1Office hoursWednesday 3-4pm304A Stanley HallReview session5pm Thursday, Dec. 11GPB100Coding sequence arrayFig. 1.13“RNA-seq”AAAAAAAAAAAAAAA counts seq maps genome pos orient 26982 TCGTTCACTTGTATTGG U0 sbaySum.fsa 3002121 R 25885 TCTCTACTGGTGGTGGT U0 chr03.fsa 138912 F 17851 GACAAACTGTCGCTGTC R0 15715 TCTCTACCGGTGGTGGT U0 sbaySum.fsa 11182634 F 14963 TCGTTCACTTGTAATGG U0 chr04.fsa 1489376 R … 610 TTGAAGCTACCACCAAC R0 607 TGTCCTTATCTATGTGT U0 sbaySum.fsa 8491512 R 602 CGTACTTTTTTAATTGT U1 chr11.fsa 145622 F 602 ATCATTGTTCCAGAAAT R0 601 TTATCAGAGGTGCTGGT U0 chr14.fsa 694435 R 600 CAAAGACCTCATTTAAT U0 sbaySum.fsa 4671591 R 599 AGGAGGGTATATATGCT U0 chr14.fsa 575509 R …Solexa sequencingExpression effects of cancerDiagnosis via transcriptional profiletranscriptspatient samplesDiagnosis via transcriptional profile2Linkage mapping of mRNA levels“Black 6” mouse x “DBA” mouse111 F2 progenyMicroarrayeach F2 liver…Genotypeeach F2Looking for linkage (coinheritance) betweenmarker and mRNA level.Marker is linked to polymorphism inexpression regulation cascadeORFTFTFGkinaseTFGGLocally acting polymorphismsPolymorphism responsible for mRNA difference is at the locusof the gene itself~25% of varying mRNAsare caused by locallyacting polymorphismNonlocal polymorphismsNonlocal polymorphismsOne polymorphism in a key regulator can affect aregulon: 100’s of related mRNAs.Clinical applications“Black 6” mouse x “DBA” mouse111 F2 progenyMicroarrayeach F2 liver…Genotypeeach F2Measure fatpad each F23Clinical applicationsClinical applicationsColored curves= fat mass atdifferent bodylocationsClinical applicationsFinding polymorphism responsible fordifference in macroscopic phenotype is hardClinical applicationsFinding polymorphism responsible fordifference in macroscopic phenotype is hardIf mRNAs change too, can learn mechanismfrom known function of encoded proteinsClinical applicationsClinical applications4Clinical applicationsClinical applicationsCounts allele 1/allele 2, casesCounts allele 1/allele 2, controls= 1.5Clinical applicationsClinical applicationsMarker predicts quantitative expression level = associationCan we map expression traitsfirst, disease afterward?Linkage of human transcripts5Linkage of human transcriptsAssociation of human transcriptsAssociation of human transcriptslinkage(families)assoc(unrelated)Association in multiple populationsHan Chinese and JapaneseEuropean-Americans in UtahAssociation in multiple populationsA new brand of geneticvariation6Fig. 8.25TranslationFig. 8.25TranslationPSI+ yeast read through STOPsPSI+ yeast read through STOPsPSI+ yeast read through STOPs40-150 bpWeird genetics of read-throughHaploid PSI+Haploid psi-Diploid PSI+7Weird genetics of read-throughHaploid PSI+Haploid psi-Diploid PSI+sporulateHaploid PSI+ Haploid PSI+Haploid PSI+ Haploid PSI+…Weird genetics of read-throughHaploid PSI+Haploid psi-Diploid PSI+sporulateHaploid PSI+ Haploid PSI+Haploid PSI+ Haploid PSI+But whywouldn’t it getdiluted overmanydivisions?Cytoplasmic aggregates of Sup35How would this explain the results?Cytoplasmic aggregates of Sup35Hard to make aggregate, easy to joinProtein inheritance8Protein inheritanceOriginal fromPSI+nucleates in allprogenyPSI+ begets more PSI+Not due to dominant genetics in the usual way.Nucleating prions in mad cow diseaseIs PSI+ the yeast analog ofmad cow or Alzheimer’s?If bad, would be lostIf bad, would be lost(Pichiamethanolicavs. S.cerevisiae)9PSI+ phenotypesPSI+ phenotypesGenetically distinct S. cerevisiae strainsPSI+ phenotypesGenetically distinct S. cerevisiae strainsPSI+ phenotypes50°CGenetically distinct S. cerevisiae strainsPSI+ phenotypes50°CGenetically distinct S. cerevisiae strainsPSI+ phenotypesWhy would aggregates result in so manynovel abilities and traits??10PSI+ phenotypesWhy would aggregates result in so manynovel abilities and traits??Apparently does not happen inAlzheimer’s…The clincherThe clincher(partial LOF)The clincher(partial LOF)What do you conclude?A. Sup35 does not cause resistanceto paraquat.B. Prions do not cause resistance toparaquat.C. Aggregation is not necessary tocause resistance to paraquat.D. Aggregation is not sufficient tocause resistance to paraquat.The clincher(partial LOF)Aggregates per se don’tcause resistance.The clincher(partial LOF)Aggregates per se don’tcause resistance.Losing function of sup35does.11Aggregation = more read-throughAggregation = more read-throughNew extra-longforms of proteinscause new traits.PSI+ allows epigeneticchange in protein sequence.Genetic effects?Phenotypes variedbetween geneticallydiverse PSI+ strains.Genetically distinct S. cerevisiae strainsGenetic effects?Phenotypes variedbetween geneticallydiverse PSI+ strains.How can the cause begenetic and non-genetic(protein aggregates) at thesame time?Genetically distinct S. cerevisiae strainsAggregation = more read-through12Aggregation = more read-throughUTR mutationsusually occur andhave no effectAggregation = more read-throughUTR mutationsusually occur andhave no effect, soorganism doesn’tdie and allele ismaintained.Aggregation = more read-throughUTR mutationsusually occur andhave no effect, soorganism doesn’tdie and allele ismaintained.Now in PSI+, theymanifest!Aggregation = more read-throughSequencedifferences in read-through regioncause phenotypicdifferencesbetween PSI+strains.Cryptic variation: DNAsequence differences betweenindividuals that are usually
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