Our laboratory is interested in identifying and understanding the dynamics of molecules that direct the formation of the brain, and how alterations in these pathways lead to neurodevelopmental and behavioral disorders.  A major effort in the lab is to leverage missense variants, which typically change a single amino acid in a protein of affected individuals.  The advantage of this approach is that missense variants are typically present in the cell (not a simple null allele), and cause diseases by changing a specific activity of a protein. We theorize that understanding how such variants affect protein function will provide deeper insights into the mechanisms of brain development, and ultimately, provide therapeutic strategies for disease treatment. Our approach is highly interdisciplinary and combines both genetic and biochemical methods in cell culture and mouse models.

understanding the mechanisms of angelman syndrome

A major focus of the lab centers on Angelman syndrome, a severe a severe form of intellectual disability characterized by epilepsy, motor deficits, microcephaly, dysmorphic facial features, sleep deficits, and a unique happy demeanor. It is caused by mutation or deletion of the maternally inherited UBE3A gene. The UBE3A gene encodes the UBE3A protein, which is a HECT (Homologous to E6AP C-terminus) domain E3 ubiquitin ligase that regulates the degradation of unnecessary proteins in the cell.  The expression pattern of UBE3A is intriguing because it is expressed biallelically in most tissues but is only expressed from the maternally-inherited gene in neurons.  In previous work, our laboratory discovered that patient-identified missense variants in UBE3A can cause both a loss and gain-of-function of the enzyme with some variants increasing UBE3A activity by over 800% above wild-type enzyme levels (Weston et al., Nature Communications, 2021).  There are several critical questions that we are currently pursuing.

1) What are the neurodevelopmental consequences of UBE3A gain-of-function?

Our initial studies indicated that individuals with UBE3A gain-of-function mutations commonly possess intellectual disability, behavioral disorders, and developmental delays. Importantly, the clinical features of these individuals suggest they possess phenotypes that are distinguishable from classic Angelman syndrome. To better model these disorders, we created several lines of mice possessing precise gain-of-function mutations in UBE3A.  In previous work, we showed that a single gain-of-function mutation in UBE3A is sufficient to cause motor, coordination, and early life communication deficits in mice (Weston et al., Nature Communications, 2021). How are brain regions and structures altered by UBE3A gain-of-function? In other words, how is brain development perturbed to cause these phenotypes? We are currently performing biochemical, anatomical, and behavioral assessments to understand how brain development is altered by excessive UBE3A ubiquitin ligase activity.

2) What are the mechanisms by which disease-causing mutations in UBE3A cause loss and gain-of-function of the enzyme?

A mystery regarding both loss and gain-of-function UBE3A variants is that many are found outside the known catalytic site of UBE3A, suggesting there are other important domains that regulate enzyme activity.  We leveraged the deep structure-function data provided by our functional variant screening and found that one of these domains comprises a ubiquitin-binding domain that is required for UBE3A to form polyubiquitin chains (Weston et al., Nature Communications, 2021).  We are continuing this work using both protein modeling and biochemical studies to identify additional domains that regulate UBE3A function.

3) Can we utilize gain-of-function mutations to understand the pathways UBE3A regulates in the developing brain?

The disease-relevant substrates of UBE3A in the developing brain remain unknown.  However, our discovery of both loss and gain-of-function variants in UBE3A provides us genetic tools to both increase and decrease UBE3A activity in the cell.  Using these insights, we are currently performing proteomics-directed studies to identify the substrates of UBE3A and map the molecular pathways that are critical for brain development.

Additional reading:

Yi lab develops assay to solve mystery genetic variants

Gaining mechanistic insights into the neuronal transcriptome

A second focus of the lab is to understand the mechanisms that govern maternal-specific expression of UBE3A in neurons.  The neuron-specific expression of UBE3A requires both epigenetic imprinting and curiously, the expression of long genes.  Thus, broadly speaking, the imprinting of UBE3A is interlinked to a unique feature of neuronal transcription programs, which is that neurons transcribe the longest genes in the genome.  To put it another way, the longest genes in the genome are neuron-specific genes.  How and why does this occur?  We are again leveraging missense variants in candidate transcriptional regulatory proteins to dissect how this phenomenon occurs.  This project utilizes both genetic and genomic manipulations, bioinformatics, and animal modeling to understand the mechanisms and neurobiological consequences of transcriptional dysfunction in developing neurons.  

Engineering novel molecular tools for the study of neuronal development

Plants have evolved numerous biochemical pathways to respond to and utilize light.  One of these light-sensitive protein domains is the Light-Oxygen-Voltage (LOV) domain.  The LOV domain is composed of a globular fold that binds a light-responsive flavin mononucleotide, and a long, C-terminal alpha helix that unwinds when this domain is exposed to light.  This structural dark/light cycle is reversible, and in previous work, we leveraged this conformational change to cage kinase inhibitory peptides into the LOV domain (Yi et al., ACS Synthetic Biology, 2014).  This provided a method to expose the inhibitory peptide only in the presence of light, thereby providing a highly precise, non-invasive method of perturbing signaling in living cells.  We are continuing our study of this fascinating domain, and looking for new ways to utilize its unique properties to engineer new light-controllable tools.   

Work in our lab is generously supported by: