Molecular Genetics

Auxiliary transcription factors consist of multiple domains. One is needed for DNA sequence-specific binding, another for transcription activation. In many transcription facctors, a third domain is required for signal transduction.

Facts

• Gal4 is a transcription regulating protein from S. cerevisiae. The protein binds specific DNA sequences in vitro.
• Substitution and deletion mutations were made in various places (indicated by x at right) along the gene for the gal4 protein. The mutated proteins were produced in vitro. DNA binding activity and ability to activate transcription were measured.
• Gal regulation requires an additional protein, gal 80. When gal 80 is deleted, gal4-dependent transcription is constitutive. Gal 80 binds galactose and part of gal4.
• Genes encoding proteins chimeric between gal4 and a metal binding transcription regulator (MBP) were constructed.
• Their activity in activating transcription of reporter genes linked to a promoter with an MRE (DNA binding target of MBP) or a GalRE upstream of the reporter was measured.
• Relative expression levels are shown at right.
• Interaction with general transcription factors can be through interaction with TAF's, through interaction with a protein complex, called mediator, that is bound to the C-terminal tail of RNA polymerase II or by other means.

Interpretations

• The part of the polypeptide responsible for activation is separate from the region required for DNA binding. Each is called a domain.
• Transcription regulators also bind other molecules that transduce signals. Likely, they have a third domain responsible for this binding.

Further information

• Many transcription regulators have been shown to have separable DNA binding and activation domains.
• As expected, binding of effector molecules is usually due to the presence of a third domain, a signal transducing domain.
• Further experiments show that the binding and activation domains of gal4 don't need to be covalently attached to one another. Activation of transcription occurs when the gal4 DNA binding domain is attached to a protein that interacts with another protein that is covalently attached to the gal4 activation domain. This is the basis for the two hybrid system for isolating genes for proteins that interact with other proteins. Reporter genes are used to report positive interactions.

This is page 2220 of Molecular Genetics by Ulrich Melcher, © 1997-1999, 2001, 2003