Dinoflagellate Genetics

Dinoflagellates have some of the largest genomes of any other organisms. Most eukaryotic algae carry approximately 0.5 pg/cell of DNA whereas dinoflagellates have been shown to carry anywhere from 3-250 pg/cell of DNA. For reference, the human genome is 3180 Mb long while dinoflagellate genomes can range from 3000-250,000 Mb depending on the species. Their genomes are also permanently condensed making dinoflagellate DNA notoriously difficult to sequence and making the organisms impossible candidates for studies involving traditional genetic techniques like CRISPR. Our lab is currently working to reduce these obstacles so that we can gain a better understanding of dinoflagellates from the genetic perspective.  

Alternative Bases

The DNA of most organisms is made up of four nucleotide bases— guanine, adenine, cytosine, and thymine. Dinoflagellates, however, have a fifth nucleotide base: 5-hydroxymethyl uracil (5hmU). This base replaces approximately 40% of thymines in the genome though this depends on the species. Additionally, 5hmU is not uniformly distributed across the genome. It is found in both the ribose and deoxyribose nucleotide pools, but is only found in DNA. Researchers have speculated what purpose 5hmU serves and have proposed that it serves some form of gene regulation.

Our lab is currently using Nanopore technology to develop methods for sequencing alternative bases like 5hmU. We have also tried combine bioinformatic and molecular data to explore whether a thymine dioxygenase enzyme might be involved in this nucleotides' synthesis. 

Gene Regulation in Dinoflagellates

Gene and transcriptional regulation is currently enigmatic in dinoflagellates with mRNA levels showing no correlation with protein production. Because dinoflagellates are believed to regulate at the translational level, rather than during transcription, translation factors are of great interest when understanding dinoflagellate metabolism. In most known eukaryotic translation systems, eIF4Es function as a rate-limiting step toward protein synthesis and our lab is exploring whether these enzymes have similar functions in dinoflagellates. Determining the role of the eIF4E family members requires a method of knocking down their expression. The unusual cell biology of dinoflagellates makes common gene knockout strategies impractical, limiting the amount of genetic research that can be applied. 

In particular, we are pursuing the use of an antisense-based knockdown approach in order to study how a decrease in target gene expression effects dinoflagellate metabolism. Our lab has also been researching dinoflagellate growth phases across a diel cycle in the hopes that with cell synchronization we can improve the antisense-based knockdown approach.