?url_ver=Z39.88-2004&rft_val_fmt=info%3Aofi%2Ffmt%3Akev%3Amtx%3Adc&rft.relation=http%3A%2F%2Fd-scholarship-dev.library.pitt.edu%2F25446%2F&rft.title=HsMCM8+and+HsMCM9%3A+Essential+for+Double-Strand+Break+Repair+and+Normal+Ovarian+Function&rft.creator=Jeffries%2C+Elizabeth+Paladin&rft.description=The+minichromosome+maintenance+(MCM)+family+of+proteins+is+conserved+from+archaea+to+humans%2C+and+its+members+have+roles+in+initiating+DNA+replication.+MCM8+and+MCM9+are+minimally+characterized+members+of+the+eukaryotic+MCM+family+that+associate+with+one+another+and+both+contain+conserved+ATP+binding+and+hydrolysis+motifs.+The+MCM8-9+complex+participates+in+repair+of+DNA+double-strand+breaks+by+homologous+recombination%2C+and+MCM8+is+implicated+in+meiotic+recombination.+We+identified+a+novel+alternatively+spliced+isoform+of+HsMCM9+that+results+in+a+medium+length+protein+product+(MCM9M)+that+eliminates+a+C-terminal+extension+of+the+fully+spliced+product+(MCM9L).+Quantitative+real-time+reverse+transcriptase+PCR+(qRT-PCR)+of+the+relative+mRNA+isoform+abundances+across+cell+lines+reveals+MCM9L+transcript+is+more+abundant+than+MCM9M.+The+expression+of+both+isoforms+is+cell+cycle+regulated%2C+as+they+are+most+abundant+in+S-phase.+Wild-type+MCM9L+forms+DNA+damage-dependent+nuclear+foci%2C+while+MCM9M+is+cytoplasmic+and+MCM9Cterm+is+diffuse+throughout+the+nucleus.+We+have+identified+and+verified+a+putative+nuclear+localization+signal+(NLS)%2C+and+Rad51-interacting+motif+(BRCv)+in+the+C-terminus+of+MCM9.+GFP-tagged+MCM9+NLS-+is+solely+cytoplasmic%2C+GFP-tagged+MCM9+BRCv-+is+diffuse+throughout+the+nucleus.+A+combination+of+SNP+arrays%2C+comparative+genomic+hybridization+arrays%2C+and+whole-exome+sequencing+analyses+identified+homozygous+pathogenic+variants+in+MCM8+(MCM8+c.446C%3EG%3B+p.P149R)+and+MCM9+(MCM9+c.1732.2T%3EC+and++MCM9+c.394C%3ET%3B+p.R132*)%2C+all+located+within+regions+of+homozygosity+in+women+afflicted+by+premature+ovarian+failure+(POF).+The+MCM9+c.1732.2T%3EC+variant+alters+a+splice+donor+site%2C+resulting+in+abnormal+alternative+splicing+and+truncated+forms+of+MCM9+that+are+unable+to+be+recruited+to+sites+of+DNA+damage.+In+the+second+family%2C+MCM9+c.394C%3ET+(p.R132*)+results+in+a+predicted+loss+of+functional+MCM9.+Compared+with+fibroblasts+from+unaffected+family+members%2C+chromosomal+break+repair+was+deficient+in+fibroblasts+from+all+affected+individuals%2C+likely+due+to+inhibited+recruitment+of+mutated+MCM8+or+MCM9+to+sites+of+DNA+damage.+Our+cumulative+results+suggest+that+MCM8-9+have+evolved+to+act+late+in+both+mitotic+recombination+pathways+to+aid+in+crossover+migration+and%2For+strand+resolution.&rft.date=2015-09-24&rft.type=University+of+Pittsburgh+ETD&rft.type=PeerReviewed&rft.format=application%2Fpdf&rft.language=en&rft.identifier=http%3A%2F%2Fd-scholarship-dev.library.pitt.edu%2F25446%2F1%2FETD_EPJeffries_v43.pdf&rft.identifier=++Jeffries%2C+Elizabeth+Paladin++(2015)+HsMCM8+and+HsMCM9%3A+Essential+for+Double-Strand+Break+Repair+and+Normal+Ovarian+Function.++Doctoral+Dissertation%2C+University+of+Pittsburgh.++++(Unpublished)++