Alan Weiner
Emeritus Professor of Biochemistry
BA 1968, Yale College
PhD 1973, Harvard University
Off.: J441
amweiner@u.washington.edu

Research

The role of CSB protein in Cockayne syndrome. Our major focus is to understand why defects in the human SWI/SNF chromatin remodeling protein known as CSB cause Cockayne syndrome (CS), a devastating childhood developmental disease characterized by neurological dysfunction, severe post-natal growth failure, and early death. The CSB protein is best understood as the central component of a DNA repair complex that performs transcription-coupled nucleotide excision repair (TC-NER), but despite continuing progress on the detailed biochemistry of TC-NER, a compelling connection between the DNA repair defect and CS disease has failed to emerge.

Our experimental evidence suggests instead that CS is not caused by a DNA repair defect (as generally assumed) but by a chromatin remodeling defect that deregulates the expression of many genes, somehow inducing a constitutive interferon-like response that resembles the early stages of Aicardi-Goutières syndrome (AGS). We are now collaborating with the Share and Care Network and the Luke O’Brien Foundation to determine whether CS newborns, infants, children, and young adults exhibit the same interferon-like response we observe in CS cell lines.

The CSB-PGBD3 fusion protein. While studying CSB, we accidentally discovered that the CSB gene actually generates two abundant proteins: intact CSB protein generated by default splicing of CSB exons 1-22, and a novel CSB-PGBD3 fusion protein in which CSB exons 1-5 are alternatively spliced in frame to a domesticated piggyBac transposase called PGBD3 which inserted into CSB intron 5 in marmosets over 43 million years ago. Conservation of the CSB-PGBD3 fusion protein for >43 My strongly suggests that it is advantageous in health, but the fusion protein may also contribute to the heterogeneous clinical presentation of CS because it continues to be expressed in some CS patients who lack functional CSB. We also found that the CSB-PGBD3 fusion protein binds to hundreds of specific genomic sites from which it regulates expression of nearby genes. Surprisingly, the fusion protein does not bind directly to DNA at these sites, but indirectly by protein/protein interactions (“tethering”) with transcriptional activators (c-Jun and AP1), transcriptional repressors and silencers (CTCF), and RNA polymerase II itself.

PGBD5: a neural-specific piggyBac transposase domesticated >500 Mya and conserved from cephalochordates to humans. We are characterizing PGBD5, the only other domesticated piggyBac element in the human genome. We’ve shown that (1) PGBD5 was first domesticated in the common ancestor of the cephalochordate Branchiostoma (aka lancelet or amphioxus) and vertebrates, and is conserved in all vertebrates including lamprey, but cannot be found in more basal urochordates, hemichordates, or echinoderms; (2) the lancelet, lamprey, and human PGBD5 genes are syntenic and orthologous as expected for a single domestication at the base of the vertebrate lineage; (3) although derived from an IS4-related transposase, PGBD5 protein is unlikely to retain enzymatic activity because the catalytic DDD motif is not conserved; (4) PGBD5 is preferentially expressed in certain granule cell lineages of the mammalian brain and central nervous system based on available mouse and human in situ hybridization data; (5) the human PGBD5 promoter and gene region is rich in bound regulatory factors including the neuron-restrictive silencer factors NRSF/REST and CoREST, as well as SIN3, KAP1, STAT3, and CTCF; and (6) despite preferential localization within the nucleus, PGBD5 protein is unlikely to bind DNA or chromatin as neither DNase I digestion nor high salt extraction release PGBD5 from fractionated mouse brain nuclei. We speculate that the neural-specific PGBD5 transposase was domesticated >500 My after cephalochordates and vertebrates diverged from urocordates, and that PGBD5 may have played a role in the evolution of a primitive deuterosome neural network into a centralized nervous system (CNS).

Publications:

Showing most recent results from Scopus…

  1. Pavelitz, T. et al. PGBD5: A neural-specific intron-containing piggyBac transposase domesticated over 500 million years ago and conserved from cephalochordates to humans Mobile DNA 2013-11-01; :23. [PubMed]
  2. Weiner, A.M. et al. What role (if any) does the highly conserved CSB-PGBD3 fusion protein play in Cockayne syndrome? Mechanisms of Ageing and Development 2013-05-01; 134(5-6):225-233. [PubMed]
  3. Gray, L.T. et al. Tethering of the Conserved piggyBac Transposase Fusion Protein CSB-PGBD3 to Chromosomal AP-1 Proteins Regulates Expression of Nearby Genes in Humans PLoS Genetics 2012-09-01; 8(9). [PubMed]
  4. Bailey, A.D. et al. The conserved Cockayne syndrome B-piggyBac fusion protein (CSB-PGBD3) affects DNA repair and induces both interferon-like and innate antiviral responses in CSB-null cells DNA Repair 2012-05-01; 11(5):488-501. [PubMed]
  5. Gray, L.T. et al. Ubiquitin Recognition by the Cockayne Syndrome Group B Protein: Binding Will Set You Free Molecular Cell 2010-06-11; 38(5):621-622. [PubMed]
  6. Cho, H.D. et al. On the role of a conserved, potentially helix-breaking residue in the tRNA-binding a-helix of archaeal CCA-adding enzymes RNA 2008-07-01; 14(7):1284-1289. [PubMed]
  7. Pavelitz, T. et al. Human U2 snRNA genes exhibit a persistently open transcriptional state and promoter disassembly at metaphase Molecular and Cellular Biology 2008-06-01; 28(11):3573-3588. [PubMed]
  8. Newman, J.C. et al. An abundant evolutionarily conserved CSB-PiggyBac fusion protein expressed in cockayne syndrome PLoS Genetics 2008-03-01; 4(3). [PubMed]
  9. Cho, H.D. et al. Reengineering CCA-adding enzymes to function as (U,G)- or dCdCdA-adding enzymes or poly(C,A) and poly(U,G) polymerases Proceedings of the National Academy of Sciences of the United States of America 2007-01-02; 104(1):54-59. [PubMed]
  10. Newman, J.C. et al. Cockayne syndrome group B protein (CSB) plays a general role in chromatin maintenance and remodeling Proceedings of the National Academy of Sciences of the United States of America 2006-06-20; 103(25):9613-9618. [PubMed]
  11. Cho, H.D. et al. A model for C74 addition by CCA-adding enzymes: C74 addition, like C75 and A76 addition, does not involve tRNA translocation Journal of Biological Chemistry 2006-04-07; 281(14):9801-9811. [PubMed]
  12. Newman, J.C. et al. L2L: a simple tool for discovering the hidden significance in microarray expression data. Genome biology 2005-12-01; 6(9):R81. [PubMed]
  13. Weiner, A.M. et al. E pluribus unum: 3′ end formation of polyadenylated mRNAs, histone mRNAs, and U snRNAs Molecular Cell 2005-10-28; 20(2):168-170. [PubMed]
  14. Cho, H.D. et al. Archaeal CCA-adding enzymes: Central role of a highly conserved ß-turn motif in RNA polymerization without translocation Journal of Biological Chemistry 2005-03-11; 280(10):9555-9566. [PubMed]
  15. Weiner, A.M. et al. tRNA maturation: RNA polymerization without a nucleic acid template Current Biology 2004-10-26; 14(20):R883-R885. [PubMed]
  16. Cho, H.D. et al. A single catalytically active subunit in the multimeric Sulfolobus shibatae CCA-adding enzyme can carry out all three steps of CCA addition Journal of Biological Chemistry 2004-09-17; 279(38):40130-40136. [PubMed]
  17. Jacobs, E.Y. et al. Role of the C-Terminal Domain of RNA Polymerase II in U2 snRNA Transcription and 3′ Processing Molecular and Cellular Biology 2004-01-01; 24(2):846-855. [PubMed]
  18. Xiong, Y. et al. Crystal structures of an archaeal class I CCA-adding enzyme and its nucleotide complexes Molecular Cell 2003-11-01; 12(5):1165-1172. [PubMed]
  19. Weiner, A. et al. Soaking up RNAi Molecular Cell 2003-09-01; 12(3):535-536. [PubMed]
  20. Cho, H.D. et al. Use of nucleotide analogs by class I and class II CCA-adding enzymes (tRNA nucleotidyltransferase): Deciphering the basis for nucleotide selection RNA 2003-08-01; 9(8):970-981. [PubMed]
  21. Li, F. et al. Crystal structures of the Bacillus stearothermophilus CCA-adding enzyme and its complexes with ATP or CTP Cell 2002-12-13; 111(6):815-824. [PubMed]
  22. Tomita, K. et al. Closely related CC- and A-adding enzymes collaborate to construct and repair the 3′-terminal CCA of tRNA in Synechocystis sp. and Deinococcus radiodurans Journal of Biological Chemistry 2002-12-13; 277(50):48192-48198. [PubMed]
  23. Newman, J.C. et al. Measuring the immeasurable Molecular Cell 2002-09-01; 10(3):437-439. [PubMed]
  24. Weiner, A.M. et al. SINEs and LINEs: The art of biting the hand that feeds you Current Opinion in Cell Biology 2002-06-01; 14(3):343-350. [PubMed]
  25. Cho, H.D. et al. U2 small nuclear RNA is a substrate for the CCA-adding enzyme (tRNA nucleotidyltransferase) Journal of Biological Chemistry 2002-02-01; 277(5):3447-3455. [PubMed]