The long-term goal of this application is to understand how chromatin structure influences gene regulation. The simple idea that all genes are packaged, irrespective of their sequence, into uniform solenoid-like chromatin structures that must be disrupted in order for gene activation to occur may not be correct. Instead, DNA may be packaged in a sequence-specific fashion. Different nucleosome arrangements may lead to the formation of different chromatin structures that facilitate gene regulation. DNA sequence variations in certain regions of non-coding DNA among individuals might influence gene expression and disease susceptibility. Preliminary findings suggest that there are signals in vertebrate DNA that influence nucleosome array formation. Specifically, there is evidence that the nucleotide triplet consensus non-T, A/T, G (VWG) existing at ten base pair multiples is a strong nucleosome localization signal. The hypothesis that oscillations specifically in periodic VWGs influences nucleosome alignment into ordered arrays will be tested using an in vitro chromatin assembly system. The VWG signal in regions of genes that strongly align nucleosomes will be disrupted by insertions, deletions, and by site-directed mutagenesis. The hypothesis that DNA sequences that exhibit strong oscillations in the average period- 10 VWG count with a dinucleosome period strongly align nucleosomes into ordered arrays, while sequences that are aperiodic in VWG do not, will be tested in vitro and in nuclei. The hypothesis that VWGs influence nucleosome positioning through anisotropic bendability will be tested. The hypothesis that the degree of regularity of the nucleosome array influences the chromatin higher-order structure will be tested using analytical sedimentation and electron microscopy. It will be determined whether the presence or absence of strong nucleosome alignment signals in transgenes influences their transcription in the mouse. Information obtained here might ultimately enable one to predict functionally important aspects of chromatin structure computationally from DNA sequence data.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM062857-03
Application #
6636604
Study Section
Cell Development and Function Integrated Review Group (CDF)
Program Officer
Carter, Anthony D
Project Start
2001-05-01
Project End
2006-04-30
Budget Start
2003-05-01
Budget End
2006-04-30
Support Year
3
Fiscal Year
2003
Total Cost
$175,877
Indirect Cost
Name
Purdue University
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
072051394
City
West Lafayette
State
IN
Country
United States
Zip Code
47907
Cioffi, Alfred; Fleury, Tomara J; Stein, Arnold (2006) Aspects of large-scale chromatin structures in mouse liver nuclei can be predicted from the DNA sequence. Nucleic Acids Res 34:1974-81
Fleury, Tomara J; Cioffi, Alfred; Stein, Arnold (2006) Detecting nucleosome ladders on unique DNA sequences in mouse liver nuclei. Methods Mol Biol 338:209-24
Fan, Yuhong; Nikitina, Tatiana; Zhao, Jie et al. (2005) Histone H1 depletion in mammals alters global chromatin structure but causes specific changes in gene regulation. Cell 123:1199-212
Dalal, Yamini; Fleury, Tomara J; Cioffi, Alfred et al. (2005) Long-range oscillation in a periodic DNA sequence motif may influence nucleosome array formation. Nucleic Acids Res 33:934-45
Cioffi, Alfred; Dalal, Yamini; Stein, Arnold (2004) DNA sequence alterations affect nucleosome array formation of the chicken ovalbumin gene. Biochemistry 43:6709-22
Stein, A; Dalal, Y; Fleury, T J (2002) Circle ligation of in vitro assembled chromatin indicates a highly flexible structure. Nucleic Acids Res 30:5103-9