Two biological questions are under investigation in my laboratory. We study the mechanism of L1 retrotransposition and its control in genetic and evolutionary time, as well as in the molecular events that determine the phenotype of a hibernating mammal. Answers to questions in both projects impact our understanding of genome evolution and the relationship between genotype and phenotype.
LINE-1, or L1, is a retrotransposon that populates the genomes of all mammals; it is the most significant dynamic force acting upon the human genome. Typically, L1 sequences comprise between 10 and 30% of a genome and are interspersed throughout all chromosomes. This highly repetitive L1 DNA family is comprised of a small number of active transposable elements and a large number of their truncated, defective pseudogene progeny. Movement into new sites in nuclear DNA occurs via reverse transcription of an RNA intermediate by a unique mechanism known as target primed reverse transcription, or tprt. Our long-range goal is to understand the retrotransposition process in detail, including the biochemical intermediates involved, as well the control of retrotransposition in genetic and evolutionary time.
LINE-1 retrotransposition begins with transcription of a full-length, sense-strand L1 RNA and requires two L1-encoded polypeptides. These proteins also catalyze the reverse transcription and integration of SINEs (short interspersed repeated sequences) and processed pseudogenes, thereby amplifying the effects of LINE-1 in mammalian genome dynamics. Specifically, our ongoing experiments are designed to enhance understanding of the structure and function of the two essential L1-encoded proteins, ORF1p and ORF2p, and the details of their interaction with each other as well as with both RNA and DNA. We are also working to define the cis- and trans-acting components of translational control that operate upon the two L1-encoded proteins in the naturally-occurring dicistronic L1 mRNA, and how the L1 RNA transitions from its role as a translation template to that of a reverse transcriptase template.
LINEs in mammals belong to a much larger family of mobile elements that are widely distributed among eucaryotes and known as the non-LTR retrotransposons. These elements have been replicating in eucaryotic genomes since the origin of eucaryotes, in some cases remaining modest in copy number, other cases amplifying to very high copy numbers (e.g., humans), and some cases becoming extinct. These dynamics reinforce the idea of the genome as an ecosystem and offer unique opportunities for gaining insight into the competition for survival among rival forces within the genome.
We also study hibernation in mammals because two aspects of the hibernating phenotype offer extraordinary potential to make an enormous impact on medicine: first, hibernators orchestrate a profound protection against cell and organ damage associated with ischemia-reperfusion injury during their hibernation season; second, these animals reversibly drop their metabolism to ~5% of basal metabolic rate. We hope that by defining the mechanisms used in this natural model will reveal novel pathways and reveal drugable strategies to mimic these responses in patients.
Mammals in deep hibernation exhibit profound reductions in their basal metabolic, heart and respiratory rates, along with core body temperature; the identical constellation of physiological parameters is inevitably lethal in a non-hibernating species such as human. At present, very little is known about the mechanisms that permit hibernators to achieve, maintain and survive these otherwise overwhelming physiological changes at the molecular level. Hibernating species are widely interspersed among monotremes, marsupials and placentals. This widespread phylogenetic distribution of hibernators suggests that hibernation is an ancestral trait that has been lost numerous times in mammalian evolution, and implies that all mammals share the genes that specify and permit the hibernating phenotype. Thus, our long term goal for this project is to understand how the mammalian genotype is expressed as a hibernating phenotype in some, but not all mammals. This project has two components: the first seeks to identify differentially expressed genes at the mRNA and protein level; the second tests predictions that derive from the hypothesis that hibernators must rewarm periodically in order to replenish gene products that are catabolized, but not synthesized, at the low body temperatures of torpor.
We have implemented various approaches over the years to identify genes that are differentially expressed at the steady-state mRNA level. These have provided some interesting candidates for further work, but it is clear that present efforts have merely scratched the surface of what remains to be discovered. We are presently exploiting new proteomics screening methods to quantify and identify proteins, metabolites and mRNAs that are differentially present or modified during the season of hibernation because these are likely responsible for the protection seen in the winter animals. Other experiments address the molecular mechanisms that control the reversible suppression of various biochemical processes during the torpor-arousal cycles of hibernation. The application of both protection and reversible hypometabolism in humans has tremendous potential to improve outcomes for victims of cardiac arrest, stroke and trauma.
Martin, S.L. (2008) Mammalian hibernation: a naturally reversible model for insulin resistance in man? Diab Vasc Dis Res 5, 76-81. PMID: 18537093
Nelson, C.J., Otis, J.P., Martin, S.L., Carey, H.V. (2009) Analysis of the hibernation cycle using LC-MS based metabolomics in ground squirrel liver. Physiol Genomics 37, 43-51. PMID: 19106184
Martin, S.L. (2009) Developmental biology: Jumping-gene roulette. Nature 460, 1087-1088.
Martin, S.L., Branciforte, D., Keller, D. and Bain, D.L. (2003) Novel trimeric structure for an essential protein in L1 retrotransposition, Proc. Natl. Acad. Sci., USA 100, 13815-13820.
van Breukelen, F., Sonenberg, N. and Martin S.L. (2004) Seasonal and state dependent changes of eIF4E and 4E-BP1 during mammalian hibernation: implications for the control of translation during torpor, Am J Physiol Regul Integr Comp Physiol. 287, R349-R353
Epperson, L.E., Dahl, T. and Martin, S.L. (2004) Quantitative analysis of liver protein expression during hibernation in golden-mantled ground squirrel, Mol. Cell. Proteomics. 3, 920-933.
Martin, S.L., Cruceanu, M., Branciforte, D., Li, P. W.-l., Kwok, S.C., Hodges, R.S. and Williams, M.C. (2005) LINE-1 retrotransposition requires the nucleic acid chaperone activity of the ORF1 protein, J. Mol. Biol. 348, 549-561.
Williams, D.R., Epperson, L.E., Li, W., Hughes, M.A., Taylor, R., Rogers, J., Martin, S.L., Cossins, A.R., Gracey, A.Y. (2005) The seasonally hibernating phenotype assessed through transcript screening. Physiol. Genomics. 24, 13-22.
Basame, S., Li, P.W-l., Howard, G., Branciforte, D., Keller, D., Martin, S.L. (2006) Spatial assembly and RNA binding stoichiometry of a protein essential for LINE-1 retrotransposition. J. Mol. Biol. 357, 351-7.
Li, P.W.-l., Li, J., Timmerman, S., Krushel, L., Martin, S. L. (2006) The dicistronic RNA from the mouse LINE-1 retrotransposon contains an internal ribosome entry site upstream of each ORF: implications for retrotransposition. Nuc. Acids Res. 34, 853-64.
Serkova, N.J., Rose, J.C., Epperson, L.E., Carey, H.V. and Martin, S.L. (2007) Quantitative analysis of liver metabolites in three stages of the circannual hibernation cycle in 13-lined ground squirrels by NMR. Physiol. Genomics, 31, 15-24.
Januszyk, K., Li, P.W.-l., Villareal, V., Branciforte, D., Wu, H., Xie, Y., Feigon, J., Loo, J.A., Martin, S.L. and Clubb,R.T. (2007) Identification and solution structure of a highly conserved C-terminal domain required for the retrotransposition of Long Interspersed Nuclear Element-1, J Biol Chem. 282, 24893-904.
Martin, S.L., Epperson, L.E., Rose, J.C., Kurtz, C.C., Ané, C. and Carey, H.V. (2008) Proteomic analysis of the winter-protected phenotype of hibernating ground squirrel intestine. Am J Physiol Regul Integr Comp Physiol 295, R316-28.
Soper, S.F.C., van der Heijden, G.W., Hardiman, T.C., Goodheart, M., Martin, S.L., de Boer, P. and Bortvin, A. (2008) Mouse maelstrom, a component of nuage, is essential for spermatogenesis and transposon repression in meiosis, Dev. Cell 15, 285-297.
Martin, S.L., Bushman, D., Wang, F., Li, P. W.-L., Walker, A., Cummiskey, J., Branciforte, D. and Williams, M.C. (2008) A single amino acid substitution in ORF1 dramatically decreases L1 retrotransposition and provides insight into nucleic acid chaperone activity. Nuc.Acids. Res., 18, 5845-5854.