for polypyrimidine tract-binding protein (PTB) at the UL37x1 polyadenylation/cleavage site and its mutation to UGGG reduced both polyadenylation and splicing of UL37x1–x2 minigene pre-mRNA, suggesting a role in both RNA processing events. To determine whether lytic HCMV infection altered the balance of RNA processing factors, which bind to UL37 pre-mRNA elements, these were investigated in permissively infected primary and immortalized human diploid fibroblasts (HFFs) and epithelial cells. Induction of polyadenylation factors in HCMV-infected, serum-starved (G) HFFs was also investigated. Permissive HCMV infection consistently increased, albeit with different kinetics, the abundance of cleavage stimulation factor 64 (CstF-64) and PTB, and altered hypo-phosphorylated SF2 in different cell types. Moreover, the preponderance of UL37x1 RNA increased during infection and correlated with CstF-64 induction, whereas the complexity of the lower abundance UL37 spliced RNAs transiently increased following reduction of hypo-phosphorylated SF2. Collectively, multiple UL37 RNA polyadenylation elements and induced cellular factors in HCMV-infected cells strongly favoured the production of UL37x1 unspliced RNA.

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2004-12-01
2021-03-08
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References

  1. Adair R., Liebisch G., Colberg-Poley A. M. 2003; Complex alternative processing of the human cytomegalovirus (HCMV) UL37 pre-mRNA. J Gen Virol 84:3353–3358 [CrossRef]
    [Google Scholar]
  2. Al-Barazi H. O., Colberg-Poley A. M. 1996; The human cytomegalovirus UL37 immediate-early regulatory protein is an integral membrane N -glycoprotein which traffics through the endoplasmic reticulum and Golgi apparatus. J Virol 70:7198–7208
    [Google Scholar]
  3. Arnoult D., Bartle L. M., Skaletskaya A., Poncet D., Zamzami N., Park P. U., Sharpe J., Youle R. J., Goldmacher V. S. 2004; Cytomegalovirus cell death suppressor vMIA blocks Bax- but not Bak-mediated apoptosis by binding and sequestering Bax at mitochondria. Proc Natl Acad Sci U S A 101:7988–7993 [CrossRef]
    [Google Scholar]
  4. Awasthi S., Isler J. A., Alwine J. E. 2004; Analysis of splice variants of the immediate-early 1 region of human cytomegalovirus. J Virol 78:8191–8200 [CrossRef]
    [Google Scholar]
  5. Back S. H., Kim Y. K., Kim W. J., Cho S., Oh H. R., Kim J.-E., Jang S. K. 2002a; Translation of polioviral mRNA is inhibited by cleavage of polypyrimidine tract-binding proteins executed by polioviral 3Cpro . J Virol 76:2529–2542 [CrossRef]
    [Google Scholar]
  6. Back S. H., Shin S., Jang S. K. 2002b; Polypyrimidine tract-binding proteins are cleaved by caspase-3 during apoptosis. J Biol Chem 277:27200–27209 [CrossRef]
    [Google Scholar]
  7. Borst E.-M., Hahn G., Koszinowski U. H., Messerle M. 1999; Cloning of the human cytomegalovirus (HCMV) genome as an infectious bacterial artificial chromosome in Escherichia coli : a new approach for construction of HCMV mutants. J Virol 73:8320–8329
    [Google Scholar]
  8. Bothwell A. L. M., Ballard D. W., Philbrick W. M., Lindwall G., Maher S. E., Bridgett M. M., Jamison S. F., Garcia-Blanco M. A. 1991; Murine polypyrimidine tract binding protein. Purification, cloning and mapping of the RNA binding domain. J Biol Chem 266:24657–24663
    [Google Scholar]
  9. Bresnahan W. A., Hultman G. E., Shenk T. 2000; Replication of wild-type and mutant human cytomegalovirus in life-extended human diploid fibroblasts. J Virol 74:10816–10818 [CrossRef]
    [Google Scholar]
  10. Castelo-Branco P., Furger A., Wollerton M., Smith C., Moreira A., Proudfoot N. 2004; Polypyrimidine tract binding protein modulates efficiency of polyadenylation. Mol Cell Biol 24:4174–4183 [CrossRef]
    [Google Scholar]
  11. Chen F., MacDonald C. C., Wilusz J. 1995; Cleavage site determinants in the mammalian polyadenylation signal. Nucleic Acids Res 23:2614–2620 [CrossRef]
    [Google Scholar]
  12. Chou M. Y., Underwood J. G., Nikolic J., Luu M. H., Black D. L. 2000; Multisite RNA binding and release of polypyrimidine tract binding protein during the regulation of c- src neural-specific splicing. Mol Cell 5:949–957 [CrossRef]
    [Google Scholar]
  13. Coats D. K., Demmler G. J., Paysse E. A., Du L. T., Libby C. 2000; Ophthalmologic findings in children with congenital cytomegalovirus infection. J AAPOS 4:110–116 [CrossRef]
    [Google Scholar]
  14. Colberg-Poley A. M., Santomenna L. D. 1988; Selective induction of chromosomal gene expression by human cytomegalovirus. Virology 166:217–228 [CrossRef]
    [Google Scholar]
  15. Colberg-Poley A. M., Patel M. B., Erezo D. P. P., Slater J. E. 2000; Human cytomegalovirus immediate early regulatory proteins traffic through the secretory apparatus and to mitochondria. J Gen Virol 81:1779–1789
    [Google Scholar]
  16. Detrick B., Rhame J., Wang Y., Nagineni C. N., Hooks J. J. 1996; Cytomegalovirus replication in human retinal pigment epithelial cells. Altered expression of viral early proteins. Invest Ophthalmol Vis Sci 37:814–825
    [Google Scholar]
  17. Dunn W., Chou C., Li H., Hai R., Patterson D., Stolc V., Zhu H., Liu F. 2003; Functional profiling of a human cytomegalovirus genome. Proc Natl Acad Sci U S A 100:14223–14228 [CrossRef]
    [Google Scholar]
  18. Furth P. A., Choe W. T., Rex J. H., Byrne J. C., Baker C. C. 1994; Sequences homologous to 5′ splice sites are required for the inhibitory activity of papillomavirus late 3′ untranslated regions. Mol Cell Biol 14:5278–5289
    [Google Scholar]
  19. Gil A., Proudfoot N. J. 1987; Position-dependent sequence elements downstream of AAUAAA are required for efficient rabbit beta-globin mRNA 3′ end formation. Cell 49:399–406 [CrossRef]
    [Google Scholar]
  20. Gil A., Sharp P. A., Jamison S. F., Garcia-Blanco M. A. 1991; Characterization of cDNAs encoding the polypyrimidine tract binding protein. Genes Dev 5:1224–1236 [CrossRef]
    [Google Scholar]
  21. Goldmacher V. S., Bartle L. M., Skaletskaya A. 10 other authors 1999; A cytomegalovirus-encoded mitochondria-localized inhibitor of apoptosis structurally unrelated to bcl-2. Proc Natl Acad Sci U S A 96:12536–12541 [CrossRef]
    [Google Scholar]
  22. Gooding C., Roberts G. C., Moreau G., Nadal Ginard B., Smith C. W. J. 1994; Smooth muscle-specific switching of alpha-tropomyosin mutually exclusive exon selection by specific inhibition of the strong default exon. EMBO J 13:3861–3872
    [Google Scholar]
  23. Gooding C., Kemp P., Smith C. W. J. 2003; A novel polypyrimidine tract binding protein paralog expressed in smooth muscle cells. J Biol Chem 278:15201–15207 [CrossRef]
    [Google Scholar]
  24. Graveley B. R. 2000; Sorting out the complexity of SR protein functions. RNA 6:1197–1211 [CrossRef]
    [Google Scholar]
  25. Gunderson S. I., Polycarpou-Schwarz M., Mattaj I. W. 1998; U1 snRNP inhibits pre-mRNA polyadenylation through a direct interaction between U1 70K and poly(A) polymerase. Mol Cell 1:255–264 [CrossRef]
    [Google Scholar]
  26. Hayajneh W. A., Colberg-Poley A. M., Skaletskaya A., Bartle L. M., Lesperance M. M., Contopoulos-Ioannidis D. G., Kedersha N. L., Goldmacher V. S. 2001a; The sequence and antiapoptotic functional domains of the human cytomegalovirus UL37 exon 1 immediate early protein are conserved in multiple primary strains. Virology 279:233–240 [CrossRef]
    [Google Scholar]
  27. Hayajneh W. A., Contopoulos-Ioannidis D. G., Lesperance M. M., Venegas A., Colberg-Poley A. M. 2001b; The carboxyl terminus of the human cytomegalovirus UL37 immediate-early glycoprotein is conserved in primary strains and is important for transactivation. J Gen Virol 82:1569–1579
    [Google Scholar]
  28. Huang T.-S., Nilsson C. E., Punga T., Akusjarvi G. 2002; Functional inactivation of the SR family of splicing factors during a vaccinia virus infection. EMBO Rep 3:1088–1093 [CrossRef]
    [Google Scholar]
  29. Jamison S. F., Pasman Z., Wang J., Will C., Lührmann R., Manley J. L., Garcia-Blanco M. A. 1995; U1 snRNP-ASF/SF2 interaction and 5′ splice site recognition: characterization of required elements. Nucleic Acids Res 23:3260–3267 [CrossRef]
    [Google Scholar]
  30. Kanopka A., Muhlemann O., Akusjarvi G. 1996; Inhibition by SR proteins of splicing of a regulated adenovirus pre-mRNA. Nature 381:535–538 [CrossRef]
    [Google Scholar]
  31. Kanopka A., Muhlemann O., Petersen-Mahrt S., Estmer C., Ohrmalm C., Akusjarvi G. 1998; Regulation of adenovirus alternative RNA splicing by dephosphorylation of SR proteins. Nature 393:185–187 [CrossRef]
    [Google Scholar]
  32. Kouzarides T., Bankier A. T., Satchwell S. C., Preddy E., Barrell B. G. 1988; An immediate early gene of human cytomegalovirus encodes a potential membrane glycoprotein. Virology 165:151–164 [CrossRef]
    [Google Scholar]
  33. Lee M., Xiao J., Haghjoo E., Zhan X., Abenes G., Tuong T., Dunn W., Liu F. 2000; Murine cytomegalovirus containing a mutation at open reading frame M37 is severely attenuated in growth and virulence in vivo. J Virol 74:11099–11107 [CrossRef]
    [Google Scholar]
  34. Liu X., Mayeda A., Tao M., Zheng Z.-M. 2003; Exonic splicing enhancer-dependent selection of the bovine papillomavirus type 1 nucleotide 3225 3′ splice site can be rescued in a cell lacking splicing factor ASF/SF2 through activation of the phosphatidylinositol 3-kinase/Akt pathway. J Virol 77:2105–2115 [CrossRef]
    [Google Scholar]
  35. MacDonald C., Wilusz J., Shenk T. 1994; The 64-kilodalton subunit of CstF polyadenylation factor binds to pre-mRNAs downstream of the cleavage site and influences cleavage site location. Mol Cell Biol 14:6647–6654
    [Google Scholar]
  36. Markovtsov V., Nikolic J. M., Goldman J. A., Turck C. W., Chou M.-Y., Black D. L. 2000; Cooperative assembly of an hnRNP complex induced by a tissue-specific homolog of polypyrimidine tract binding protein. Mol Cell Biol 20:7463–7479 [CrossRef]
    [Google Scholar]
  37. Mavinakere M. S., Colberg-Poley A. M. 2004a; Dual targeting of the human cytomegalovirus UL37 exon 1 protein during permissive infection. J Gen Virol 85:323–329 [CrossRef]
    [Google Scholar]
  38. Mavinakere M. S., Colberg-Poley A. M. 2004b; Internal cleavage of the human cytomegalovirus UL37 immediate early glycoprotein and divergent trafficking of its proteolytic fragments. J Gen Virol 85:1989–1994 [CrossRef]
    [Google Scholar]
  39. Moreira A., Takagaki Y., Brackenridge S., Wollerton M., Manley J. L., Proudfoot N. J. 1998; The upstream sequence element of the C2 complement poly(A) signal activates mRNA 3′ end formation by two distinct mechanisms. Genes Dev 12:2522–2534 [CrossRef]
    [Google Scholar]
  40. Murthy K. G., Manley J. L. 1992; Characterization of a multisubunit cleavage polyadenylation specificity factor from calf thymus. J Biol Chem 267:14804–14811
    [Google Scholar]
  41. Niwa M., MacDonald C. C., Berget S. M. 1992; Are vertebrate exons scanned during splice-site selection?. Nature 360:277–280 [CrossRef]
    [Google Scholar]
  42. Pérez I., Lin C. H., McAfee J. G., Patton J. G. 1997; Mutation of PTB binding sites causes misregulation of alternative 3′ splice site selection in vivo. RNA 3:764–778
    [Google Scholar]
  43. Pérez Cañadillas J. M., Varani G. 2003; Recognition of GU-rich polyadenylation regulatory elements by human CstF-64 protein. EMBO J 22:2821–2830 [CrossRef]
    [Google Scholar]
  44. Poncet D., Larochette N., Pauleau A.-L. 12 other authors 2004; An anti-apoptotic viral protein that recruits Bax to mitochondria. J Biol Chem 279:22605–22614 [CrossRef]
    [Google Scholar]
  45. Rambhatla L., Chiu C.-P., Glickman R. D., Rowe-Rendelman C. 2002; In vitro differentiation capacity of telomerase immortalized human RPE cells. Invest Ophthalmol Vis Sci 43:1622–1630
    [Google Scholar]
  46. Ryan K., Calvo O., Manley J. L. 2004; Evidence that polyadenylation factor CPSF-73 is the mRNA 3′ processing endonuclease. RNA 10:565–573 [CrossRef]
    [Google Scholar]
  47. Sciabica K. S., Dai Q. J., Sandri-Goldin R. M. 2003; ICP27 interacts with SRPK1 to mediate HSV splicing inhibition by altering SR protein phosphorylation. EMBO J 22:1608–1619 [CrossRef]
    [Google Scholar]
  48. Shen H., Kan J. L. C., Ghigna C., Biamonti G., Green M. R. 2004; A single polypyrimidine tract protein (PTB) binding site mediates splicing inhibition at mouse IgM exons M1 and M2. RNA 10:787–794 [CrossRef]
    [Google Scholar]
  49. Staknis D., Reed R. 1994; SR proteins promote the first specific recognition of pre-mRNA and are present together with the U1 small nuclear ribonucleoprotein particle in a general splicing enhancer complex. Mol Cell Biol 14:7670–7682
    [Google Scholar]
  50. Su Y., Testaverde J. R., Davis C. N., Hayajneh W. A., Adair R., Colberg-Poley A. M. 2003a; Human cytomegalovirus UL37 immediate early target minigene RNAs are accurately spliced and polyadenylated. J Gen Virol 84:29–39 [CrossRef]
    [Google Scholar]
  51. Su Y., Adair R., Davis C. N., DiFronzo N. L., Colberg-Poley A. M. 2003b; Convergence of RNA cis elements and cellular polyadenylation factors in the regulation of human cytomegalovirus UL37 exon 1 unspliced RNA production. J Virol 77:12729–12741 [CrossRef]
    [Google Scholar]
  52. Takagaki Y., Manley J. L. 1997; RNA recognition by the human polyadenylation factor CstF. Mol Cell Biol 17:3907–3914
    [Google Scholar]
  53. Tenney D. J., Colberg-Poley A. M. 1991; Expression of the human cytomegalovirus UL36–38 immediate early region during permissive infection. Virology 182:199–210 [CrossRef]
    [Google Scholar]
  54. Venkatramana M., Ray P. S., Chadda A., Das S. 2003; A 25 kDa cleavage product of polypyrimidine tract binding protein (PTB) present in mouse tissues prevents PTB binding to the 5′ untranslated region and inhibits translation of hepatitis A virus RNA. Virus Res 98:141–149 [CrossRef]
    [Google Scholar]
  55. Wagner E. J., Garcia-Blanco M. A. 2002; RNAi-mediated PTB depletion leads to enhanced exon definition. Mol Cell 10:943–949 [CrossRef]
    [Google Scholar]
  56. Wang X., Huong S.-M., Chiu M. L., Raab-Traub N., Huang E.-S. 2003; Epidermal growth factor receptor is a cellular receptor for human cytomegalovirus. Nature 424:456–461 [CrossRef]
    [Google Scholar]
  57. Xiao S.-H., Manley J. L. 1998; Phosphorylation–dephosphorylation differentially affects activities of splicing factor ASF/SF2. EMBO J 17:6359–6367 [CrossRef]
    [Google Scholar]
  58. Yu D., Silva M. C., Shenk T. 2003; Functional map of human cytomegalovirus AD169 defined by global mutational analysis. Proc Natl Acad Sci U S A 100:12396–12401 [CrossRef]
    [Google Scholar]
  59. Zarkower D., Wickens M. 1988; A functionally redundant downstream sequence in SV40 late pre-mRNA is required for mRNA 3′ end formation and for assembly of a precleavage complex in vitro . J Biol Chem 263:5780–5788
    [Google Scholar]
  60. Zhao J., Hyman L., Moore C. 1999; Formation of mRNA 3′ ends in eukaryotes: mechanism, regulation and interrelationships with other steps in mRNA synthesis. Microbiol Mol Biol Rev 63:405–445
    [Google Scholar]
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