Alpha satellite at the centromere: It’s not junk DNA!
The centromere is a specialized chromosomal site involved in chromosome architecture and movement, kinetochore function, heterochromatin assembly, and sister chromatid cohesion. Our experiments have uncovered a unique type of chromatin (CEN chromatin) formed exclusively at the centromere by replacement of core histone H3 with the histone variant CENP-A. We are exploring the composition of CEN chromatin, its relationship to the underlying highly repetitive alpha satellite DNA, and the dynamics of CENP-A loading and distribution during mitosis using a variety of cell biological, molecular, genomic, and chromosome engineering approaches.
Non-coding RNAs produced from human alpha satellite arrays
Our lab is also interested in identifying and functionally characterizing non-coding RNAs produced from human alpha satellite arrays. All human chromosome contain at least one region of alpha satellite DNA, a homogenous repetitive array that can extend up to 5 megabases in size. The centromere, defined by CENP-A chromatin and other centromere proteins, is assembled on alpha satellite DNA. Although the DNA sequence itself does not appear to be necessary or sufficient for centromere identity, other features of alpha satellite may be important for centromere function. We have identified alpha satellite transcripts that are chromosome-specific and array-specific and tested their role(s) in centromere architecture and function. Our ongoing work is focused on understanding their distinct interactions with centromere proteins and relevance to centromere variation.
Genomic variation within repetitive DNA and centromeric epialleles
Some human chromosomes have more than one distinct alpha satellite array. We showed using human chromosome 17 as a model that, depending on the individual, the centromere can form at either array. The molecular basis for these centromeric epialleles and such centromere plasticity is not clear. We discovered that the amount of genomic variation within alpha satellite DNA correlates with where a centromere is formed on chromosome 17 and affects how stable the chromosome is. Variation within the repetitive portion of the human genome has not been well studied, primarily because alpha satellite DNA is part of the 10% of the human genome that has been excluded from the contiguous genome assembly. Our studies emphasize that the repetitive portion of the genome is rich in unappreciated variation that has functional significance. Furthermore, alpha satellite variation could be a useful biomarker for aneuploidy, a hallmark of many cancers. We are currently using endogenous chromosomes and human artificial chromosomes (HACs) to investigate how alpha satellite variation affects centromeric transcription, recruitment of centromere proteins, de novo centromere assembly, kinetochore architecture, and chromosome stability.
Dicentric chromosomes: Formation and fate
The lab also studies dicentric chromosomes, that is, genome rearrangements that produce chromosomes that have two centromeres. Originally described by Nobel prizewinner Barbara McClintock in the 1930s, dicentrics were considered to be inherently unstable chromosomes that trigger genome instability and cancer. Dicentric chromosomes in humans, however, are very stable and are often transmitted through multigenerational families. The mechanisms by which human dicentric chromosomes are stabilized are unclear. To understand mechanisms of dicentric stabilization, we use multiple chromosome engineering approaches to experimentally produce dicentrics in human cells. We have observed that some dicentrics undergo inactivation of one centromere within 20 weeks after formation, while other dicentrics retain two active centromeres and do not exhibit chromosome instability. These latter dicentrics are particularly intriguing, so we are investigating the molecular basis of dicentric stabilization immediately after formation using quantitative microscopy and high resolution imaging.