Research

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Alison Pidoux 2012

Centromeres and Kinetochores

Eukaryotic organisms segregate their chromosomes to newly forming daughter cells by attaching the chromosomes to the microtubules of the mitotic spindle. The nucleoprotein complex that binds chromosomes to the spindle is termed the kinetochore and the site of kinetochore assembly on each chromosome is the centromere. Our work focuses on how cells establish and maintain centromeres and how centromeres promote the formation of kinetochores as cells enter mitosis. Of particular interest is how genetic and epigenetic information defines the position of the centromere on each chromosome. One key epigenetic feature of centromeres in many eukaryotes is the replacement of histone H3 in centromeres with a centromere specific histone variant termed CEntromere Protein A (CENP-A). We study the mechanisms of CENP-A chromatin assembly at centromeres, how the underlying DNA of the centromere impacts the process and how CENP-A chromatin is utilized to build the centromere and kinetochore.

People: Eline Hendrix, Jacob Schwartz, Kelsey Fryer, Rae Brown

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David Goodsell 2013

Non-coding RNAs

The majority of DNA sequences in the human genome are transcribed into RNA yet only a small fraction of those RNAs encode protein. Generally termed noncoding RNAs, these transcripts play diverse and essential roles in cells. We have been working to understand how one class of these noncoding RNAs, chromosome associated RNA, interacts with and regulates the function of the genome. We are interested in how chromatin associated RNAs govern the differentiation and development of embryonic stem cells into different cell types. We are also studying how noncoding RNAs help to control the transcription of repetitive elements in the genome and the function of repeat transcripts in the biology of cells. Toward this end we have developed a novel method termed Chromatin Associated RNA Sequencing (ChAR-Seq) that enables the identification of all RNAs in a cell that bind to chromatin and maps their binding sites genome wide. We are using a combination of this genomic approach with microscopic and biochemical approaches to understand how noncoding RNAs bind to and regulate chromosomes.

People: Kelsey Fryer

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Genome Stability

The replication, expression, repair, and segregation of genomes pose major challenges to the cell to ensure the stability of the genome. Errors in these processes that cause genome damage or chromosome missegregation lead to mutation and aneuploidy that promote tumorigenesis. Mutations in proteins that correct these errors result in susceptibility to genetic disease and cancer. During transcription, RNA polymerase melts the DNA double helix as the nascent RNA is generated. Normally this process resolves through transcriptional termination but in cases where RNA polymerase stalls, an R-loop can form in which the nascent RNA re-hybridizes to the DNA, displacing the opposite DNA strand. These lesions are known to cause DNA damage and mutations in proteins that process and repair these structures including RNaseH1 and the breast cancer genes BRCA1 and BRCA2, are associated with human disease. In collaboration with the lab of Karlene Cimprich we are working to understand how R-loops are generated and processed and how BRCA1/2, RNaseH and other repair factors resolve these structures to maintain genome stability. 

People: Pragya Sidhwani

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Lois DeAntonio 2020

Chromosome Structure

The 3-dimensional organization of the genome compartmentalizes chromosomes into functional domains. That chromosomes are not randomly organized in the nucleus has been appreciated since the early studies of Rabl and Boveri in the late 19th century. Interactions between chromosomes and between chromosomes and nuclear compartments position different chromosomes in nuclear space. Interactions between different regions of individual chromosomes help to organize and fold the genome as well as segregate domains such as heterochromatin from transcriptionally active regions of the genome. We are interested in understanding the molecular mechanisms that drive the organization of genomes into functional domains. One example of this organization is the coalescence of centromeric nucleosomes into a cohesive unit at the centromere. CENP-A nucleosomes are interspersed with H3 nucleosomes yet are brought together into close proximity while excluding H3 chromatin. It is thought that this orientation is important for microtubule capture and for resisting force applied at the kinetochore during chromosome segregation. How the cell brings together the active centromeric nucleosomes into a cohesive unit is unknown. We are working to understand how the proteins of the centromere can bring together centromeric nucleosomes in 3-dimensions even when those nucleosomes are distant from one another along the DNA strand.

People: Jacob Schwartz, Eline Hendrix, Rae Brown, Alex Leffell

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Joy Division 1979

Epigenetic Regulation

Centromeres contain CENP-A-rich “core” centromeric domains surrounded by heterochromatin-rich pericentromeric domains. Despite nucleosome disassembly during replication, both regions are heritably maintained through cell divisions. Our group is focused on 1) how the centromere is epigenetically defined and maintained by centromeric chromatin, 2) how heterochromatin is formed at repetitive regions of genomes, 3) how chromatin associated RNAs regulate epigenetic states and 4) how chromatin states change during embryonic stem cell differentiation gives rise to specific cell types. In collaboration with the Streets lab at UC Berkeley and the Altemose lab at Stanford, we have developed a technique called Directed Methylation and Long read Sequencing (DiMeLo-seq) to study epigenetic signatures at repetitive regions of the genome at single molecule resolution. 

People: Rae Brown, Kelsey Fryer, Pragya Sidhwani