Daniel R. Foltz
Primary AppointmentAssociate Professor,
- BA, Physiology, University of Minnesota, Minneapolis, Mn
- PhD, Cell Biology, Northwestern University, Chicago, IL
- Postdoc, Ludwig Institute for Cancer Research
Assembly and Function of Centromeric Chromatin
The faithful segregation of chromosomes through each round of cell division is imperative to guard the human genome against errors in chromosome number (a.k.a. chromosomal instability) that can lead to birth defects, cell death or cancer. Chromosomal instability is not only a hallmark of cancer but can be an instigating event in the disease. Integral to the process of chromosome segregation is a specialized chromatin domain known as the centromere. The centromere directs the assembly of the kinetochore during mitosis, which in turn mediates and senses the interaction between the chromosome and spindle microtubules. The most basic unit of the centromere is a unique nucleosome containing Centromere protein- A (CENP-A), which replaces histone H3 within the histone octamer (and includes histones H4, H2A and H2B). This CENP-A nucleosome is present specifically in centromeric chromatin. We are interested in the mechanism by which the CENP-A nucleosome directs the assembly of the centromere and contributes to proper chromosome segregation. Using a proteomics approach we have identified the CENP-A nucleosome associated complex (CENP-ANAC, an assemblage of six centromere proteins (CENP's) as the most proximal component of the human centromere. We are now working to determine how this complex is assembled, how it distinguishes CENP-A nucleosomes from general histone H3 containing chromatin as well as how these proteins contribute both individually and as a complex to the stable segregation of chromosome and avoid a state of aneuploidy.
Molecular Cellular and Developmental Biology
The process of new centromeric nucleosome assembly is a cell cycle regulated process. Incorporation of new CENP-A nucleosomes is tightly restricted to the early-G1 phase of the cell cycle and requires the cells to successfully transit through mitosis. We are interested in the mechanism by which this process is temporally controlled and the consequences of misregulating this process. In addition, the centromere exerts its recognized function during mitosis, at the time when chromosomes are interacting with microtubules of the mitotic spindle in order to provide the motive forces required for segregating sister chromatids into daughter cells. However, the CENP-ANAC is present at the centromere throughout the cell cycle. We are interested in determining what function this complex plays during interphase, and how that function may affect the ability of the centromere to fulfill its roll during mitosis.
Biochemistry, Molecular Biology and Genetics
The mechanism by which a region of the chromosome is specified as a centromere and how centromeric chromatin assembly is accomplished is another focus of our lab. The location of the mammalian centromere on each chromosome is stably inherited through each cell cycle however, its location is not dictated by the underlying DNA sequence, and therefore the mechanism of centromere identification is thought to be epigenetic. Epigenetic inheritance can be governed by histone modification or variant histone incorporation, both of which may play a role in centromere identity. Each round of DNA synthesis presents a challenge for its stable propagation, since replication of the chromosome requires that new CENP-A nucleosomes are assembled in the proper location in order to maintain the epigenetic mark. We seek to understand how centromeric chromatin is assembled and how this process is restricted to sites already determined to be active centromeres.
Our laboratory uses a combination of biochemical, molecular biological and microscopic techniques in order to investigate the structure and function of the proteins that comprise the human centromere. Specifically, we are using high power fluorescence microscopy on living and fixed mammalian cells in conjunction with siRNA mediated gene silencing to examine centromere assembly. Proteomics using tandem affinity purification of epitope-tagged complexes from mammalian cells coupled with mass spectrometry is used to investigate the complexes that are required for the process of centromere specification. In addition, we use a variety of classic molecular biology and biochemistry techniques including bacterial protein expression, chromatography and in vitro complex assembly.