Current Students

Supervisor: Stephen Wright, Ecology and Evolutionary Biology

Supervisor: Leslie Buck, Department of Cell and Systems Biology

Supervisor: Belinda Chang, Department of Cell and Systems Biology

The infraorder Cetacea (whales and dolphins) has emerged from perhaps the most dramatic evolutionary transformation that has occurred within mammals. While modern cetaceans are obligately aquatic, foraging by breath-hold diving and intermittently resurfacing to breathe, they are in fact descended from terrestrial artiodactyls; their closest extant relatives are the land-dwelling hippopotamids. Following their emergence about 54 million years ago, cetaceans underwent major morphological and sensory changes as they transitioned from a terrestrial to an aquatic habitat. The visual system in particular underwent a number of changes, likely driven in large part by the need to forage via diving. As light availability underwater decreases with increasing depth, the cetacean eye has become highly adapted to dim-light conditions, with features such as a reflective tapetum lucidum and a rod-dominated retina. As well, since the spectrum of visible light deep underwater is blue-shifted relative to the surface, the absorption spectra of some cetacean visual pigments have shifted accordingly, while others have become non-functional. With the sequencing of many cetacean genomes over the past decade, the specific molecular mechanisms underlying the adaptation of cetacean visual systems to their aquatic environments can now be investigated. First, computational analyses can be conducted to detect signatures of positive selection acting on specific codon sites within protein-coding genes. Then, experimental analyses can be conducted to determine how the substitutions observed at positively selected sites affect the function of the encoded protein. Several studies have begun to apply these methods to cetacean visual proteins, and have identified key substitutions that may have contributed greatly to visual adaptation. Much of this research has been focused on rhodopsin, the light-sensitive pigment that initiates the phototransduction cascade in the rod cells of the retina. While these studies have provided great insight into the molecular evolution of vision in cetaceans, much more remains unknown. Hundreds of mammalian proteins are known to be involved in phototransduction and the visual cycle, and hundreds more are known to be required for eye development. Amino acid changes in any number of these proteins could have contributed to the adaptation of cetacean visual systems to their aquatic environments, but large-scale computational analyses of selection have not yet been performed on the genes encoding these proteins in cetaceans. Such analyses would provide insight into potential positively selected sites within these proteins, and provide a more complete picture of the molecular basis of this adaptation.

This project will use various computational methods to detect positive selection on a large number of cetacean visual genes, including those involved in rod and cone phototransduction, the visual cycle, photoreceptor cell development, eye structure, and other vision-related functions. Sets of visual genes will first be established using current literature, and mammalian eye-derived gene expression datasets. Genome assemblies for dozens of cetacean species will be obtained from genomic databases, including NCBI and the Zoonomia Project. The protein-coding sequences of the genes of interest will then be extracted from these genomes using existing gene annotation, together with an automated computational pipeline that has recently been developed in the Chang laboratory. The resulting coding sequences will be aligned using a multiple sequence alignment algorithm such as MUSCLE or PRANK. Using these alignments together with phylogenies of the relevant species, patterns of selection will be inferred by estimating dN/dS (denoted ω), the ratio of the nonsynonymous and synonymous substitution rates; ω<1 is indicative of purifying selection, ω=1 indicates neutral evolution, and ω>1 is a signature of positive selection. Codon-based models of sequence evolution, such as those available in the PAML and HyPhy programs, will be fitted to the sequence data and compared against their respective null models to statistically test for positive selection. Models will be implemented that allow ω to vary between codon sites; this will enable the detection of specific amino acid residues that have been the targets of positive selection. Models that additionally allow ω to vary between specified groups of branches of the phylogeny will be implemented to test for differences in selection pressure between cetacean lineages. When implementing these models, the phylogeny will be partitioned based on factors such as habitat, diet, feeding mode, and foraging depth, in order to assess the effects of these ecological factors on visual protein evolution in cetaceans. This project will identify visual proteins that have been the targets of positive selection in cetaceans, and will determine the functional effects of amino acid substitutions at positively selected sites within them. The results will provide great insight into the molecular mechanisms behind the adaptation of cetacean visual systems to the aquatic environment; they will also reveal much about the functions of visual proteins, and the functional importance of certain sites within these proteins, in cetaceans and beyond.

Supervisor: Yan Wang, Ecology and Evolutionary Biology

Supervisory: John Calarco, Department of Cell and Systems Biology

Alternative splicing plays a central role in pre-mRNA maturation, influencing gene expression and generating proteomic diversity. Distinct patterns of alternative splicing, largely governed by RNA-binding proteins (RBPs), give rise to unique transcriptomic identities. These RBPs tend to bind functionally related pre-mRNAs, forming dynamic post-transcriptional operons. Notably, neuronal tissue is characterized by an abundance of alternative isoforms and a unique profile of splicing factor expression, which serves as a model to study the regulation of alternative splicing networks by RBPs. One aspect of alternative splicing regulation that remains poorly understood is its connection to intracellular signalling, particularly in neuronal tissue. To address this gap, I aim to conduct a reverse genetics screen within C. elegans neuronal tissue to identify alternative splicing events responsive to intracellular signals. In parallel, I am analyzing available RNA-seq datasets to uncover neuronal alternative splicing influenced by the daf-2 insulin-like signalling pathway. To identify the splicing factors affected by these signalling pathways, I am adapting an RNA-tethered proximity-labelling method coupled with mass spectrometry. This approach will characterize the proteins associated with transcripts undergoing signal-regulated alternative splicing. Neuronal transcriptome profiling in conjunction with transcriptome-wide identification of the targets of signal-regulated splicing factors will then be used to reveal how regulation of the RBPs affects their post-transcriptional operons. This project will offer insights into the mechanisms governing the regulation of alternative splicing networks by intracellular signalling pathways and shed light on the complex interplay between distinct regulatory mechanisms that determine neuronal function and identity.