Ravi Allada

Ravi Allada (born 1967) is an Indian-American chronobiologist studying the circadian and homeostatic regulation of sleep primarily in the fruit fly Drosophila. He is currently the executive director of the Michigan Neuroscience Institute (MNI), a collective which connects neuroscience investigators across the University of Michigan to probe the mysteries of the brain on a cellular, molecular, and behavioral level. Working with Michael Rosbash, he positionally cloned the Drosophila Clock gene.

Early life Allada was born on August 20, 1967, in Midland, Michigan, to Indian immigrant parents, Sambasiva Rao and Jayalakshmi. Allada has two brothers, Vivek and Gopal, who both currently work as physicians. At the age of 11, Allada won 3rd place in a free throw competition. Allada's interest in sports also led him to track baseball statistics, which triggered Ravi Allada's interest in math and later, his research on jet lag for MLB players. While attending University of Michigan Medical School, Allada spent two years as an HHMI-NIH Research Scholar working with Howard Nash on a molecular genetics project relating to general anesthesia in Drosophila.

The Allada lab focuses on finding molecular components of the circadian clock and their impacts on neurodegenerative diseases, sleep, jet lag, and memory processing.

Early research

Drosophila circadian rhythms

Molecular identification of the Drosophila Clock Gene (1998) Using Drosophila melanogaster as a model organism, Allada and his team used forward genetics to discover a circadian rhythm gene called Drosophila Clock (dClock; Clk). Forward genetics screens for observable phenotypes that could potentially correspond to underlying genetic differences typically resulting from randomly induced mutagenesis. dClock (Clk) was discovered when Allada and his colleagues were completing a forward genetic screen of EMS mutagenized flies. The mutation found by Allada, that abolishes fly circadian rhythms is termed Jrk. While all homozygous flies showed arrhythmic activity in constant darkness. Coupled with complementation data with a null deletion. Data suggests that the Jrk mutation has a negative dominant effect, meaning that only one copy of the gene is sufficient for phenotype interference. Clk and other circadian genes are predominantly expressed in the "lateral" neurons (LN), the pacemaker neurons of the Drosophila central circadian clock.

Allada's research team worked to understand how the expression of certain clock genes influenced the expression of other clock genes and where these genes were expressed. They used the GAL4/UAS system to examine how the expression of certain clock genes, pigment dispersing factor gene (pdf) and long and short versions of the cry promoter DNA sequence, affected neuronal gene expression. Pdf-GAL4 and long cry-GAL4 were only active in known clock neurons, but the shorter cry sequence was also active in "non-circadian" neurons. Additionally, transgenic flies with misexpression of Clk displayed different patterns in locomotor activity to wild type files under light-dark conditions: they displayed a single peak of activity in the daytime, opposed to two peaks of activity in the morning and evening. The receptor was found to be a class II peptide G protein-coupled receptor. TIM and PER proteins form a heterodimer that serves to inhibit the further transcription of clock genes tim and per. Allada studied fruit flies with a mutant CK2 gene, termed CK2 Tik , and observed an abnormally long behavioral rhythm of about 33 hours. The Drosophila gene na codes for an ion channel with homology to the mammalian sodium leak channel, nonselective (NALCN). Mutants of na show poor circadian rhythms, yet oscillations of the clock protein PER remain; pacemaker neurons were found to express na and inducing na in pacemaker neurons is sufficient to restore normal locomotor activity rhythms. This served as an indication that NA likely functions on the clock output and "the mutant is a result of disruption in the coupling between the central clock and the neuronal networks controlling locomotion." Rescue of na in a cluster of posterior DN1 neurons implicates its role in "mediating the acute response to the onset of light" and anticipatory behavior, and pdf expression in DN1 partially rescues morning and free-running rhythms—including in DN1 neurons.

Posterior DN1 pacemaker neurons demonstrate rhythmicity in firing rate throughout the day to facilitate behaviors of sleeping and waking, firing a lot in the morning and scarcely in the evening. DN1 membrane potentials and conductance of sodium and potassium have daily rhythms, indicating that they are potentially under circadian clock control. The researchers refer to this antiphase activity of sodium and potassium currents as a "bicycle" mechanism, and its discovery in nocturnal mice suggests this mechanism is ancient, well-conserved through evolution, and thus likely present in humans. Allada's research team conducted one of the first unbiased neurogenetic screens for neurons identified the Drosophila mushroom bodies (MBs) as a major sleep regulatory center with an effect on wakefulness and sleep duration. The MBs are also well known for their role in learning and memory, connecting sleep regulation to memory consolidation. They also further demonstrated the significance of the GABAA receptor gene, Resistant to dieldrin (Rdl), in promoting sleep. RDL's role in PDF pacemaker neuron inhibition was supported by electrophysiological evidence that GABA induced an inward current of chloride and GABA antagonist picrotoxin blocked this current in lLNv neurons—PDF-secreting, arousal promoting pacemaker neurons. These findings outline one of first proposed wake-promoting circuits in Drosophila, which posits that PDF neuron activation in controlled by the circadian clock to time waking behavior and GABA serves an inhibitor of these neurons to promote sleep.

Rebound sleep (2012-2017) Allada has continued to uncover the molecular basis of sleep homeostasis to further understand why sleep occurs. Focusing on when an organism is sleep deprived leads to what is known as compensatory sleep mechanisms. Sleep deprived organisms engaging in compensatory sleep or sleep rebound is a good indicator of homeostatic sleep regulation. It is known that cul3 and inc are implicated in protein ubiquitination, it is unclear how reduced activity of these genes impact sleep. Allada et al. propose that Inc/Cul3 proteins may "impact dopaminergic modulations of sleep", given that loss of cul3 and inc results in "hyper-arousability to a mechanical stimulus in adult flies" like flies with increased dopaminergic signaling. For example, east coast teams that had travelled west for games recorded decreased game performance after flying back home for home games. In order to receive an assessment of one's biological time, a dim light melatonin onset test is often used. This requires the patient to stay in low light condition, while numerous blood or saliva samples are taken from the patient. However, a new blood test that only requires two blood draws may be able to provide physicians with accurate patient chronotypes. Computational biologist Rosemary Braun, Allada et al, of Northwestern University published a new study in the Proceedings of the National Academy of Sciences USA. The study claimed that their blood draw test can be easily generalized to more patients. The major obstacle in developing a blood test is to find a reliable gene expression biomarker. Due to the diversity of measurement platforms and inherent variability, many biomarkers perform well in original data sets but cannot be universally applied to new samples. Machine learning algorithms can now learn which gene gives the best indication of biological time. With the help of TimeSignature, a computer algorithm that infers circadian time from gene expression, the team was able to yield highly accurate results with a wide population without renormalizing that data.

Neurodegenerative research

Huntington's Disease (2019) Allada and his team have recently been studying a neurodegenerative disease called Huntington's disease and its relationship with circadian rhythm. Using a Drosophila model impaired with Huntington's disease, they found evidence that environmental and genetic perturbations of the circadian clock alter the neurodegeneration caused by Huntington's disease. The results suggested that the knockdown of the clock-regulated protein called Heat Shock Protein 70/90 Organizing Protein (HOP) reduces the mutant Huntington's disease aggregation and toxicity, providing evidence for the casual relationship between circadian clock and neurodegenerative disease.

Sleep waste (2021) Allada has been studying proboscis extension sleep, a deep sleep stage in Drosophila similar to the human deep sleep. The study identified that the prevention of proboscis extensions increased injury related mortality and reduced waste clearance. Allada and his lab team administered luciferin, a substrate for firefly luciferase reporters and discovered evidence of a functional role in Drosophila proboscis extension sleep related to waste clearance. In subsequent experiments, Allada has emphasized the implication of the waste clearance functionality in maintaining brain health and preventing neurodegenerative disease.

Positions and honors

2023 - Executive Director, Michigan Neuroscience Institute (MNI) 2016 - Edward C. Stuntz Distinguished Professorship in Neuroscience 2011 - Brain Research Foundation Seed Grant Award, Circadian Clock and Neurodegeneration 2008 - Distinguished Service Award, Society for Research on Biological Rhythms 2003-05 National Alliance for Research on Schizophrenia and Depression Young Investigator Award 1999-2003 Burroughs Welcome Career Award in the Biomedical Sciences