(1) Ribosomal RNA (rRNA) Structure
Our lab sequenced the first metazoan 28S rRNA. We showed that certain sequences are highly conserved, even between bacterial and eukaryotic rRNA, thus defining functionally important sites of rRNA. Furthermore, we found that the secondary structure of rRNA is highly conserved in all domains of life. We discovered “expansion segments” that are inserted into the conserved core secondary structure; they are variable in length and sequence in eukaryotic rRNA. Expansion segments have proven useful for phylogenetic analysis and are the focus of several recent cryo-EM and x-ray crystallography studies by other groups. Our recent bioinformatic analysis of rRNA sequences from all domains of life has identified domain-specific sequence signatures (e.g., conserved nucleotide elements in eukaryotes but not in bacteria or archaea). These recent investigations opens the possibility for a new class of antibiotics targeted against conserved nucleotide elements.
Selected Papers
Gourse RL, Thurlow DL, Gerbi SA and Zimmermann RA (1981) Specific binding of a prokaryotic ribosomal protein to a eukaryotic ribosomal RNA: implications for evolution and autoregulation. Proc. Nat. Acad. Sci. 78: 2722-2726. PMID: 6265904; PMCID: PMC326536.
Stebbins-Boaz B and Gerbi SA (1991) Structural analysis of the peptidyl transferase region in ribosomal RNA of the eukaryote Xenopus laevis. J. Mol. Biol.217: 93-112. PMID: 1988683.
Gerbi SA (1996) Expansion segments: regions of variable size that interrupt the universal core secondary structure of ribosomal RNA. In Ribosomal RNA: Structure, Evolution, Processing and Function in Protein Synthesis, eds: R.A. Zimmermann and A.E. Dahlberg. Telford – CRC Press, Boca Raton, FL. pp. 71-87.
Doris SM, Smith DR, Beamesderfer JN, Raphael BJ, Nathanson JA and Gerbi SA (2015) Universal and domain-specific sequences in 23S-28S ribosomal RNA identified by computational phylogenetics. RNA 21: 1719-1730. PMID: 26283689; PMCID: PMC4574749.
(2) Ribosomal RNA (rRNA) Processing
Using oocytes from the frog Xenopus to study ribosome biogenesis, we were the first to show a role in vivo for any small nucleolar RNA (snoRNA). Specifically, by injection of antisense oligonucleotides into Xenopus oocytes we demonstrated that U3 snoRNA plays a role in rRNA processing. We found that U3 influences the order of cleavages to process pre-rRNA and it guides cleavage of pre-rRNA. Moreover, U3 snoRNA may act as a chaperone to prevent premature pseudoknot formation in 18S rRNA, and is a molecular bridge to draw together the 5′ and 3′ ends of 18S rRNA in the precursor. U3 snoRNA docks on pre-rRNA by base-pairing interactions that are species-specific, and may allow us to design a new class of antibiotics against eukaryotic pathogens. We found that rRNA processing in Xenopus has several features that are distinct from rRNA processing in yeast and provides a foundation for current studies by other groups on rRNA processing in humans.
Selected Papers
Savino R and Gerbi SA (1990) In vivo disruption of Xenopus U3 snRNA affects ribosomal RNA processing. EMBO J.9: 2289-2308. PMID: 2357971; PMCID: PMC551956.
Borovjagin AV and Gerbi SA (2000) The spacing between functional cis-elements of U3 snoRNA is critical for rRNA processing. J. Mol. Biol. 300: 57-74. PMID: 10864498.
Borovjagin AV and Gerbi SA (2001) Xenopus U3 snoRNA GAC-Box A’ and Box A sequences play distinct functional roles in rRNA processing. Mol. Cell. Biol. 21: 6210-6221. PMID: 11509664; PMCID: PMC87338.
Borovjagin AV and Gerbi SA (2004) Xenopus U3 snoRNA docks on pre-rRNA through a novel base-pairing interaction. RNA 10: 942-953. PMID: 15146078; PMCID: PMC1370586.
(3) Nuclear Localization of Small RNAs
We have developed methodology to allow in vivo studies on localization of small RNAs in nuclei. The technique involves injection into Xenopus oocytes of a fluorescently tagged small RNA to follow its nuclear localization by fluorescence microscopy. With this approach, we showed that conserved boxes C/D or boxes H/ACA are the nucleolar localization elements (NoLEs) in snoRNAs. We also showed that snRNAs of the spliceosome traffic through the nucleolus. Our lab identified NoLEs for U4 and U6 snRNAs and the elements needed for Cajal body localization.
Selected Papers
Lange TS, Borovjagin A, Maxwell ES and Gerbi SA (1998) Conserved Boxes C and D are essential nucleolar localization elements of U8 and U14 snoRNAs. EMBO J. 17: 3176-3187. PMID: 9606199; PMCID: PMC1170656.
Lange TS, Ezrokhi M, Amaldi F and Gerbi SA (1999) Box H and Box ACA are nucleolar localization elements of U17 snoRNA. Mol Biol. Cell 10: 3877-3890. PMID: 10564278; PMCID: PMC25686.
Gerbi SA and Lange TS (2002) All snRNAs of the (U4/U6.U5) tri-snRNP localize to nucleoli; identification of the NoLE of U6 snRNA. Mol. Biol. Cell 13: 3123-3137. PMID: 12221120; PMCID: PMC124147.
Gerbi SA, Borovjagin AV, Odreman FE and Lange TS (2003) U4 snRNA nucleolar localization requires the NHPX/15.5 KD protein binding site but not Sm protein or U6 snRNA association. J. Cell Biol. 162: 821-832. PMID: 12939253; PMCID: PMC2172826.