How messenger RNAs (mRNAs) code for proteins is determined by the DNA sequence that encodes the mRNA and post-transcriptional events, known as mRNA processing, that rearrange the existing RNA sequence (mRNA splicing) or change it (mRNA editing). My lab studies mRNA editing as it occurs in liver, but there are many tissues in humans and animals in which different mRNAs are changed by editing in ways that are essential for normal tissue function and dysfunction in disease.

      One area of active research involves computational modeling and screening of DNA microarrays for mRNAs that may serve as new editing substrates. This is an important question to answer as editing changes the ability of mRNAs to code for proteins, consequently the existence of these proteins cannot be predicted directly from genomic information. mRNA editing is an essential part of the new field of ribonomics, which contributes vital information on how mRNAs are processed and therefore, in terms of the field of proteonomics, provides solutions to questions in metabolism, therapeutics and toxicology. (For more details see: http://www.rna.ucla.edu/gordon2000/.)

      Apolipoprotein B coordinates lipid as VLDL and LDL and is translated from a 14.1 kb mRNA. ApoB mRNA can be translated as either of two apoB protein isoforms, depending on whether a cytidine at nucleotide position 6666 has been edited to a uridine. ApoB mRNA editing is tissue-specific and is subject to developmental, dietary and hormonal regulation. VLDL containing the apoB isoform synthesized form edited mRNA do not become LDL and hence, editing is associated with a reduction in this atherogenic risk factor.

       The ability to carry out RNA editing in vitro using tissue extracts and recombinant apoB RNA substrates has led to the discovery of the multimeric macromolecular assembly believed to be responsible for carrying out the editing reaction (the editosome). Specific protein-RNA interactions 3' of the editing site are necessary for site-specific editing and the assembly of the editosome. The current model is that 66 and 44 kDa proteins interact with the an 11 nucleotide motif, the 'mooring sequence' to form the editing site recognition complex. Interaction of a 27 kDa cytidine deaminase, 'APOBEC-1' with these RNA-protein complexes leads to alteration of the enzymes substrate specificity and editing of apoB mRNA.

       An additional complex of proteins containing a 240 kDa antigen has been proposed to facilitate the assembly of fully functional editosomes. RNA sequences 5' of the editing site enhance editing activity through unknown mechanisms. RNA foot-printing, site-directed mutagenesis and monoclonal and peptide-specific polyclonal antibody production are a few of the techniques being used to develop probes for studying the mechanism of apoB mRNA editing and its regulation. Genes for the editing factors are being cloned and sequenced in order to evaluate possibilities for transcriptional regulation and to engineer gene knock-out constructs for studies in tissue culture cells and eventually transgenic animals.

      Regulation of apoB mRNA editing is central to liver activity in terms of health and disease. Editing is regulated developmentally in a tissue-specific manner and can be significantly altered by diet, hormonal changes, and toxins such as ethanol. We are studying the regulation of editing at the level of cell signaling and trafficking of the editing factors between the cytoplasm and the nucleus (ultimately the intracellular site where mRNA editing takes place). Animal model systems and whole cell microinjections analyses are being used to determine the molecules and mechanisms involved in apoB mRNA editing.

My laboratory page.

Jin, X., Wu, H., Smith, H.C. 2007. APOBEC3G levels predict rates of progression to AIDS. Retrovirology 4:20-27.

Smith. H.C. 2007. Measuring editing activity and identifying C to U mRNA editing factors in cells and biochemical isolates. In Methods in Enzymology, RNA Editing & Modification (Gott, J., eds) Academic Press, NY. 424:387-416.

Smith, H. C. 2006. Editing Informational Content of Expressed DNA Sequences and Their Transcripts. In: The Implicit Genome, Edited by Lynn Helena Caporale, Oxford University Press.

Lehmann DM, Galloway CA, Macelrevey C, Sowden MP, Wedekind JE, Smith HC. 2007. Functional characterization of APOBEC-1 complementation factor phosphorylation sites. Biochim Biophys Acta. Mar;1773(3):408-1418.

Wedekind JE, Gillilan R, Janda A, Krucinska J, Salter JD, Bennett RP, Raina J, and Smith HC. 2006. Nanostructures of APOBEC3G support a hierarchical assembly model of high molecular mass ribonucleoprotein particles from dimeric subunits. J Biol Chem. Dec 15;281(50):38122-38126.

Lehmann, DM, Galloway, CA, Sowden, MP, and Smith, HC. 2006. Metabolic regulation of ApoB mRNA editing is associated with phosphorylation of APOBEC-1 complementation factor. Nucleic Acids Research, Vol. 34, No. 11 3299-3308.

Ichikawa, HT, Sowden, MP, ATorelli, AT, Bachl, J, Huang, P, Dance, GSC, Marr, SH, Robert, J, Wedekind, JE, Smith, HC, and Bottaro, A. 2006. Structural phylogenetic analysis of activation-induced deaminase function. J. Immunol. Jul 1;177(1):355-361

Jin, X., Brooks, A., Chen, H., Bennett, R. Reichman, R., and Smith, H.C. 2005. APOBEC3G-CEM15/hA3G mRNA Levels Associates Inversely with HIV Viremia.Ê J. Virol. 79:11513-11516.

Xie K, Sowden MP, Dance GS, Torelli AT, Smith HC, and Wedekind JE. 2004. The structure of a yeast RNA-editing deaminase provides insight into the fold and function of activation-induced deaminase and APOBEC-1. Proc Natl Acad Sci U S A. May 25;101(21):8114-8119.

Smith HC, Bottaro A, Sowden MP, Wedekind JE. 2004. Activation induced deaminase: the importance of being specific. Trends Genet. Jun;20(6):224-227. Review

Sowden MP, Lehmann DM, Lin X, Smith CO, and Smith HC. 2004. Identification of novel alternative splice variants of APOBEC-1 complementation factor with different capacities to support apolipoprotein B mRNA editing. J Biol Chem. Jan 2;279(1):197-206.

Wedekind JE, Dance GS, Sowden MP, and Smith HC. 2003. Messenger RNA editing in mammals: new members of the APOBEC family seeking roles in the family business. Trends Genet. Apr;19(4):207-216. Review.

Galloway CA, Sowden MP, and Smith HC. 2003. Increasing the yield of soluble recombinant protein expressed in E. coli by induction during late log phase. Biotechniques. Mar;34(3):524-526, 528, 530

Smith HC. 2002. A biochemistry course director’s perspectives on student learning and faculty teaching in lecture-based and problem-based learning curricula. Academic Medicine Dec;77(12 Pt 1):1189-1198.

Dance GS, Sowden MP, Cartegni L, Cooper E, Krainer AR, and Smith HC. 2002. Two proteins essential for apolipoprotein B mRNA editing are expressed from a single gene through alternative splicing.J Biol Chem Jan 28; [epub ahead of print].

Sowden MP, Ballatori N, Jensen KL, Reed LH, and Smith HC. 2002. The editosome for cytidine to uridine mRNA editing has a native complexity of 27S: Identification of intracellular domains containing active and inactive editing factors. J Cell Sci Mar 1;115(Pt 5):1027-1039.

Yang Y, Ballatori N, and Smith HC. 2002. Apolipoprotein B mRNA editing and the reduction in synthesis and secretion of the atherogenic risk factor, apolipoprotein B100 can be effectively targeted through TAT-mediated protein transduction. Mol Pharmacol Feb;61(2):269-276.

Giangreco A, Sowden MP, Mikityansky I, and Smith HC. 2001. Ethanol stimulates apolipoprotein B mRNA editing in the absence of de novo RNA or protein synthesis. Biochem Biophys Res Commun Dec 21;289(5):1162-1167.

Sowden MP and Smith HC. 2001. Commitment of apolipoprotein B RNA to the splicing pathway regulates cytidine-to-uridine editing-site utilization. Biochem J Nov 1;359(Pt 3):697-705.

Yang Y, Sowden MP, Yang Y, Smith HC. 2001. Intracellular Trafficking Determinants in APOBEC-1, the Catalytic Subunit for Cytidine to Uridine Editing of Apolipoprotein B mRNA. Exp Cell Res Jul 15;267(2):153-64.