1. Bloom’s Syndrome and Bloom helicase Alexandra Otto March 16, 2004

2. Bloom Syndrome Syndrome was first described by New York dermatologist David Bloom in 1954 Syndrome was first described by New York dermatologist David Bloom in 1954 Extremely rare Extremely rare ~ 220 cases worldwide Death before age 30 Death before age 30 Mean age of cancer diagnosis ~ 24 Mean age of cancer diagnosis ~ 24 BS is associated with a predisposition to cancers of all types BS is associated with a predisposition to cancers of all types Autosomal recessive disorder Autosomal recessive disorder Arises from a mutation in the gene BLM Arises from a mutation in the gene BLM

3. Clinical Features of BS Proportional dwarfism Proportional dwarfism Sun-induced erythema Sun-induced erythema Type-II diabetes Type-II diabetes Narrow face and prominent ears Narrow face and prominent ears Male infertility and female sub-fertility Male infertility and female sub-fertility Frequent infections Frequent infections http://www.skinsite.com/erythema

4. How was BLM identified? Prevalence of BS among the Ashkenazi Jewish population (carrier rate of 1%) Prevalence of BS among the Ashkenazi Jewish population (carrier rate of 1%) Used positional cloning (like Rb) Used positional cloning (like Rb) Maps to chromosome 15q26.1 Maps to chromosome 15q26.1 ) http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=OMIM

5. BLM encodes a helicase Helicases are enzymes that separate the complementary strands of nucleic-acid duplexes Helicases are enzymes that separate the complementary strands of nucleic-acid duplexes essential for all aspects of DNA metabolism essential for all aspects of DNA metabolism http://www.blc.arizona.edu/marty/411/Modules/Lectures/Figures/helicase.GIF

6. The RecQ helicase family BLM helicase is a member of the RecQ family BLM helicase is a member of the RecQ family RecQ family gets its name from the recQ gene in E. coli. RecQ family gets its name from the recQ gene in E. coli. Family members share a homologous region with E. coli Family members share a homologous region with E. coli Conserved region is flanked by stretches of amino acids called the N-terminal region and the C-terminal region Conserved region is flanked by stretches of amino acids called the N-terminal region and the C-terminal region

7. http://www.nature.com/nrc/journal/v3/n3/images/nrc1012-f1.jpg

8. RecQ helicases Unicellular organisms express 1 RecQ enzyme whereas humans express 5 Unicellular organisms express 1 RecQ enzyme whereas humans express 5 Defects in 3 of these human RecQ helicases (BLM, WRN, and RECQ4) give rise to clinical disorders associated with cancer predisposition Defects in 3 of these human RecQ helicases (BLM, WRN, and RECQ4) give rise to clinical disorders associated with cancer predisposition Bloom’s syndrome, Werner’s syndrome, and Rothmund-Thomson syndrome Bloom’s syndrome, Werner’s syndrome, and Rothmund-Thomson syndrome

9. Role of Bloom helicase Required for the maintenance of genomic integrity Required for the maintenance of genomic integrity Duplex unwinding Duplex unwinding ‘Caretaker’ tumor-suppressor ‘Caretaker’ tumor-suppressor Caretakers influence genomic stability without directly regulating tumorigenesis Caretakers influence genomic stability without directly regulating tumorigenesis Repair of double-strand breaks Repair of double-strand breaks

10. http://nar.oupjournals.org/cgi/content/full/31/21/6272

11. Role of BLM helicase  bloom helicase normally plays a role in the repair of DSB by the homologous recombination pathway  In Bloom’s syndrome cells, repair may occur through the error-prone NHEJ pathway  increased genomic instability and predisposition to malignancy.  BLM helicase has not been placed at an exact step in the HR pathway

12. Possible roles of BLM Ability to process recombination intermediates during DNA replication Ability to process recombination intermediates during DNA replication - G-quadruplexes, hairpins  Bloom helicase could reset the replication fork by branch migration http://www.nature.com/nrc/journal/v3/n3/images/nrc1012-f1.jpg

13. BLM helicase as a roadblock remover. http://www.biochemj.org/bj/374/0577/bj3740577.htm

14. Branch migration http://www.biochemj.org/bj/374/0577/bj3740577.htm

15. Interaction with crucial proteins  BLM has not been definitively placed at a certain step in the homologous recombination pathway, but is known to interact with a number of crucial proteins recombination pathway, but is known to interact with a number of crucial proteins http://www.nature.com/nrc/journal/v3/n3/images/nrc1012-f1.jpg

16. Features of BLM helicase mutants Abnormal DNA replication Abnormal DNA replication Elevated level of homologous recombination Elevated level of homologous recombination In Bloom’s syndrome cells → accumulation of abnormal replication intermediates → accumulation of abnormal replication intermediates → increase in the frequency of reciprocal exchanges → increase in the frequency of reciprocal exchanges → ~ 10 fold increase in sister-chromatid exchanges → ~ 10 fold increase in sister-chromatid exchanges

17. Mouse Model Knockout mice - death by extreme anemia at 13.5 days - death by extreme anemia at 13.5 days - immortalized cell line showed a high frequency of sister chromatid exchange exchange - characteristic short stature is seen in early stages of embryo development development Viable BLM-/- Mouse - elevated rate of mitotic recombination - high frequency of sister-chromatid exchanges and somatic loss of heterozygosity - high cancer incidence (lymphomas, carcinomas, sarcomas) http://www.weizmann.ac.il/home/ligivol/publications/PNAS%201999.pdf

18. Cancer Predisposition What features of hyper-recombination underlie the cancer predisposition?  Recombination events are not carried out with perfect fidelity  Events are not carried out to completion This leads to:  Chromosomal duplication or breakage  Genomic instability and therefore cancer

19. BLM helicase and cancer Concluding points  BLM helicase is a caretaker tumor suppressor  Proposed to act in HR pathway  Homologous recombination exists to repair double strand breaks and damaged replication forks  Sister chromatid exchanges arise during HR from the crossing over of chromatid arms  BS cells have high frequency of SCE  This hyper-recombination results from defective replication  Without BLM helicase, replication cannot proceed smoothly  Genomic instability → CANCER predisposition  Cancer of all types because all cells need to repair damages in replication machinary

20. Works Cited http://www.ncbi.nlm.nih.gov http://www.nature.com/nrc/journal/v3/n3/images/nrc1012-f1.jpg http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abs tract&list_uids=12691817 http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abs tract&list_uids=12691817 http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abs tract&list_uids=12427531 http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abs tract&list_uids=12427531 http://www.biochemj.org/bj/374/0577/bj3740577.htm http://www.mssm.edu/jewish_genetics/genetic_diseases.shtml http://www.nature.com/cgi- taf/DynaPage.taf?file=/onc/journal/v21/n58/full/1205959a.htm http://tmm.trends.com http://www.biomedcentral.com/1471-2121/4/15