# Administrative Supplement to : Functional crosstalk between the Fanconi Anemia and ATRX/DAXX histone chaperone pathways

> **NIH NIH R01** · UNIVERSITY OF MINNESOTA · 2021 · $59,228

## Abstract

PROJECT SUMMARY
Studies from our laboratory provided evidence that the Fanconi Anemia (FA) and Alpha Thalassemia Retardation
Syndrome X-linked (ATRX) pathways are interconnected to promote genome stability in the face of DNA
replication stress by supporting homologous recombination (HR) mechanisms. Our previous observations led us
to hypothesized that ATRX cooperates with FANCD2 to promote HR-mediated DNA replication fork rescue, but
also possesses additional, independent activities to support a distinct subset of DNA repair steps.
So far, our study results were mostly obtained using human cancer-derived cell lines. Intriguingly, our recent
findings indicate that the FANCD2- and ATRX-dependent DNA replication stress responses activities differ
mechanistically between human cancer cells and non-cancer (“normal”) cells.
To test our hypothesis, we propose to investigate FANCD2 and ATRX functions in normal human cell lines,
namely RPE1 and BJ cells. To this end, we will generate normal human cells genetically null for ATRX,
FANCD2, or both, as well as cells carrying ATRX loss-of-function mutations. Importantly, efficient ATRX gene
knock-out or knock-in in normal human cells requires the use of a gentle cell sorter that (a) allows single live
cell sorting while maintaining high ATRX mutant cell viability and (b) allows for specific bulk purification of S-
phase cells to increase subsequent ATRX knock-in targeting efficiency. We propose to use a Sony benchtop
SH800 cell sorter equipped with two lasers and microfluidics sorting chips, in order to:
 (1) Perform single live cell sorting of CRISPR/Cas9 targeted, ATRX- and ATRX/FANCD2-null RPE1 and
 BJ cells
 (2) Perform S-phase-specific bulk pre-sorting of RPE1 and BJ cells, followed by CRISPR/Cas9 mediated
 ATRX gene knock-in.
Generating these mutant RPE1 and BJ cells utilizing the SONY SH800 sorter will then allow us to (i) elucidate
the molecular and structural makeup of ATRX-FANCD2 protein complexes; (ii) Determine molecular
mechanisms of ATRX/FANCD2-mediated replication fork recovery; (iii) Dissect FANCD2-dependent and -
independent roles of ATRX during DNA repair.

## Key facts

- **NIH application ID:** 10387846
- **Project number:** 3R01GM132596-03S1
- **Recipient organization:** UNIVERSITY OF MINNESOTA
- **Principal Investigator:** Alexandra Theresia Sobeck
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $59,228
- **Award type:** 3
- **Project period:** 2019-05-01 → 2023-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10387846

## Citation

> US National Institutes of Health, RePORTER application 10387846, Administrative Supplement to : Functional crosstalk between the Fanconi Anemia and ATRX/DAXX histone chaperone pathways (3R01GM132596-03S1). Retrieved via AI Analytics 2026-05-28 from https://api.ai-analytics.org/grant/nih/10387846. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*