DNA double-strand breaks: a complicated legacy

“Well, you know, in a way I wish I hadn't met you two. It's much more convenient to think of the opposition as a nice homogeneous, dead-wrong mass. Now I've got to muddy my thinking with exceptions.” -Kurt Vonnegut

“Among the most dangerous damage that can occur is DNA [double-strand] breaks. Like tiny scissors, radiation, UV light, and certain drugs, as well as unavoidable errors during cell division, can snip the DNA strand in two, causing mutations that may lead to cancer or cell death.” So I wrote in this column just a few months ago while describing the work of Dr. Gerry Smith, professor in the Basic Sciences Division at Fred Hutch, to understand the cellular mechanisms responsible for patching broken DNA back together. And those words remain true. Except when they don’t. Here’s another quote, from Dr. Smith himself: “In the absence of crossovers [initiated by double-strand breaks]…[most species] often produce gametes with an improper (aneuploid) set of chromosomes that give rise to inviable or disabled progeny.” They’re good. They’re bad. You need them. They’ll kill you. Double-strand breaks are, to use a favored word in the scientist’s lexicon, complicated. In this month’s article, I highlight new work from Dr. Smith’s lab, led by former student Mélody Wintrebert and technician Mai-Chi Nguyen and recently published in the Journal of Cell Science, on the beneficial aspects of double-strand breaks.

Reaping the benefits of double-strand breaks, and avoiding their dangers, is a matter of limiting them to the right time and place. One such time and place is during meiosis, which produces gametes such as sperm and eggs or, in the case of Dr. Smith’s organism of choice, the yeast Schizosaccharomyces pombe, spores. During meiosis, pairs of homologous chromosomes are physically broken up and recombined, a process that is important both to ensure that meiosis proceeds correctly and produces healthy gametes and to promote evolution, as mixing and matching chromosomal DNA generates new mutations and new gene combinations upon which natural selection can act. “During meiosis, recombination in the several species examined is initiated by the formation of DNA double-strand breaks (DSBs)”, write the authors. Interestingly, “DSBs do not occur at random across the genome; rather, they occur at higher than genome-average frequency at special sites, called DSB hotspots.” This process requires a group of 4 proteins Rec25, Rec27, Mug20, and Rec10, collectively called Linear Element (LinE) proteins, which interact to form a complex and collectively activate  the double strand break machinery for recombination. But questions regarding the function of these proteins remain.

In the current work, Dr. Gerry Smith’s group examined how these proteins make their way from the cytoplasm, in which they are made, into the nucleus in order to encounter DNA and perform their meiotic role. “Only Rec10 appears to have a special amino acid sequence, the nuclear localization signal (or NLS), which directs proteins to the nucleus,” said Dr. Smith. To test the hypothesis that the Rec10’s NLS is responsible for nuclear import, the group mutated this portion of the protein and examined effects on the localization of the LinE components. They observed that all 4 of the LinE proteins no longer could get into the nucleus at normal levels. “Instead, they formed lumpy aggregates in the cytoplasm,” Dr. Smith described. They also observed a reduction in the amount of meiotic recombination, which requires double-strand breaks, in mutant cells, leading to a loss of spore viability. The authors concluded that the four LinE proteins bind together in the cytoplasm, and then collectively rely on Rec10’s NLS to get them into the nucleus. Dr. Smith describes this process as analogous to a group of co-workers commuting to work. The LinE proteins, he notes, represent a case of molecular carpooling, with Rec10 being responsible for picking up its co-workers and driving them to work on the back of its NLS. He notes, though, that if Rec10 calls in sick the rest of the group may not be completely without other options. “Without the NLS, the complex gets into the nucleus at only low level, but it still seems to work better at hotspots than elsewhere. So, we think there is some NLS-independent way to get in occasionally, but the few LinEs that get in do their work properly.”

Although this work was done in yeast cells, Dr. Smith is excited about the prospect of the findings informing us about how human double-strand breaks are regulated. “Our research here likely pertains to other species, including humans, because LinEs have structural and functional similarities to the meiotic synaptonemal complex (SC) found in nearly all species. But an NLS has not, to our knowledge, been demonstrated for the SC proteins of any other species. There are excellent candidate NLSs in some species we discuss, and we await someone testing these NLSs, perhaps using methods like the ones we used.”

This work was supported by the National Institutes of Health and philanthropic contributions to the Fred Hutchinson Cancer Research Center.

Fred Hutch/UW Cancer Consortium member Gerald Smith contributed to this work.

Wintrebert M, Nguyen M-C, Smith GR. (2021) Activation of meiotic recombination by nuclear import of the DNA hotspot-determining complex in fission yeast. Journal of Cell Science 134 doi: 10.1242/jcs.253518