Revealing the Mechanism of HSP104 Transcription Initiation in the ...

More documents

Recommendations

Info

(70, 73, 79, 122). It is believed that one of the major events leading to the "opening" of chromatin is the acetylation of histones on conserved lysine residues. This acetylation probably reduces interactions between histones and DNA. Acetylated histones (generally H3 and H4) then facilitate access of other chromatin remodeling factors which impose additional spatial changes on chromatin, followed by the recruitment of basal transcription factors and RNA PolII [reviewed in (28, 78)]. These modifications are actually critical for proper gene activation. The above concept is supported by numerous studies (2, 13, 21, 36, 64, 100, 103), but recent studies reported that on some promoters it is not histone acetylation, but rather histone deacetylation, that is related to the activation of these promoters. Particularly, Deckert and Struhl showed that in yeast, histones H3 and H4 undergo deacetylation on some stress-activated and galactose-induced promoters (31). They further showed that, depending on the inducer, one single promoter can undergo different chromatin modifications. For example, an increase in histone acetylation in response to one stress and a decrease in histone acetylation in response to another stimulus (31). Following chromatin remodeling and establishment of the preinitiation complex on the promoter, transcription initiation could be further enhanced by coactivator complexes such as SRB/MED (8, 53, 63, 125). It should be appreciated that a plethora of factors could join any of these multi-protein complexes changing their composition and catalytic properties from one promoter to another. The transcriptional activators responsible for initiating the cascade of events leading to transcription initiation are even more specific, usually involved in activation of a limited number of genes. Many different transcriptional activators are expressed in the cell, each responding to a narrow subset of signals, and some to a single signal (i.e., hormones, growth factors). The different modifications occurring on the chromatin and the enzymes involved in these modifications are also different from promoter to promoter. Similarly, components composing the co-activator complex SRB/MED could also vary on promoters [reviewed in (8, 10)] and even components making up the RNA PolII holoenzyme could differ according to cellular conditions and the particular promoter. In fact, even one subunit of RNA PolII, Rpb4, is dispensable for cell proliferation and seems to be required only under stress conditions (23, 89, 97, 98). 5
Transcription under stress in S.cerevisiae Although several signal transduction pathways and some stress-induced transcriptional activators have been identified (37, 38, 41, 87, 92, 96, 102, 133), we have only partial answers to some of the major questions raised above with respect to sensing the stress and in turn activating transcription. This issue is nevertheless better understood in S.cerevisiae than in any other experimental system. A large number of stress responsive systems have been discovered in this organism including several transcriptional activators whose activity is induced by specific stresses (e.g. yAP1, Gcn4, Gln3) (41, 84, 88, 92). Another activator, Hsf1, that is induced mainly by elevated temperature, but also by other stresses [(52, 83, 90, 133) and see below] and yet two more activators, Msn2 and Msn4 that are activated in response to any stress [(17, 87, 110) and see below]. As promoters are usually complex, containing several enhancer elements, it is most probable that none of these activators is acting alone on target promoters, but cooperate with one of the other stress-induced activators (4, 29, 47), or with other activators, not necessarily induced by stress (39, 53, 65, 72, 80, 93, 123). It is still unclear how two or more activators co-act on the same promoter. Recent studies addressed the changes in the organization of chromatin that occur on stress responsive promoters upon activation (21, 36, 128, 135, 136). It was found that many promoters undergo extensive chromatin remodeling (i.e., nucleosomal disassembly following histone acetylation) upon activation and that the complexes responsible for this modification are in fact recruited by transcriptional activators (21, 36, 128, 135, 136). It should be noted that most studies address the question at the whole genome level and their conclusions are therefore grossly general. The epistatic relationships between recruitment of transcriptional activators and changes in chromatin structure are not well established in many cases. Also, it is also not fully understood how RNA PolII and the factors of the basal transcription machinery function under stresses such as heat shock, when many proteins are denatured. One of the RNA PolII subunits, Rpb4, is essential only under stress, and seems to be involved in the induction of some stress related genes (23, 89, 97, 98). It may also function as a stabilizer of RNA PolII under stress (23, 104). It is not clear whether other components of the PIC are specifically important for transcription under stress. In an attempt to understand these aspects of the mechanisms of transcriptional activation under stress, we have been focusing on the HSP104 promoter. This 6
Page 1 and 2: Revealing the Mechanism of HSP104 T
Page 3 and 4: This thesis is dedicated to my fami
Page 5 and 6: genetic approach, we identified com
Page 7 and 8: INTRODUCTION The cellular stress re
Page 9: for their transcription, some 16% o
Page 13 and 14: In mammalian cells, Hsf1 is a monom
Page 15 and 16: Hsf1 and Msn2/4 can exclusively or
Page 17 and 18: A) 5’-XhoI -713 BamHI-3’ ATG La
Page 19 and 20: 1 We first noticed that the activit
Page 21 and 22: 1 1 A) B) 700 600 500 ras2∆msn2
Page 23 and 24: Table 1. Summary of the role of var
Page 25 and 26: same enzymes. BS-HA-HSF was obtaine
Page 27 and 28: β-Galactosidase assay Overnight cu
Page 29 and 30: to the fact that the HSEs are prese
Page 31 and 32: The systematic 5’ deletion analys
Page 33 and 34: 1 A) units 100.00 90.00 80.00 70.00
Page 35 and 36: nucleosome disassembly involving sp
Page 37 and 38: A) Strain W.T. msn2∆msn4∆ ras2
Page 39 and 40: Hsf1 constitutively binds the HSP10
Page 41 and 42: Next, we attempted to directly meas
Page 43 and 44: Table 3. List of mutants in which r
Page 45 and 46: Table 4. Strains required for posit
Page 47 and 48: Overall, the results nevertheless s
Page 49 and 50: Regulation of Hsf1 As explained in
Page 51 and 52: Table 7. Comparison between HSE-Lac
Page 53 and 54: promoter. III) The identification o
Page 55 and 56: to stress. This change in nucleosom
Page 57 and 58: perhaps not surprisingly, overexpre
Page 59 and 60: 21. Chandy, M., J. L. Gutierrez, P.
Page 61 and 62:
56. Hietakangas, V., J. K. Ahlskog,
Page 63 and 64:
89. Miyao, T., J. D. Barnett, and N
Page 65 and 66:
121. Stanhill, A., N. Schick, and D
show all

Revealing the Mechanism of HSP104 Transcription Initiation in the ...

You also want an ePaper? Increase the reach of your titles

Delete template?

Save as template?