# Genomic Analysis of Centromere Assembly and Function

> **NIH NIH R01** · DUKE UNIVERSITY · 2022 · $384,796

## Abstract

Centromeres are essential to genome inheritance. Abnormal centromere function is associated with birth defects,
infertility, and cancer. Human centromeric (CEN) chromatin typically forms on alpha satellite DNA, a 171bp
monomeric sequence that is organized into tandem arrays extending for several megabases. The Telomere-to-
Telomere (T2T) Consortium that recently produced the first complete human genome assembly revealed that
nearly all endogenous human chromosomes contain multiple alpha satellite arrays. We have shown that on some
chromosomes, multiple arrays are competent for centromere assembly. Using Homo sapiens chromosome 17
(HSA17) as a model, we demonstrated that centromere location is dictated by genomic variation within alpha
satellite DNA. On HSA17 that has three distinct higher order repeat (HOR) unit arrays, when the largest array
contains size and sequence variation, centromere assembly shifts to a nearby array. If the centromere forms at a
highly variant array, fewer centromere proteins are present and the chromosome experiences instability. We
hypothesize that placement/organization of variant HORs within an alpha satellite array influences centromere
location and kinetochore assembly. Since most human chromosomes must choose between two (or more) sites
at which to build a stable centromere, our work addresses a fundamental gap in the knowledge of basic processes
that influence centromere location, competence, and long-term stability. However, we still lack a comprehensive
view of the extent of alpha satellite variation within the population and thus the range of functional centromere
outcomes. In this competing renewal application, the proposed work builds on our classification of specific alpha
satellite variants identified in diverse human populations to assemble stable, de novo centromeres. We will also
systematically test centromere competency of long-range alpha satellite organization, coupled with variant
content and proximity to mobile elements. Our project goals are to: 1) produce new genomic assemblies of
functionally characterized centromeres using targeted long read sequencing approaches, 2) use human artificial
chromosome assays to assess competency of different alpha satellite DNA arrangements for de novo centromere
formation, 3) explore the molecular basis for variant centromere defects, and 4) use genome engineering to
rehabilitate and/or rescue defective variant centromeres. Successful completion of this work will result in major
advancement of our basic understanding of genomic variation within large repetitive DNA arrays in humans and
its link to specific centromere outcomes and long-term chromosome maintenance and stability.

## Key facts

- **NIH application ID:** 10520534
- **Project number:** 2R01GM124041-05
- **Recipient organization:** DUKE UNIVERSITY
- **Principal Investigator:** BETH A SULLIVAN
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $384,796
- **Award type:** 2
- **Project period:** 2017-09-01 → 2026-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10520534

## Citation

> US National Institutes of Health, RePORTER application 10520534, Genomic Analysis of Centromere Assembly and Function (2R01GM124041-05). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10520534. Licensed CC0.

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