7.342: Week 3 Questions

Post discussion, questions, or comments about the Week 3 course material here.

Amber

Question for Brownell et al:

At the end of the section Tetrahymena is Homologous to Yeast Gcn5p, the authors say that Gcn5p "migrates anomolously in SDS gels". Why would that happen?

Question for Taunton et al:

I noticed that they related RbAp48, the protein that binds to the retinoblastoma gene product, to histone deacetylase. They also found that inhibition of the deacetylase arrests the cell cycle. Could any of the research be useful for cancer treatment?

Georgi

Comments from the second paper (Taunton et al)

There are three classes of HDAC proteins with class I being homologous to Rpd3 and containing 4 members (H. Santos-Rosa, C. Caldas/ European Journal of Cancer 41 (2005) 2381-2402). Why did they identified only one protein? Maybe some of the others were among the six bands that initialy appeared?

They predicted that RbAp48 is an adaptor subunit targeting HDAC to chromatin domains; it is found in the HATB complex in the cyoplasm, with CAF-1 in the hucleus, participating in nuclesome assembly and with HDAC. What is its precise role in all these processes?

Also, can we have a more in depth discussion about the various HAT and HDAC complexes, their specific functions, etc.?

Holly

Taunton et al paper: Why did RbAp48 not co-precipitate with the recombinant HD 1 protein? Did this mean that the HD 1 was not targeted effectively? What would the effect of this be on the cells?

Brownell et al paper: It is suggested that a conserved bromodomain is a source of HAT A specificity. Presumably the genes that need transcribing change during the lifetime of the cell (e.g. during differentiation, apoptosis etc) – how is this accommodated? Is the bromodomain modified in any way?

Kathy

Taunton: How does the charcoal precipitation assay work? Has any subsequent work regarding "cell cycle checkpoints may exist that monitor histone acetylation" been done?

Brownell: I don't quite understand why this paper was important. It had already been known that histone acetylation was important for transcription. The main finding seems to be of this conserved bromodomain, but it seemed like the authors only speculated/theorized on its function. Did they really present any experimental evidence that this domain is indeed what directs HAT A to specific sites? What data did Marcus et al, 1994 and Georgakopoulos et al, 1995 show?

Manpreet

Comments from the first paper (Brownell et al)

The authors show that p55 and Gcn5p both contain bromodomain; and that Hat1p doesn't possess a bromodomain. They speculate that the bromodomain tethers HAT A to other factors at specific chromosomal sites. This is remarkably accurate - later research has shown that bromodomains bind to acetylated lysine residues; helping regulate transciptional remodelling and transciptional activation (Zeng, L., Zhou, M.M. (2002)Bromodomain: an acetyl lysine binding domain. FEBS Letters. Vol 213, 124-128)

Question about the second paper (Taunton et al.)

On p. 410, they say "RPD3 has yet to be implicated in silencing at telomeres or at the mating loci." Presumably this work has been done since 1996. Do you know what the outcomes were?