Question

Understanding regulation of transcription in eukaryotes. The images below depict how the expression of a gene required to break down lactose in yeast is regulated. The top image illustrates the key regulators of the process. The bottom two panels illustrate what happens in the presence and absence of galactose. Describe how the expression of this gene is regulated. Include the roles of all key players illustrated below in this process. Key players in the process GAL4 GAL GAL80 UAS Upstream activating sequence for GAL4 TATA Gene promoter Coding sequence RNA polymerase Transcription factors Galactose present: No galactose: GAL3 — GAL4 UAS GALA VASG GAL80 GAL80 GAL3 TATA Galactose Basal transcription apparatus

Describe how the expression of this gene is regulated. Include the roles of all key players (GAL3, GAL4, GAL80, Gene promoter, RNA polymerase, and transcription factors) illustrated.

Answer 1

In yeast, the Gal4 system regulates the transcription of all Gal genes (GAL1, GAL10, GAL2 and GAL7) involved in galactose metabolism. Gal 4 (purple) is a transcriptional activator of all GAL genes. It can bind to the upstream activator sequence in GAL gene promoter (UASG) on genes for galactose pathway. It binds via its N-terminal domain as a dimer. As a result, there is activation of transcription via effects on the C-terminal transactivation (transcription activation) domain (AD).

Gal 80 (peach) protein is an inhibitor that can bind to Gal4 dimer and inhibit its binding to UASG sequence present in Gal genes. Gal3 (blue) protein can bind to Gal80 and remove its inhibitory effect on Gal4 dimer. Promoter contains the TATA box to which RNA polymerase and other transcription factors bind to initiate transcription of coding sequence of GAL genes.

Absence of Galactose (lower Left panel): When the medium lacks galactose, Gal4 dimer is bound by Gal80 dimer. Gal 80 binds to a small peptide within the transactivation domain of Gal4. This will inhibit the transactivation domain. As a result, Gal4 dimer can bind to UASG, but cannot activate the transactivation domain. This will inhibit the transactivation domain from interacting with the basal transcription apparatus (RNA polymerase bound to TATA box along with transcription factors). Basal transcription apparatus involves binding of RNA polymerase to TATA box in the promoter, followed by binding of transcription factors such as TFIID, TBP etc. RNA polymerase cannot initiate transcription of GAL genes. Thus, GAL genes are not expressed when galactose is absent.

Presence of Galactose (lower right panel): Galactose binds to Gal3. Gal3 is converted to an active form which then binds to Gal80 dimer. As a result, Gal4 dimer binds to UASG and is not bound to Gal80 dimer at the transactivation domain. Thus, the Gal4-UASG binding activates the transactivation domain which then interacts with the basal transcription apparatus. RNA polymerase now initiates transcription and GAL genes are expressed. Thus, GAL genes are only transcribed when galactose is present in the medium.