Transcription Co-activator Family Proteins Combiz Richard Abdolrahimi 06-09-2005 HC 70AL Gene Discovery Lab Dr. Bob Goldberg UCLA/HHMI Institute

What are the Structures of RNA and DNA?

What is Transcription/Transcription Factors? Transcription Factors: Any protein required to initiate or regulate transcription RNA Polymerase II Transcription factor activity-enhancer binding, functions to initiate or regulate RNA polymerase II by binding a promoter or enhancer region of DNA During transcription, mRNA is synthesized following the sequence of nucleotides in the coding strand of DNA. This will require the enzyme RNA polymerase and nucleotides from which RNA will be constructed. RNA polymerase will unwind the DNA, separate the strands, and assemble the mRNA molecule using the coding strand of DNA as a template.

TATA BOX & Function of Transcription Co-Activators When a protein binds to a promoter, it automatically activates transcription? Not entirely, it needs the help of transcription co-activators Regulating when/where gene is transcribed Stop Stop TATA Box 3’ UTR 5’ UTR 3’ UTR Promoter of the Gene: “On Switch”

Hypothetical Example of the influence of Transcription Co-Activators Structure of the upstream region of a typical eukaryotic mRNA gene that hypothetically contains 2 exons and a single intron. The diagram indicates the TATA-box and CCAAT-box basal elements at positions -25 and -100, respectively. The transcription factor TFIID has been shown to be the TATA-box binding protein, TBP. Several additional transcription factor binding sites have been included and shown to reside upstream of the 2 basal elements and of the transcriptional start site. The location and order of the variously indicated transcription factor-binding sites is only diagrammatic and not indicative as being typical of all eukaryotic mRNA genes. There exists a vast array of different transcription factors that regulate the transcription of all 3 classes of eukaryotic gene encoding the mRNAs, tRNAs and rRNAs.

What do Transcription Co-activators do? Why are they important? Transcription co-activators activate transcription from a RNA polymerase II promoter, does not bind DNA itself 43 genes in the Arabidopsis Genome that have proteins that regulate transcription How do you know when/where your gene of interest is transcribed in plants? Activator proteins bind to genes known as enhancers which help determine which genes are switched on and speed up transcription. Repressor proteins bind to genes called silencers which interfere with activator proteins and slow down transcription. “Co-activators, adapter molecules which coordinate signals from activator and repressor proteins, relay information to basal factors which then tell RNA polymerase where and when to start transcription.” http://www.faculty.uaf.edu/ffdew2/geneticsWeb/lecture/lecture10.pdf

A Patent on Transcription Co-activator PC4 proteins FIELD OF THE INVENTION [0002] This invention is in the field of plant molecular biology. More specifically, this invention pertains to nucleic acid fragments encoding PC4 transcription coactivators in plants and seeds. BACKGROUND OF THE INVENTION [0003] Activation of transcription in eukaryotes depends upon the interplay between sequence specific transcriptional activators and general transcription factors. While direct contacts between activators and general factors have been demonstrated in vitro, an additional class of proteins, termed coactivators, appear to be required for transcriptional activation of some genes. For example, transcription of class II genes depends upon the assembly of basal transcription machinery containing RNA polymerase II and the general transcription factors (GTFs): TFIIA, TFIIB, TFIID, TFIIE, TFIIF, and TFIIH. Class II genes contain core-promoter elements recognized by the general transcription factors and gene-specific sequences recognized by the activators. Coactivators mediate the interaction between the transcriptional activators the GTFs. Transcription activation is the output of the interaction between the sequence-specific activator and basal transcription machinery, which increases the efficiency and/or stability of the entire transcription machinery complex. [0004] The positive cofactor 4 (PC4) functions as both an activator-dependent, and a general transcription factor-dependent coactivator. It interacts with activation domains such as VP16 and the general transcription factors such as TFIIA, TFIIB, TFIIH and TAFs in TFIID. PC4 is a bridge or signal mediator between a set of specific activators and general transcription factors in transcription initiation complex (Wu et al. (1998): EMBO J. 17:4478-4490; and Zhu et al. (1995) Plant Cell 7:1681-1689.) Positive Cofactor 4 has been purified from the Upstream Stimulatory Fraction of HeLa cells and found to mediate activator dependent transcriptional activation. PC4 has been demonstrated to be a promiscuous and potent coactivator interactng with several activators, including Ga14NP 16. PC4 itself is a non-specific DNA binding protein that binds to both ssDNA and dsDNA, but has a higher affinity for ssDNA (Ge et al. (1994) Cell 78:513-523; Henry et al. (1996) J. Biol. Chem. 271:21842-21847; Kaiser et al. (1995) EMBO J. 14:3520-3527; Kretzschmar et al. (1994) Cell 78:525-534; and Werten et al. (1998) EMBO J. 5:5103-51 11. PC4 has also been shown to interact with members of the basal transcriptional machinery. Specifically, the TFIIA-DNA and TFIIA-TFIIB-DNA complexes. Phosphorylation of PC4 by TFIIH or TATA associated factors abolish PC4 DNA-binding activity. Additionally, PC4 and Ga14/VP 16 have been shown to be required during TFIID-TFIIA-DNA complex formation (D-A complex) in order to stimulate transcription. This ability to affect D-A complex formation is linked to PC4's dsDNA-binding characteristic. www.freshpatents.com

Summary of Transcription Co-Activators in Arabidopsis thaliana Identification of transcription co-activators and factors are a necessary prerequisite for the study of the molecular mechanisms of gene activation in plants KIWI and KELP: 2 putative transcriptional co-activators from Arabidopsis Single-stranded DNA binding activity of PC4/p15 and all proteins between 98 – 165 amino acids in length Transcription factors include many protein families like MYBs and …more Research is currently being conducted to gather more information on these pertinent co-activators