Inducing and Detecting Mitotic DNA Synthesis at Difficult-to-Replicate Loci

Lorenza Garribba; Wei Wu; Özgün Özer; Rahul Bhowmick; Ian D Hickson; Ying Liu

doi:10.1016/bs.mie.2017.11.025

Inducing and Detecting Mitotic DNA Synthesis at Difficult-to-Replicate Loci

Lorenza Garribba, Wei Wu, Özgün Özer, Rahul Bhowmick, Ian D Hickson, Ying Liu

7 Citations (Scopus)

Abstract

Our conventional understanding of the process of DNA replication is that it occurs in the S-phase of the cell division cycle. However, during investigations into the mechanism by which common fragile sites (CFSs) drive genome instability, we observed that some DNA synthesis was still occurring in early mitosis at these loci. This curious phenomenon of mitotic DNA synthesis (which we now term "MiDAS") appears to be a form of break-induced DNA replication (BIR), a DNA repair process based on homologous recombination that has been characterized in detail only in lower eukaryotes. During MiDAS, it is proposed that parts of the human genome that are not fully replicated when cells enter mitotic prophase complete their replicative cycle at that point. To date, the loci that most depend upon this process are those whose replication can be affected by oncogene-induced DNA replication stress (RS), most notably, CFSs. From our studies, it is clear that the successful completion of MiDAS at CFSs can minimize chromosome missegregation and nondisjunction. Nevertheless, it is still not clear which loci that can undergo MiDAS, whether MiDAS is associated with mutations or genome rearrangements, or whether MiDAS really is a form of BIR. In this review, we describe methods for detecting MiDAS both in prometaphase cells and directly on isolated metaphase chromosomes. In addition, we have included methods for combining MiDAS detection either with immunofluorescence (IF) detection of proteins that are recruited to the MiDAS loci, or with fluorescence in situ hybridization using probes that target specific genomic loci.

Original language	English
Title of host publication	Methods in Enzymology : Mechanisms of DNA Recombination and Genome Rearrangements: Intersection between Homologous Recombination, DNA Replication and DNA Repair
Editors	Marie Spies, Anna Malkova
Number of pages	14
Volume	601
Publisher	Elsevier
Publication date	2018
Pages	45-58
Chapter	3
ISBN (Print)	978-0-12-813979-0
DOIs	https://doi.org/10.1016/bs.mie.2017.11.025
Publication status	Published - 2018

Series	Methods in Enzymology
Number	601
ISSN	0076-6879

Keywords

Common fragile sites
DNA replication stress
EdU
Fluorescence in situ hybridization
Immunofluorescence
Mitosis

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10.1016/bs.mie.2017.11.025

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Garribba, L., Wu, W., Özer, Ö., Bhowmick, R., Hickson, I. D., & Liu, Y. (2018). Inducing and Detecting Mitotic DNA Synthesis at Difficult-to-Replicate Loci. In M. Spies, & A. Malkova (Eds.), Methods in Enzymology: Mechanisms of DNA Recombination and Genome Rearrangements: Intersection between Homologous Recombination, DNA Replication and DNA Repair (Vol. 601, pp. 45-58). Elsevier. https://doi.org/10.1016/bs.mie.2017.11.025

Inducing and Detecting Mitotic DNA Synthesis at Difficult-to-Replicate Loci. / Garribba, Lorenza; Wu, Wei; Özer, Özgün et al.
Methods in Enzymology: Mechanisms of DNA Recombination and Genome Rearrangements: Intersection between Homologous Recombination, DNA Replication and DNA Repair. ed. / Marie Spies; Anna Malkova. Vol. 601 Elsevier, 2018. p. 45-58 (Methods in Enzymology; No. 601).

Research output: Chapter in Book/Report/Conference proceeding › Book chapter › Research › peer-review

Garribba, L, Wu, W, Özer, Ö, Bhowmick, R , Hickson, ID & Liu, Y 2018, Inducing and Detecting Mitotic DNA Synthesis at Difficult-to-Replicate Loci. in M Spies & A Malkova (eds), Methods in Enzymology: Mechanisms of DNA Recombination and Genome Rearrangements: Intersection between Homologous Recombination, DNA Replication and DNA Repair. vol. 601, Elsevier, Methods in Enzymology, no. 601, pp. 45-58. https://doi.org/10.1016/bs.mie.2017.11.025

Garribba L, Wu W, Özer Ö, Bhowmick R , Hickson ID , Liu Y. Inducing and Detecting Mitotic DNA Synthesis at Difficult-to-Replicate Loci. In Spies M, Malkova A, editors, Methods in Enzymology: Mechanisms of DNA Recombination and Genome Rearrangements: Intersection between Homologous Recombination, DNA Replication and DNA Repair. Vol. 601. Elsevier. 2018. p. 45-58. (Methods in Enzymology; No. 601). doi: 10.1016/bs.mie.2017.11.025

Garribba, Lorenza ; Wu, Wei ; Özer, Özgün et al. / Inducing and Detecting Mitotic DNA Synthesis at Difficult-to-Replicate Loci. Methods in Enzymology: Mechanisms of DNA Recombination and Genome Rearrangements: Intersection between Homologous Recombination, DNA Replication and DNA Repair. editor / Marie Spies ; Anna Malkova. Vol. 601 Elsevier, 2018. pp. 45-58 (Methods in Enzymology; No. 601).

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Inducing and Detecting Mitotic DNA Synthesis at Difficult-to-Replicate Loci

Abstract

Keywords

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