Bloom deficiency in humans leads to Bloom Syndrome autosomal recessive growth retardation “butterfly” rash due to sun exposure compromised immune system elevated risk of cancer This syndrome is AUTOSOMAL RECESSIVE, meaning that humans need TWO null alleles in order to show symptoms of the syndrome. In general, there is a wide range of mutations that have been found from sequencing in human bloom syndrome patients. Many are deletions or frameshifts that lead to the Nuclear Localization Sequence not being expressed in the protein, which would completely ablate protein function because it wouldn’t be able to get to the DNA http://www.hxbenefit.com/bloom-syndrome.html

Bloom deficiency leads to cancer susceptibility in humans Cause of death is almost always cancer Average age of death is 27 https://www.jewishgenetics.org/bloom’s-syndrome Maureen M Sanz and James German. 2013. Gene Reviews. http://www.ncbi.nlm.nih.gov/books/NBK1398/

Bloom Syndrome Protein is a helicase unwinds DNA 3’ to 5’ ATPase preferentially binds to single stranded DNA catalyzes ssDNA annealing The image is of a Bloom dimer attached to a double Holliday Junction. A cartoon of how Bloom can dissolve these structures is on slide seven K. Kitano. 2014. Front Genet. 29;5:366

Bloom is a member of the RecQ family of helicases These five helicases shown are all HUMAN proteins in the HUMAN RecQ family of helicases. RecQ C terminus domain: mediates binding to G Quadruplex DNA and stabilizes binding to other DNA structures HDRC domain: plays a role to laser induced double stranded DNA breaks NLS: nuclear localization sequence Raymond J. Monnat Jr. 2010. Seminars in Cancer Biology. 20:329-339

Bloom Helicase interacts with myriad proteins to maintain genomic integrity Base Excision Repair Double Strand Break Repair Deborah L. Croteau, Venkateswarlu Popuri, Patricia L. Opresko, and Vilhelm A. Bohr. 2014. Annu. Rev. Biochem. 83:519-52

Bloom Helicase plays a key role in repairing DNA double strand breaks via homologous repair. double stranded breaks can be repaired through non homologous end joining, alternative non homologous end joining, or homologous repair, which is the pathway in which BLM functions. SDSA: break strands undergo resection, which then invade the homologous chromosome forming a D loop. Synthesis extends the invading strand, which then dissociates and anneals to the other half of the DNA double strand break, allowing completion of repair or dHJ dissolution: 5’ end undergoes resection, one tail then commits rad51 mediated strand invasion, 2nd end capture occurs when D-loop is displaced by synthesis, and then ligation of the free end results in formation of a double HJ - this can be resolved with possible gene conversion by nicking the sides of the HJ (or can result in crossing over when BLM isn’t present) Deborah L. Croteau, Venkateswarlu Popuri, Patricia L. Opresko, and Vilhelm A. Bohr. 2014. Annu. Rev. Biochem. 83:519-52

BLM functions in homologous repair both through the SDSA and dHJ dissolution pathways Bloom acts as a disruptase during SDSA, in which it pushes the free end of the DNA off of the homologous chromosome so that it can re anneal to the proper chromosome Bloom acts as a dissolvase during dHJ dissolution, in which it forces the crosses in the double holliday junction closer to each other, allowing the strands to be cut without a crossover event. Synthesis Dependent Strand Annealing Double Holliday Junction Dissolution M. McVey, S. L. Anderson, Y. Broze, and J. Sekelsky. 2007. Genetics. 176:1979-1992

Bloom deficiency leads to an increased rate of sister chromatid exchanges m1 and m3 are null alleles of bloom, so the cell on the right is Bloom deficient - these images were generated from BLM deficient mice Homologous chromosomes were differentially labeled with Brdu, which is taken up into the DNA itself during synthesis and acts as a fluorescent marker In wild type cells, no mitotic crossing over occurs, but in bloom deficient mice, mitotic crossing over occurs at a very high rate, indicating that double strand breaks could not be repaired through the SDSA or dHJ dissolution models in which bloom is required, but instead end up in a mitotic recombination event Luo G, Santoro IM, McDaniel LD, Nishijima I, Mills M, Youssoufian H, Vogel H, Schultz RA, Bradley A. 2000. Nat. Genet. 26:424–29

Bloom deficiency facilitates chromosomal instability A different lab also generated Bloom null mice - these are cells that have been DAPI stained, which marks the nucleus This is showing all the types of chromosomal aberrations that were found in cells lacking bloom MN = micronuclei AL = anaphase laggards m-nuc = multinucleated m-lobed = multilobed mTL = metacentric translocations SMC = small marker chromosome - indicative of a break, smaller than the smallest chromosome Chester N, Babbe H, Pinkas J, Manning C, Leder P. 2006. Mol Cell Biol. 26(17):6713-26