15. Genomes and Genomics

Functional Genomics

Open Question
In conducting the study described in Problem 24, you have noted that a set of S. cerevisiae genes are repressed when yeast are grown under high-salt conditions.How might you determine whether this set of genes is regulated by a common transcription factor?
Open Question
What is the difference between biochemical and biological function?
Open Question
Using the two-hybrid system to detect interactions between proteins, you obtained the following results: A clone encoding gene A gave positive results with clones B and C; clone B gave positive results with clones A, D, and E but not C; and clone E gave positive results only with clone B. Another clone F gave positive results with clone G but not with any of A–E. Can you explain these results?To follow up your two-hybrid results, you isolate null loss-of-function mutations in each of the genes A–G. Mutants of genes A, B, C, D, and E grow at only 80% of the rate of the wild type, whereas mutants of genes F and G are phenotypically indistinguishable from the wild type. You construct several double-mutant strains: The ab, ac, ad, and ae double mutants all grow at about 80% of the rate of the wild type, but af and ag double mutants exhibit lethality. Explain these results.How do the two-hybrid system and genetic interaction results complement one another? Can you reconcile your two-hybrid system and genetic interaction results in a single model?
Open Question
As shown in Figure 14.1, mutations in the Drosophila Ultrabithorax (Ubx) gene result in wings developing from two thoracic segments, rather than just one as in wild-type flies. In the mouse genome there are two Ubx orthologs (see Genetic Analysis 14.2 for definition). How would you determine whether the two mouse genes have distinct or redundant functions?
Open Question
Although a single activator may bind many enhancers in the genome to control several target genes, in many cases, the enhancers have some sequence conservation but are not all identical. Keeping this in mind, consider the following hypothetical example:

- Undifferentiated cells adopt different fates depending on the concentration of activator protein, Act1.
- A high concentration of Act1 leads to cell fate 1, an intermediate level leads to cell fate 2, and low levels to cell fate 3.
- Research shows that Act1 regulates the expression of three different target genes (A, B, and C) with each having an enhancer recognized by Act1 but a slightly different sequence that alters the affinity of Act1 for the enhancer. Act1 has a high affinity for binding the enhancer for gene A, a low affinity for the gene B enhancer, and an intermediate affinity for the gene C enhancer.

From these data, speculate on how Act1 concentrations can specify different cell fates through these three target genes? Furthermore, which target genes specify which fates?
Open Question
Hereditary spherocytosis (HS) is a disorder characterized by sphere-shaped red blood cells, anemia, and other abnormal traits. Ankyrin-1 (ANK1) is a protein that links membrane proteins to the cytoskeleton. Loss of this activity is associated biochemically to HS. However, Gallagher et al. (2010) (J. Clin. Invest. 120:4453–4465) show that HS can also be caused by mutations within a region from -282 to -101 relative to the transcriptional start site, which lead to constitutive transcriptional repression in erythroid cells due to local chromatin condensation. Propose a hypothesis for the function of the -282 to -101 region of the ANK1 gene.
Open Question
Through a forward genetics screen in Arabidopsis you have identified a mutation that results in leaves curling upward, rather than being flat as in wild type. You have cloned the corresponding gene and note that it is a member of a small gene family composed of three additional members in Arabidopsis. How will you determine if the other three members of the gene family have similar or distinct functions as compared with the gene you first identified?
Open Question
Transcription factors play key roles in the regulation of gene expression, but to do so, they must act within the nucleus. Like most proteins, however, transcription factors are translated in the cytoplasm. To enter the nucleus, transcription factors contain nuclear localization signals, which in some cases can work only when bound to some other molecule such as a steroid hormone. After entering the nucleus, transcription factors must bind to appropriate DNA sites and must interact with other transcription proteins at promoters, enhancers, and silencers. Transcription factors then activate or repress transcription through their activation or repression domains. Many drug therapies target transcription factors. Based on the information provided above, suggest three specific mechanisms through which a successful drug therapy, targeted to a transcription factor, might work.