Ruth Epstein
Ruth Epstein
Faculty Advisor: Wojtek Pawlowski
Year: 2019

Epigenetic Control of Meiotic Recombination in Maize

Meiotic recombination is a major force of genetic variability in the natural world. Recombination creates new combinations of alleles which creates new phenotypes. Control of recombination machinery allows design of favorable combinations of useful genes, such as those controlling drought and pests resistant for more effective crop production. Understanding recombination of crops is becoming quite important in challenges of global warming or increasing global population.
It is known DNA and histone methylation affect the mechanisms of recombination, but exact details of this relationship are not clear. In this project we focus on the mutant, decreased in DNA methylation I (ddm1). DDM1 encodes a helicase Snf2 protein, responsible for the maintenance of DNA and specific histone methylation. Maize has two DDM1 gene copies, however, double mutant is not viable. This situation forces us to investigate a ddm1-1/ddm1-1/ddm1-2/+ heterozygote, which displays a milder mutant phenotype.
To understand how the ddm1 mutations affect the meiotic recombination landscape during meiosis, the Pawlowski lab is performing crossover mapping, cytological, and molecular studies. My part of this project is characterizing localization and dynamics of meiotic and chromatin proteins in the ddm1 mutant in comparison to wild-type plants. For that, I tested a novel meiotic spread slide preparation method and performed a number of immunolocalization experiments at different meiotic stages. In the experiments, I used RAD51 protein as a marker recombination initiation, HEI10 and MLH3 as crossover markers, and H3K4me3 and H3K9me2 as a chromatin state markers.
I evaluated a novel meiotic spread slide preparation. This new method tracks the dynamics of localization of investigated antibodies. Using immunolocalization, I found that the number of HEI10 foci in ddm1 pachytene cells is lower in comparison to the B73 WT; MLH3 foci number is opposite. MLH3 foci is elevated in ddm1 and lower in WT pachytene cells. From immunolocalization images and statistics, I found that there is an increase in co-localization of MLH3 and HEI10 through prophase I, which was not observed before. Finally, H3K9me2 foci points were elevated in the ddm1 mutant in comparison to the B73 WT indicating different chromatin organization in ddm1 mutant.

 

My Experience

Before coming to BTI, I knew I was interested in the plant sciences. My undergraduate university does not have a large plant biology department so the BTI internship was extremely helpful in exploring the plant science field. Weekly seminars and networking within the plant community was crucial to my education and made me feel prepared as a researcher. From these events, I have become interested in plant breeding and its impact in global health.
Something else I enjoyed was being surrounded by like-minded undergraduates who were just as passionate as I am about plant science while also wanting to pursue a higher degree. Most of all, I loved my time in the Pawlowski lab. My mentor, Mateusz, was paramount in my understanding of the lab’s research and my own project. I feel I have improved my laboratory skills, learned new techniques, and most of all realized the importance in asking for help. In the future I want to pursue a Ph.D in either plant biology or plant breeding. Without this internship I would have not realized my potential or my true passions in the plant sciences. I am thankful for the wonderful people I have meet and will always remember my summer in Ithaca.