Plants In 1985, Kay and his colleagues found that the
Cab gene was under circadian control in wheat and transgenic tobacco plants during his postdoctoral research. In 1991, Kay extended this research into a suitable model plant,
Arabidopsis thaliana and found that Cab mRNA levels are also under circadian control in
Arabidopsis. He then developed Cab2:luc fusion, the fusion of luciferase
open reading frame downstream of the Cab2 promoter region, as a marker for monitoring the circadian phenotype. This fusion marker was widely used in later studies and contributed enormously to the understanding of
circadian rhythm regulation in
Arabidopsis. Based on this Cab:luc fusion technology, Kay set up
luciferase imaging assays for large scale
forward genetics screening and identified the first short period mutant of
TOC1 gene.
TOC1 was proved to be a core clock gene in
Arabidopsis and was cloned by Kay lab after a long period of time Kay also revealed the biochemical function of
TOC1 and found that
TOC1 and LHY/CCA1 reciprocally regulate each other, and further studied the mechanism of this regulation. Kay identified ELF3, GI, Lux, CHE and PRRs as core clock genes and studied their role in the circadian regulation loop. He also profiled
clock controlled genes (ccg) in
Arabidopsis with several technologies and identified key pathways temporally controlled by
circadian clock. His work on functional analyses of core clock genes, as well as ccg, successfully connected
circadian rhythm with the control of development, like seedling, growth and flowering. His work on these clock genes contributed significantly to the understanding of repression-based clock regulation loops in plants, which is distinct to the ones in animals that are composed of both positive and negative elements. Kay discovered the mechanism of seasonal time and day-length measurement and flowering time determination in
Arabidopsis through the
GI/FKF1-CO-FT pathway. Kay found evidence that there are multiple phototransduction pathways, and contributed to the discovery and functional analysis of many
photoreceptors, including
phytochrome,
cryptochrome,
ZTL and
LKP2 and their roles in
circadian rhythms.
Flies Kay applied the first clock gene fusion, Per:luc, in
Drosophila melanogaster which allows monitoring of its rhythm at the single animal level. Per:luc fusion also helped him understand the phase relationship in
mRNA and protein oscillation. He further improved the mathematical method of
bioluminescence analysis and made the results quantified. In 1997, his
Per promoter driven
Green Fluorescent Protein (GFP) study suggested that
Per is widely expressed throughout the fly body in a rhythmic pattern, and all body parts are capable of light perception. This is one of the first pieces of evidence for a peripheral self-sustaining circadian clock. In 1998, he proposed the translational transcriptional feedback loop model of the
circadian clock in flies, analogous to other labs that proposed a same model in mammals and fungi. Kay discovered that
cryptochrome is the circadian
photoreceptor that directly acts with and sequesters
TIM in response to light. Kay did one of the pioneering microarray analyses to study
clock controlled genes (ccg), and revealed tissue-specific nature of
circadian rhythms by analyzing the ccg of heads and bodies separately.
Mice Kay began his extensive research on mice in 1999 at the
Genomics Institute of the Novartis Research Foundation, with a primary focus on
melanopsin (Opn4) and visual photoreceptors. It was here, with the use of automation and large-scale genomics technology, that Kay and collaborating colleagues found that the mammalian clock consisted of more than just one feedback loop. In 2002, Kay and his team were able to show the role of
melanopsin, a photosensitive photopigment in retinal ganglion cells, in detecting light for the master circadian oscillator located in the
suprachiasmatic nucleus (SCN) in the hypothalamus of the brain. Both
melanopsin and visual
photoreceptors, such as rods and cones, were required for
entrainment. However, removing each individually did not result in total blindness in mice, as they retained non-visual photoreception. The enzyme
luciferase was utilized by Kay's lab to research
clock gene expression in single culture cells and revealed that a variety of cells, including those of the liver and fibroblasts, demonstrate
circadian rhythm. As time went on, these rhythms became increasingly out of phase as local oscillators desynchronized and each cell expressed their own pace. In 2007, these findings demonstrated the need to examine single-cell phenotypes along with behaviors of experimental clock mutants. In 2009, inspired by his mother's fatal motor neuron disease, Kay and some colleagues performed a study manipulating the
ubiquitin ligase protein
Listerin in mice which led to the conclusion that mutations in Listerin caused neurodegeneration.
Humans Kay's research on intercellular networks has the potential to contribute to drug therapies by identifying compounds that affect the circadian pathways. His findings and analyses of this mammalian oscillator contribute to our medical understanding of how the clock controls downstream processes and holds clinical significance as a variety of diseases and biological processes are involved, such as aging, immune response, and metabolism. For instance,
diabetes and the circadian clock may correlate based on the findings of circadian expression in the liver and glucose output. Using a cell-cased circadian phenotypic screen, Kay and a team of
chronobiologist researchers identified a small molecule,
KL001, that interacts with
cryptochrome to prevent
ubiquitin-dependent degradation, which results in a longer circadian period.
KL001-mediated
cryptochrome stabilization (of both CRY1 and CRY2) was found to restrain
glucagon-activated
gluconeogenesis. These findings bear the potential to aid in the development of circadian-based diabetic therapeutics. Circadian clocks have also been shown to influence cancer treatments, where circadian disruption accelerates processes and drug responses are affected by the time of administration with respect to the circadian cycle. ==Positions and honors==