The Genom of Homo sapiens.pdf

HAPLOTYPE EXPRESSION ASSOCIATIONS 105track 69 (Ttk69) protein are important for Ttk69 activity;single amino acid changes in the domain abolish transcriptionalrepression activity (Wen et al. 2000). Furthermore,the zinc fingers in the carboxyl terminus of theTtk69 protein were also important for DNA binding andfunction. ZNF295 is expressed in a wide variety of tissues(Unigene cluster Hs.157079) and is also conserved inmouse (ZNP295, 83.7% identity, Homologene) and in rat(LOC304056, 85.1% identity, Homologene).One interesting observation (Fig. 2) is that the directionof change of the expression levels is not the same for allgenes. In some cases, the Marker(+) group has higher expressionlevels than the Marker(–) group, whereas in others,the opposite is seen. One can speculate that the codingchanges seen in this marker are directly responsiblefor altering the activity of a transcription complex that includesZNF295. If this is the case, one would assume thatthe sign of this activity change would be consistent acrosstranscripts regulated by the complex. However, it couldalso be true that some of these associations are secondordereffects; i.e., ZNF295 is responsible for the transcriptionof some number of the associated genes, but theremaining associated genes are themselves induced bythe directly transcribed genes. We prefer the second explanationbecause, as discussed below, many associationsof ZNF295 haplotypes are with transcription factors,coactivators and genes involved in transcriptional regulation.This induction could be positively or negatively affectedby changes in expression levels of the originalgenes.There are several interesting observations to be made inthe set of genes whose expression levels are associatedwith markers in ZNF295. There is a statistically significantexcess of observed genes on Chromosomes 17 and22. Of the transcripts on Chromosome 22, several clusteron sub-band 22q13.1. As shown in Table 6, ZNF295 haplotypemarkers are associated with 36 genes that are involvedin transcriptional regulation; have DNA-bindingproperties; or are involved in DNA repair and metabolism.Expression levels of 14 genes that are involved in RNAprocessing or that have RNA-binding properties are alsoassociated to markers within the ZNF295 gene. Furthermore,expression levels of several members of very importantclasses of genes such as GTPases (10 genes), immuneresponse genes (10 genes), ATPase or ATP-binding(9 genes), genes involved in cell cycle or cell division orcell–cell signaling (10 genes), protein kinases (13 genes),protein trafficking (11 genes), genes involved in proteinbinding and folding (13 genes), genes involved in signaltransduction (11 genes), and genes involved in apoptosis(5 genes) are highly significantly and reproducibly associatedwith markers within the ZNF295 gene. One examplethat illustrates the impact of ZNF295 haplotypes on expressionlevels of potential target genes is the associationthat links ZNF295 with expression levels of the transcriptionfactor SP100. The 7 individuals in the Marker(+)group have a mean expression level for SP100 of 90.8,whereas the 82 individuals in the Marker(–) group have amean expression level of 52.8. The raw and permutationadjustedp values for the association are 0.00024 and0.0034, respectively. In the replication experiment, theMarker(+) and Marker(–) groups had 6 and 61 individuals,respectively. The respective means were 65.6 and21.2, and the adjusted p value was 0.00016. Three otherinstances are noteworthy because of the involvement ofgenes that have the POZ domain; the association betweenmarkers in the ZNF295 gene and expression levels in theENIGMA, BTBD2, and TAZ genes (Guy et al. 1999;Kanai et al. 2000; Carim-Todd et al. 2001). The BTBD2and the TAZ genes are involved in transcriptional regulation,whereas the ENIGMA gene is involved in receptormediatedendocytosis.Another noteworthy feature of our data is that the twoamino acid changes (Asn181Ser and Lys218Gln) fall outsideof the POZ and the zinc finger domains. Classic mutationalapproaches (such as the changes in the POZ domainof the Tramtrack protein mentioned above) usuallyabolish transcriptional activity such that subtle relationshipsbetween members of the transcriptional machineryare not revealed. By exploiting the naturally occurringvariants in ZNF295, we have not only obtained clues tothe biological function of ZNF295 itself, but also to howit is networked into the larger transcriptional machineryand the extent of its influence on the different cellularprocesses. These results demonstrate the utility of combiningexpression analyses with genetic analyses (such asthe use of haplotype markers) to unravel gene networksand discover gene functions.Figure 4. Fraction of associations withZNF295 that are confirmed (p ≤0.05 in thereplication experiment) as a function of p value(adjusted) in the original experiment. Thedashed line represents the expectation underthe null hypothesis of no confirmation.Fraction Confirmed70%60%50%40%30%20%10%0%10 15 20 25 30 35 40 45 50-10 log(p)