The Libuda Lab investigates how developing sperm and eggs repair DNA breaks with the proper chromosome template to ensure faithful genome inheritance from one generation to another.
Double strand DNA breaks (DSBs), potentially catastrophic events, occur during both mitosis and meiosis and must be repaired to maintain genomic stability. The inaccurate repair of DSBs contributes to the development and progression of cancer. During meiosis, DSBs are intentionally induced, and their formation and repair occur under precise regulation both to restore genome integrity prior to cell division and to promote proper chromosome segregation. Meiotic errors in chromosome segregation contribute to miscarriages, stillbirths, and birth defects. While repair of DSBs with the appropriate chromosome template (homolog) is necessary for genomic integrity, very little is known as to how germ cells achieve this repair template preference in the presence of other potential templates with nearly identical sequences (sister chromatids; Figure 1).
To understand how chromosomes are able to access distinct repair templates and pathways depending on chromosomal and cellular context, the Libuda Lab combines genetic, cytological, molecular, and biochemical methods in the Caenorhabditis elegans model system. The C. elegans system is advantageous for investigating how DSB repair decisions are imposed and regulated during meiosis, in part because the events of meiotic prophase are organized in a spatial-temporal gradient along the distal-proximal axis of the gonad, enabling visualization of all the stages of meiosis in a single gonad (Figure 2). Utilizing high-resolution and super-resolution microscopy, the germ line allows imaging of DNA repair events along chromosome structures of intact germ cells where the three-dimensional nuclear architecture is preserved (Figure 3).
Some of our current interests include:
Temporal features of DNA repair template choice
We have developed a set of assays that: 1) induce formation of a DSB at a defined site; and, 2) detect and distinguish DSB repair outcomes with either the homolog and sister chromatid template at specific stages of meiotic prophase. Utilizing these assays, we are testing for a hypothesized switch in partner choice during meiotic prophase progression. Further, we are identifying chromosome-structural features and bound proteins that influence partner choice.
Molecular mechanisms of DNA repair template choice
Utilizing a variety of genetic and cytological assays, we are investigating how specific chromosome structures, checkpoint kinases, and repair pathways affect the partner choice decision. In addition, we are performing genetic screens to identify additional factors involved in the DNA repair partner choice decision.
Dynamics of DSB repair
Utilizing live imaging of ongoing DSB repair events in whole, intact worms, we are revealing how duration and dynamic localization of early DSB repair stages influence and ensure DSBs are repaired in the appropriate context (Figure 5).
Male germline-specific responses to temperature
Our lab has identified a temperature condition that induces a phenotype only found in male germlines. We are investigating this phenotype to understand how chromosomes in male germlines differ in their response to changes in temperature.
(pulled from pubmed)
(pulled from pubmed)