Search tips
Search criteria 


Logo of narLink to Publisher's site
Nucleic Acids Res. 1997 March 15; 25(6): 1117–1122.
PMCID: PMC146575

A highly conserved nucleotide in the Alu domain of SRP RNA mediates translation arrest through high affinity binding to SRP9/14.


Binding of the signal recognition particle (SRP) to signal sequences during translation leads to an inhibition of polypeptide elongation known as translation arrest. The arrest activity is mediated by a discrete domain comprised of the Alu portion of SRP RNA and a 9 and 14 kDa polypeptide heterodimer (SRP9/14). Although very few nucleotides in SRP RNA are conserved throughout evolution, the remarkable conservation of G24, which resides in the region of SRP9/14 interaction, suggests that it is essential for translation arrest. To understand the functional significance of the G24 residue, we made single base substitutions in SRP RNA at this position and analyzed the ability of the mutants to bind SRP9/14 and to reconstitute functional SRPs. Mutation of G24 to C reduced binding to SRP9/14 by at least 50-fold, whereas mutation to A and U reduced binding approximately 2- and 5-fold respectively. The mutant RNAs could nevertheless assemble into SRPs at high subunit concentrations. SRPs reconstituted with mutant RNAs were not significantly defective in translation arrest assays, indicating that the conserved guanosine does not interact directly with the translational machinery. Taken together, these results demonstrate that G24 plays an important role in the translation arrest function of SRP by mediating high affinity binding of SRP9/14.

Full Text

The Full Text of this article is available as a PDF (94K).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Walter P, Johnson AE. Signal sequence recognition and protein targeting to the endoplasmic reticulum membrane. Annu Rev Cell Biol. 1994;10:87–119. [PubMed]
  • Görlich D, Hartmann E, Prehn S, Rapoport TA. A protein of the endoplasmic reticulum involved early in polypeptide translocation. Nature. 1992 May 7;357(6373):47–52. [PubMed]
  • Görlich D, Rapoport TA. Protein translocation into proteoliposomes reconstituted from purified components of the endoplasmic reticulum membrane. Cell. 1993 Nov 19;75(4):615–630. [PubMed]
  • Poritz MA, Bernstein HD, Strub K, Zopf D, Wilhelm H, Walter P. An E. coli ribonucleoprotein containing 4.5S RNA resembles mammalian signal recognition particle. Science. 1990 Nov 23;250(4984):1111–1117. [PubMed]
  • Ribes V, Römisch K, Giner A, Dobberstein B, Tollervey D. E. coli 4.5S RNA is part of a ribonucleoprotein particle that has properties related to signal recognition particle. Cell. 1990 Nov 2;63(3):591–600. [PubMed]
  • Althoff S, Selinger D, Wise JA. Molecular evolution of SRP cycle components: functional implications. Nucleic Acids Res. 1994 Jun 11;22(11):1933–1947. [PMC free article] [PubMed]
  • Schatz G, Dobberstein B. Common principles of protein translocation across membranes. Science. 1996 Mar 15;271(5255):1519–1526. [PubMed]
  • Siegel V, Walter P. Each of the activities of signal recognition particle (SRP) is contained within a distinct domain: analysis of biochemical mutants of SRP. Cell. 1988 Jan 15;52(1):39–49. [PubMed]
  • Kurzchalia TV, Wiedmann M, Girshovich AS, Bochkareva ES, Bielka H, Rapoport TA. The signal sequence of nascent preprolactin interacts with the 54K polypeptide of the signal recognition particle. Nature. 1986 Apr 17;320(6063):634–636. [PubMed]
  • Krieg UC, Walter P, Johnson AE. Photocrosslinking of the signal sequence of nascent preprolactin to the 54-kilodalton polypeptide of the signal recognition particle. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8604–8608. [PubMed]
  • Siegel V, Walter P. Removal of the Alu structural domain from signal recognition particle leaves its protein translocation activity intact. Nature. 1986 Mar 6;320(6057):81–84. [PubMed]
  • Gundelfinger ED, Krause E, Melli M, Dobberstein B. The organization of the 7SL RNA in the signal recognition particle. Nucleic Acids Res. 1983 Nov 11;11(21):7363–7374. [PMC free article] [PubMed]
  • Larsen N, Zwieb C. SRP-RNA sequence alignment and secondary structure. Nucleic Acids Res. 1991 Jan 25;19(2):209–215. [PMC free article] [PubMed]
  • Siegel V, Walter P. Binding sites of the 19-kDa and 68/72-kDa signal recognition particle (SRP) proteins on SRP RNA as determined in protein-RNA "footprinting". Proc Natl Acad Sci U S A. 1988 Mar;85(6):1801–1805. [PubMed]
  • Walter P, Lingappa VR. Mechanism of protein translocation across the endoplasmic reticulum membrane. Annu Rev Cell Biol. 1986;2:499–516. [PubMed]
  • Wood H, Luirink J, Tollervey D. Evolutionary conserved nucleotides within the E.coli 4.5S RNA are required for association with P48 in vitro and for optimal function in vivo. Nucleic Acids Res. 1992 Nov 25;20(22):5919–5925. [PMC free article] [PubMed]
  • Strub K, Moss J, Walter P. Binding sites of the 9- and 14-kilodalton heterodimeric protein subunit of the signal recognition particle (SRP) are contained exclusively in the Alu domain of SRP RNA and contain a sequence motif that is conserved in evolution. Mol Cell Biol. 1991 Aug;11(8):3949–3959. [PMC free article] [PubMed]
  • Liao X, Selinger D, Althoff S, Chiang A, Hamilton D, Ma M, Wise JA. Random mutagenesis of Schizosaccharomyces pombe SRP RNA: lethal and conditional lesions cluster in presumptive protein binding sites. Nucleic Acids Res. 1992 Apr 11;20(7):1607–1615. [PMC free article] [PubMed]
  • Selinger D, Brennwald P, Althoff S, Reich C, Hann B, Walter P, Wise JA. Genetic and biochemical analysis of the fission yeast ribonucleoprotein particle containing a homolog of Srp54p. Nucleic Acids Res. 1994 Jul 11;22(13):2557–2567. [PMC free article] [PubMed]
  • Andreazzoli M, Gerbi SA. Changes in 7SL RNA conformation during the signal recognition particle cycle. EMBO J. 1991 Apr;10(4):767–777. [PubMed]
  • Bovia F, Fornallaz M, Leffers H, Strub K. The SRP9/14 subunit of the signal recognition particle (SRP) is present in more than 20-fold excess over SRP in primate cells and exists primarily free but also in complex with small cytoplasmic Alu RNAs. Mol Biol Cell. 1995 Apr;6(4):471–484. [PMC free article] [PubMed]
  • Chang DY, Sasaki-Tozawa N, Green LK, Maraia RJ. A trinucleotide repeat-associated increase in the level of Alu RNA-binding protein occurred during the same period as the major Alu amplification that accompanied anthropoid evolution. Mol Cell Biol. 1995 Apr;15(4):2109–2116. [PMC free article] [PubMed]
  • Chang DY, Nelson B, Bilyeu T, Hsu K, Darlington GJ, Maraia RJ. A human Alu RNA-binding protein whose expression is associated with accumulation of small cytoplasmic Alu RNA. Mol Cell Biol. 1994 Jun;14(6):3949–3959. [PMC free article] [PubMed]
  • Chang DY, Hsu K, Maraia RJ. Monomeric scAlu and nascent dimeric Alu RNAs induced by adenovirus are assembled into SRP9/14-containing RNPs in HeLa cells. Nucleic Acids Res. 1996 Nov 1;24(21):4165–4170. [PMC free article] [PubMed]
  • Maraia RJ. The subset of mouse B1 (Alu-equivalent) sequences expressed as small processed cytoplasmic transcripts. Nucleic Acids Res. 1991 Oct 25;19(20):5695–5702. [PMC free article] [PubMed]
  • Maraia RJ, Driscoll CT, Bilyeu T, Hsu K, Darlington GJ. Multiple dispersed loci produce small cytoplasmic Alu RNA. Mol Cell Biol. 1993 Jul;13(7):4233–4241. [PMC free article] [PubMed]
  • Sinnett D, Richer C, Deragon JM, Labuda D. Alu RNA secondary structure consists of two independent 7 SL RNA-like folding units. J Biol Chem. 1991 May 15;266(14):8675–8678. [PubMed]
  • Hsu K, Chang DY, Maraia RJ. Human signal recognition particle (SRP) Alu-associated protein also binds Alu interspersed repeat sequence RNAs. Characterization of human SRP9. J Biol Chem. 1995 Apr 28;270(17):10179–10186. [PubMed]
  • Chang DY, Maraia RJ. A cellular protein binds B1 and Alu small cytoplasmic RNAs in vitro. J Biol Chem. 1993 Mar 25;268(9):6423–6428. [PubMed]
  • Ullu E, Weiner AM. Upstream sequences modulate the internal promoter of the human 7SL RNA gene. Nature. 318(6044):371–374. [PubMed]
  • Zopf D, Bernstein HD, Walter P. GTPase domain of the 54-kD subunit of the mammalian signal recognition particle is required for protein translocation but not for signal sequence binding. J Cell Biol. 1993 Mar;120(5):1113–1121. [PMC free article] [PubMed]
  • Walter P, Blobel G. Disassembly and reconstitution of signal recognition particle. Cell. 1983 Sep;34(2):525–533. [PubMed]
  • Walter P, Blobel G. Subcellular distribution of signal recognition particle and 7SL-RNA determined with polypeptide-specific antibodies and complementary DNA probe. J Cell Biol. 1983 Dec;97(6):1693–1699. [PMC free article] [PubMed]
  • Morrissey JH. Silver stain for proteins in polyacrylamide gels: a modified procedure with enhanced uniform sensitivity. Anal Biochem. 1981 Nov 1;117(2):307–310. [PubMed]
  • Strub K, Walter P. Assembly of the Alu domain of the signal recognition particle (SRP): dimerization of the two protein components is required for efficient binding to SRP RNA. Mol Cell Biol. 1990 Feb;10(2):777–784. [PMC free article] [PubMed]
  • Siegel V, Walter P. Elongation arrest is not a prerequisite for secretory protein translocation across the microsomal membrane. J Cell Biol. 1985 Jun;100(6):1913–1921. [PMC free article] [PubMed]
  • Murray AW, Solomon MJ, Kirschner MW. The role of cyclin synthesis and degradation in the control of maturation promoting factor activity. Nature. 1989 May 25;339(6222):280–286. [PubMed]
  • Janiak F, Walter P, Johnson AE. Fluorescence-detected assembly of the signal recognition particle: binding of the two SRP protein heterodimers to SRP RNA is noncooperative. Biochemistry. 1992 Jun 30;31(25):5830–5840. [PubMed]
  • Carey J. Gel retardation. Methods Enzymol. 1991;208:103–117. [PubMed]
  • Jaeger JA, Turner DH, Zuker M. Improved predictions of secondary structures for RNA. Proc Natl Acad Sci U S A. 1989 Oct;86(20):7706–7710. [PubMed]
  • Siegel V, Walter P. The affinity of signal recognition particle for presecretory proteins is dependent on nascent chain length. EMBO J. 1988 Jun;7(6):1769–1775. [PubMed]
  • Powers T, Walter P. The nascent polypeptide-associated complex modulates interactions between the signal recognition particle and the ribosome. Curr Biol. 1996 Mar 1;6(3):331–338. [PubMed]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press