Current research profile
The research in the BA laboratory has for many years focused on the molecular genetics and the molecular pathology of human disease with a special emphasis on regulation of normal and aberrant alternative pre-mRNA splicing. Initially, the research was mainly focused on studies of how mutations can disrupt the splicing regulatory mechanisms and cause disease. We have characterized numerous splicing regulatory elements and their binding proteins by using splicing reporter mini genes, expression systems for splicing regulatory factors and in vitro techniques.
In recent years, our laboratory has focused intensely on development of techniques for Next Generation Sequencing (NGS) of RNA, development of data-analysis pipelines, as well as global mapping and characterization of in vivo binding sites for splicing regulatory factors using iCLIP and HITS-CLIP techniques.
We have now taken our projects to a new level, where we use global analysis of mRNA splicing in patient cells and cell lines, and global identification of binding motifs for splicing regulatory proteins in manipulated cells. This allows us to answer questions of a more general nature. One of our overall goals is the deciphering of the “splicing code”.
We employ RNA-affinity purification techniques and Surface Plasmon Resonance (SPR) based binding assays to identify and characterize binding of splicing regulatory proteins. Recently we have developed methods for modulation of splicing using splice shifting oligonucleotides (SSOs) to modulate splicing and to target disease-associated splicing regulatory elements aiming at development of new individualized therapies for inherited diseases as well as cancer.
Very recently we have initiated several projects studying splicing by the use of bioinformatics, including employing the supercomputer at SDU, Abacus 2.0, to use neural networks to analyze protein binding sites.
Our research is funded by grants from the National Danish Research Councils (FNU and FSS) and Foundations like Novo Nordisk Fonden and Lundbeck Fonden.
The BA group consists of: 1 PI, 4 postdoc/AC-TAP, 3 PhD students, 2 technician trainees, 4 master students and several students on (Bachelor/ITEK) projects.
Head of research: Professor Brage Storstein Andresen
Postdoc Maja Dembic
Some of our publications on aberrant splicing in human disease:
We have reported how mutations in the non-variant positions of splice sites can cause disease (Korman et al. 2004), how mutations in exons can disrupt splice sites (Andresen et al. 2000, Maydan et al. 2006) and how apparently benign changes in splice sites may instead cause aberrant splicing and human disease (Madsen et al. 2006; Roca et al. 2008). Moreover, we have shown how mutations located outside of the splice sites may affect splicing regulatory elements and cause missplicing (Nielsen et al. 2007, Dobrowolski et al. 2010) and that even deep intronic mutations may cause aberrant splicing and disease by pseudoexon activation (Homolova et al. 2010). We have shown that the affected splicing regulatory elements are general – As illustrated by a splicing regulatory element, which is mutated both in MCAD deficiency and spinal muscular atrophy (SMA). We have shown that a neutral single nucleotide polymorphism (SNP) in this general element in the MCAD gene (ACADM) can protect against missplicing (Nielsen et al. 2007), and we have just finished a study showing that manipulation of this element or the splicing regulatory factors that bind to it can correct aberrant splicing of the SMN2 gene (Doktor et al. 2011). A review of splicing and disease can be found in Andresen and Krainer 2009.
1. Andresen BS, Christensen E, Corydon TJ, Bross P, Pilgaard B, Simonsen H, Knudsen I, Schroeder LD, Gregersen N, Skovby F (2000) Isolated 2-methylbutyrylglycinuria caused by short/branched-chain acyl-CoA dehydrogenase (SBCAD) deficiency: Identification of a new enzyme defect, resolution of its molecular basis and evidence for distinct acyl-CoA dehydrogenases in isoleucine and valine metabolism. Am J Hum Genet 67:1095-1103.
2. Korman SH, Gutman A, Brooks R, Sinnathamby T, Gregersen N, Andresen BS (2004) Homozygosity for a severe novel medium-chain acyl-CoA dehydrogenase (MCAD) mutation IVS3-1G>C that leads to introduction of a premature termination codon by complete missplicing of MCAD mRNA, and is associated with phenotypic diversity ranging from sudden neonatal death to asymptomatic status. Molecular Genetics and Metabolism. 82:121-129.
3. Madsen PP, Kibæk M, Roca X, Sachidanandam R, Krainer AR, Christensen E, Steiner R, Gibson KM, Corydon TJ, Knudsen I, Wanders RJA, Ruiter JPN, Gregersen N, Andresen BS (2006) Short/branched-chain acyl-CoA dehydrogenase deficiency due to an IVS3+3A>G mutation that causes exon skipping. Human Genetics 118(6):680-690.
4. Maydan G, Andresen BS, Madsen PP, Zeigler M, Raas-Rothschild A, Zlotogorski A, Gutman A, Korman SH (2006) TAT gene analysis in three Palestinian kindreds with oculocutaneous tyrosinemia II: characterization of a silent exonic transversion that causes complete missplicing by exon 11 skipping J Inher Metab Dis. 29:620-626.
5. Nielsen KB, Sørensen S, Cartegni L, Corydon TJ, Doktor TK, Schroeder LD, Reinert LS, Elpeleg ON, Krainer AR, Gregersen N, Kjems J, Andresen BS (2007) Seemingly neutral polymorphic variants may confer immunity to splicing inactivating mutations Am J Hum Genet. 80(3):416-32.
6. Roca X, Olson AJ, Rao AR, Enerly E, Kristensen VN, Borresen-Dale AL, Andresen BS, Krainer AR, Sachidanandam R (2008) Disease-causing mutations at 5’ splice sites and comparative genomics help identify features determining splice-site efficiency. Genome Research –18(1):77-87.
7. Andresen BS and Krainer AR (2009) When the genetic code is not enough – How sequence variations can affect pre-mRNA splicing and cause (complex) disease. Chapter 15 (pp.165-182) in Genetics of Complex Human Diseases. Edited by Laura Almasy and Ammar Al-Chalabi. Cold Spring Harbor Laboratory Press. Cold Spring Harbor, New York, USA. Paperback: ISBN 978-087969883-6 / Hardcover: ISBN 978-087969882-9).
8. Homolova K, Zavadakova P, Doktor TK, Schroeder LD, Kozich V, Andresen BS (2010) The prevalent intronic c.903+469T>C mutation in the MTRR gene creates an SF2/ASF binding ESE, which leads to pseudoexon activation and causes the cblE type of homocystinuria. Human Mutation 31:437–444. See also editorial comment in same issue.
9. Dobrowolski SF, Andersen HS, Doktor TK, Andresen BS (2010) The PAH c.30C>G synonymous variation (p.G10G) creates a common exonic splicing silencer. Molecular Genetics and Metabolism 100:316–323.
10. Doktor TK, Schröder LD, Jensen AV, Palmfeldt J, Andersen HS, Gregersen N, Andresen BS (2011) SMN2 exon 7 splicing is inhibited by binding of hnRNPA1 to a common ESS motif that spans the 3’ splice site. Human Mutation. 32(2):220-30. See also editorial comment in same issue.
Some of our publications on molecular genetic diagnostic methods, newborn screening and potential therapy:
BSA is involved in mutation diagnosis of inherited metabolic disorders (in particular MCAD and VLCAD deficiency) in routine newborn screening programs in Denmark and other countries (Andresen et al. 2001, Khalid et al. 2008). We also work with development of mutation detection methods (Olsen et al. 2010) and possible treatments (Gobin-Limballe et al. 2007).
11. Strauss AW, Andresen BS, Bennett MJ (2009) Chapter 5: Fatty-acid oxidation defects. pp. 51-70. In: Pediatric Endocrinology and Metabolism. Edited by Sarafoglou K. McGraw-Hill. ISBN: 0071439153/9780071439152.
12. Andresen BS, Dobrowolski SF, O`Reilly L, Muenzer J, McCandless S, Frazier D, Udvari S, Bross P, Knudsen I, Banas R, Chace D, Engel P, Naylor EW, Gregersen N (2001) The spectrum of medium-chain acyl-coa dehydrogenase (MCAD) mutations in newborns identified by MS/MS based prospective newborn screening is different from that observed in patients with clinical symptoms – Identification and characterization of a new prevalent mutation that results in mild MCAD deficiency. Am J Hum Genet 68:1408-18.
13. Khalid JM, Oerton J, Cortina-Borja M, Andresen BS, Besley G, Dalton N, Downing M, Foo Y, Green A, Henderson M, Leonard J, Dezateux C - On behalf of the UK Collaborative Study of Newborn Screening for MCADD (2008) Ethnic-specific birth prevalence of c.985A>G homozygous Medium-Chain Acyl-CoA Dehydrogenase Deficiency (MCADD): results from screening ~ 1.1 million newborn infants. The Journal of Medical Screening. 15(3):112-117.
14. Olsen RKJ, Dobrowolski SF, Kjeldsen M, Hougaard DM, Simonsen H, Gregersen N, Andresen BS (2010) High resolution amplicon melting analysis, A simple and effective method for reliable mutation scanning and frequency studies in the ACADVL gene. J Inherited Metabolic Disease 33:247–260.
15. Gobin-Limballe S, Djouadi F, Aubey F, Olpin S, Andresen BS, Yamaguchi S, Mandel H, Fukao T, Ruiter JPN, Wanders RJA, McAndrew R, Kim JJ, Bastin J (2007) Genetic basis for correction of Very Long Chain Acyl-CoA Dehydrogenase deficiency by bezafibrate in patient fibroblasts: towards a genotype-based therapy. Am J Hum Genet 81(6):1133-43.