Initiation of Silencing

The Central Question

A fundamental question in biology is how cells distinguish self from non-self genetic elements. How do sophisticated silencing pathways selectively target non-self elements like viruses and transgenes while sparing endogenous genes? Complicating this further are transposable elements (TEs)- intrinsic genomic parasites that must be silenced despite being part of the organism's own DNA, as their mobilization can disrupt genes and genome function.

I like to think of the genome as embodying a Yin-Yang philosophy, both the 'light' (genes) and 'dark' (transposons) elements are essential to life. This perspective highlights how transposons, despite needing to be silenced, play crucial roles in positively regulating genome integrity and generating variation that enhances organismal fitness.

DNA Methylation and the RdDM Pathway

My research focuses on identifying these elusive "primary small RNAs" that initiate the RNAi feed-forward cycle, establishing DNA methylation and long-term chromatin modifications that lead to stable epigenetic silencing.

Why Arabidopsis?

I use Arabidopsis thaliana because it's exceptionally suited for genetic manipulation and has vast resources of mutants and genomic information. Since my work relies heavily on genetic approaches, Arabidopsis can survive more severe genetic alterations compared to other models. The findings also translate well to crops, helping tackle transgene silencing that causes significant economic losses to farmers and industry. Plus, plants are a fantastic system to study transposons, they were first discovered in corn by Barbara McClintock in the 1940s!

Genome Engineering and Technology Development

CRISPR has become my go-to tool for generating mutants in Arabidopsis, whether single mutants or complex combinations. During the early years of my PhD, I was lucky to be part of the team developing transposase-assisted target-site integration, which ended up being published in Nature (pretty cool right?!).

These days, I'm constantly experimenting with different transformation techniques like protoplast transformation, transient transformation of Arabidopsis seedlings, basically whatever works best to develop high-throughput screening tools for genome editing outcomes.

During my graduate school rotations, I also got to play around with targeted methylation using CRISPR/SunTag systems for epigenome engineering, with the goal of engineering viral disease resistance in plants.

Mechanism of Genome Dosage Sensitivity and Regulation (Undergraduate research)

Genome dosage sensitivity describes how changes in the ratio of maternal to paternal genetic contributions trigger developmental responses, with imbalances causing dramatic effects on growth and development. The endosperm, a triploid tissue in flowering plants containing two maternal and one paternal genome copies, is exceptionally sensitive to dosage changes, making it an ideal model system to study how organisms sense and respond to genomic imbalances.

During my time as a Khorana Scholar at MIT with Dr. Mary Gehring, I investigated a fascinating question: how do plants sense and respond to genomic imbalances? The Gehring lab had discovered that mutations in RNA Polymerase IV, an enzyme that produces small RNAs guiding DNA methylation, can rescue seed lethality caused by paternal genome excess. This finding suggested that RNA Pol IV plays a critical role in sensing genome dosage.

This discovery raised intriguing questions about the mechanisms underlying genome dosage sensitivity in plants. To explore this, I generated extensive interploidy crosses in Arabidopsis thaliana, systematically testing mutants for various methylation readers and writers. By analyzing patterns of seed viability across different genetic backgrounds, I aimed to identify which genes and epigenetic pathways are essential for proper dosage sensing in the endosperm.

This work highlighted the intricate connections between small RNA pathways, DNA methylation, and reproductive success in plants, demonstrating how epigenetic mechanisms can regulate genome dosage.

Annotation of Domain of Unknown Function Proteins (DUFs) (Undergraduate research)

During my undergraduate and master's research, I investigated the role of DUF1645 proteins in multiple stress tolerance and grain yield in rice. Our work revealed DUF1645 as a novel intrinsically disordered transcription factor family specific to plants, contributing to our understanding of plant stress responses at the molecular level.

I used various computational tools and publicly available transcriptome data extensively to annotate the DUF1645 protein family. I was fortunate to work with multiple protein structure prediction tools prior to AlphaFold during my undergraduate research, and then validate my computational findings through wet-lab experiments during my master's thesis work.

Intrinsically disordered proteins continue to capture my attention and interest, and I hope to return to studying them in the future!