Progress and challenges in profiling the dynamics of chromatin and transcription factor binding with DNA microarrays

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ChIP-chip, or chromatin immunoprecipitation followed by DNA microarray analysis, has proven to be an efficient means of mapping protein-genome interactions. Recent experiments using this tool are beginning to reveal the complex dynamics of transcription factor binding and chromatin organization, and how these processes interact with each other to generate a cellular response to environmental and developmental cues. Data derived from this approach, particularly data involving chromatin components and histone modifications, might be affected by assumptions underlying the procedure, and the data might be made more useful by adoption of standardized whole-genome microarray platforms.

Introduction

The development of DNA microarrays has provided a detailed view of how cells coordinate the transcription of their genome in response to environmental changes and developmental cues. As informative and spectacular as those global views are, measurement of RNA levels provides an indirect readout of the transcription factor activity and chromatin dynamics underlying those responses. To probe the interaction between proteins and DNA directly, a technique fusing chromatin immunoprecipitation (ChIP) and microarray analysis (chip), known as ChIP-chip, was developed (Figure 1a) 1., 2.. Early ChIP-chip studies examined the genome-wide location of several well-studied yeast transcription factors including Ste12p, Gal4p, Swi4p, Swi6p and Rap1p 1., 2., 3. and the binding pattern of several transcription factors required for cell-cycle progression 4., 5.. In addition to a large number of novel binding sites, these initial transcription factor localization studies identified known binding sites, empirically verifying the effectiveness of the method 1., 2., 3.. Since then, ChIP-chip has been applied to scores of transcription factors in organisms ranging from bacteria to humans.

In this review, we highlight some of the recent papers that explore how patterns of transcription factor binding and chromatin organization change under different biological conditions. We then discuss some of the possible limitations of the technique, particularly as they relate to mapping chromatin components and histone modifications.

Section snippets

Profiling the dynamics of transcription factor binding

Recent ChIP-chip studies have begun to address the question of how the cell is able to use a single transcription factor to elicit multiple downstream transcriptional responses (Figure 1b). This complex issue is one of the fundamental, over-arching questions in the field. Zeitlinger et al. [6••] have made an important contribution using the Saccharomyces cerevisiae transcription factor Ste12p as a model. Ste12p has multiple functions in the cell. In the presence of mating pheromone, Ste12p

ChIP-chip analysis of the general transcription machinery

In addition to examining the profile of sequence-specific DNA binding proteins, the occupancy of the general transcription machinery has been probed by ChIP-chip. These studies are important because recruitment of general transcription machinery is often the endpoint in a cascade of regulatory events. Progress in this area can be illustrated by several recent ChIP-chip studies designed to elucidate the set of transcripts regulated by RNA Polymerase (Pol) III in yeast 9.•, 10.•, 11.•. The

ChIP-chip analysis of chromatin structure

In addition to transcription factor binding, ChIP-chip has been used to monitor the distribution of the structural components of chromatin 12., 13., 14.•, 15.••, enzymes involved in histone modification 16., 17., 18.•, and post-translational histone modifications themselves 19., 20., 21.•, 22.••. These chromatin components and determinants of chromatin dynamics are often important regulators of transcription (Figure 1c; reviewed in [23]).

The challenges of applying ChIP-chip to chromatin

The ChIP-chip protocol has been largely standardized (Figure 1a), and is now used routinely by laboratories pursuing a wide spectrum of biological research. Despite the outward appearance of a simple and streamlined method, there are technical aspects of the procedure that are difficult to control for experimentally and which could complicate analysis (Figure 2). These factors include fixation, epitope accessibility, antibody specificity and choice of microarray content. All ChIP-chip

Conclusions

As evidenced by the exciting advances outlined in the first half of this review, ChIP-chip will continue to be the method of choice for uncovering relationships between transcription-factor dynamics and chromatin organization. The challenges described in the second half of this review do not diminish the power of ChIP-chip, which has been thoroughly demonstrated. Rather, we call attention to the possibility that there are underlying technical considerations that may affect interpretations of

Update

Recently, ChIP-chip was used to map the distribution of cohesin proteins on four Saccharomyces cerevisiae and two Schizosaccharomyces pombe chromosomes [48]. The authors report that, in both species, cohesins are primarily localized to intergenic regions between convergently transcribed genes and that this positioning is dependent on active transcription of the genes. These results are similar to the findings of Glynn et al. [14] and further illustrate both the dynamic positioning of cohesins

References and recommended reading

Papers of particular interest, published within the annual period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

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