Research

Genome instability is a defining feature of cancer, yet many of the most dramatic genome rearrangements arise not gradually, but in sudden, catastrophic events. In chromothripsis, one or a few chromosomes are shattered into many fragments following errors in cell division, rapidly reshaping the cancer genome and promoting oncogene amplification, tumor evolution, and therapy resistance.

The Krupina Lab investigates the molecular mechanisms that initiate chromosome shattering, with a particular focus on cytoplasmic DNA nucleases that gain access to chromosomes after nuclear envelope rupture. Our work aims to define how “all-at-once” genome damage occurs, how it is regulated, and how it contributes to tumorigenesis.

Research Directions

• Molecular mechanisms of chromosome shattering by N4BP2
  Our work identified N4BP2, a cytoplasmic endonuclease, as a key trigger of chromosome shattering when chromosomes become exposed in ruptured micronuclei (Fig). We are now defining how this enzyme damages DNA at a mechanistic level. We combine biochemistry, genome-wide DNA break mapping, and super-resolution live-cell imaging to define the spatiotemporal dynamics of N4BP2-mediated chromosome fragmentation.
• Regulation of cytoplasmic nucleases during genome instability
  N4BP2 is a large, poorly characterized protein whose activity must be tightly regulated to prevent inappropriate genome damage. We study how nuclease activity is controlled in space and time, focusing on protein–protein interactions, regulatory complexes, localization.
  
  This work addresses how cells normally protect their genomes — and how that protection fails in cancer.
• Nuclear envelope rupture as a gateway to genome damage
  Chromosome shattering requires a breach of the nuclear–cytoplasmic barrier. We investigate how nuclear envelope rupture during micronucleation and chromatin bridge formation exposes chromosomes to cytoplasmic enzymes.
  
  Our goals are to understand why only certain micronuclei undergo catastrophic damage, whether DNA damage is spatially restricted to rupture sites, and how replication stress and nuclear architecture influence chromosome vulnerability. This direction links mitotic errors, nuclear integrity, and genome instability.
• Consequences for tumor growth and therapeutic targeting
  Chromosome shattering promotes aggressive tumor behavior by generating genome instability and oncogene amplification, including extrachromosomal DNA. We study how nuclease-driven genome damage impacts tumor growth and aggressiveness, intratumoral heterogeneity, patient outcomes across cancer types.
  
  Long-term, we aim to target genome instability at its source, by inhibiting nucleases or regulatory pathways that enable chromosome shattering.

Approaches

We combine:

• high-content and live-cell imaging
• genome engineering and functional genetic screens
• genome-wide DNA break mapping and cancer genomics
• in vitro and in vivo cancer models

This integrative strategy allows us to connect molecular mechanisms to genome-wide outcomes in cancer.