Header

UZH-Logo

Maintenance Infos

Role of the RNA-binding Protein NONO in Regulating Circadian Gene Expression and Metabolism


Benegiamo, Giorgia. Role of the RNA-binding Protein NONO in Regulating Circadian Gene Expression and Metabolism. 2017, University of Zurich, Faculty of Science.

Abstract

Circadian rhythms represent a fundamental property of living organisms. Even when they are held in temporal isolation, most organisms from bacteria to humans exhibit behavioral and physiological rhythms with a period of approximately 24 h. These rhythms are driven by internal biological clocks. In mammals, the molecular clock mechanism is driven by transcriptionaltranslational feedback loops of activator and repressor proteins. The transcriptional activators Circadian Locomotor Output Cycles Kaput (CLOCK) and Brain and Muscle ARNT-Like Protein 1 (BMAL1) heterodimerize and activate the transcription of the repressors Periods (Per1, Per2 and Per3) and Cryptochromes (Cry1 and Cry2) that in turn repress the activity of the CLOCK:BMAL1 complex. These cycles of activation and repression take about 24 h to complete and they generate oscillations in several clock-controlled genes (CCGs) with periods of ~24 h. It has been estimated that in any mammalian tissue, about 5-20% of the expressed genes exhibit circadian oscillations in their messenger RNA (mRNA) levels. Rhythms in mRNA abundance are the result of the interplay between the autonomous cellular clock and external time cues (i.e. the light/dark cycle and the fasting/feeding cycle). It has been demonstrated that the fasting/feeding cycle is the dominant Zeitgeber (“time giver”) that drives rhythms of mRNA abundance in peripheral tissues. It has been estimated that most cycling mRNAs do not have a corresponding rhythm in their nascent RNA, this suggested that post-transcriptional mechanism play an important role in generating mRNA and protein oscillation. However, little is known about the molecular mechanisms involved in generating mRNA rhythms at post-transcriptional level. The nuclear RNA binding protein Non-POU Domain Containing Octamer Binding (NONO) is a multifunctional protein that has been involved in many different steps of the mRNA life cycle from transcription, to processing, RNA stability and export. Furthermore, NONO is one of the main components of a subset of nuclear speckles called paraspeckles, whose function in the cell remains unclear. Importantly, NONO was found to interact with PER proteins and affect circadian ! ! Summary ! ! ! 2 rhythmicity in flies and mammals, however the specific mechanisms of its function in circadian RNA expression are unknown. In my thesis I have investigated the role of NONO in mammalian circadian gene expression and physiology. We found that feeding induces an increase in the number of NONO-containing, speckle-like structures in the nuclei of liver cells. In order to investigate NONO function in the fasting-feeding transition, we performed immunoprecipitation of NONO complexes in the liver nucleus and characterized bound RNAs and proteins. We found that NONO interacts with several RNA processing factors and that it primarily binds promoter-proximal introns of transcripts. This led us to hypothesize that NONO might be involved in the processing of its target RNAs. Furthermore, we found that the number of NONO-bound RNAs increases upon feeding and that about 40% of NONO target genes display circadian rhythmicity in their mRNA expression. We then showed that NONO regulates the rhythmicity and phase of these genes post-transcriptionally. To further explore the role of NONO in liver physiology we characterized the function of its bound RNAs. We found that a large fraction of circadian NONO-bound RNAs encodes proteins implicated in glucose uptake and macronutrient metabolism. The absence of NONO-mediated regulation of target RNAs profoundly impacts metabolic health. Indeed, NONO-deficient mice exhibit impaired glucose tolerance, which can be rescued by re-expression of NONO specifically in the liver. NONOdeficient mice also show reduced capacity to store glycogen and lipids in the liver. We also found that NONO-deficient mice burn more fat during the fasting (resting) phase and maintain a lean phenotype throughout their life. We therefore propose that NONO coordinates pre-mRNA processing of metabolic genes with the fasting/feeding cycle, thus allowing efficient energy utilization and storage. Disruption of the temporal coordination between metabolic demand and gene expression leads to the development of metabolic diseases, like obesity and diabetes. Our findings help to better understand how metabolic homeostasis is maintained in mammals, and identify novel therapeutic targets for treating diabetes and other associated metabolic dysfunctions.

Abstract

Circadian rhythms represent a fundamental property of living organisms. Even when they are held in temporal isolation, most organisms from bacteria to humans exhibit behavioral and physiological rhythms with a period of approximately 24 h. These rhythms are driven by internal biological clocks. In mammals, the molecular clock mechanism is driven by transcriptionaltranslational feedback loops of activator and repressor proteins. The transcriptional activators Circadian Locomotor Output Cycles Kaput (CLOCK) and Brain and Muscle ARNT-Like Protein 1 (BMAL1) heterodimerize and activate the transcription of the repressors Periods (Per1, Per2 and Per3) and Cryptochromes (Cry1 and Cry2) that in turn repress the activity of the CLOCK:BMAL1 complex. These cycles of activation and repression take about 24 h to complete and they generate oscillations in several clock-controlled genes (CCGs) with periods of ~24 h. It has been estimated that in any mammalian tissue, about 5-20% of the expressed genes exhibit circadian oscillations in their messenger RNA (mRNA) levels. Rhythms in mRNA abundance are the result of the interplay between the autonomous cellular clock and external time cues (i.e. the light/dark cycle and the fasting/feeding cycle). It has been demonstrated that the fasting/feeding cycle is the dominant Zeitgeber (“time giver”) that drives rhythms of mRNA abundance in peripheral tissues. It has been estimated that most cycling mRNAs do not have a corresponding rhythm in their nascent RNA, this suggested that post-transcriptional mechanism play an important role in generating mRNA and protein oscillation. However, little is known about the molecular mechanisms involved in generating mRNA rhythms at post-transcriptional level. The nuclear RNA binding protein Non-POU Domain Containing Octamer Binding (NONO) is a multifunctional protein that has been involved in many different steps of the mRNA life cycle from transcription, to processing, RNA stability and export. Furthermore, NONO is one of the main components of a subset of nuclear speckles called paraspeckles, whose function in the cell remains unclear. Importantly, NONO was found to interact with PER proteins and affect circadian ! ! Summary ! ! ! 2 rhythmicity in flies and mammals, however the specific mechanisms of its function in circadian RNA expression are unknown. In my thesis I have investigated the role of NONO in mammalian circadian gene expression and physiology. We found that feeding induces an increase in the number of NONO-containing, speckle-like structures in the nuclei of liver cells. In order to investigate NONO function in the fasting-feeding transition, we performed immunoprecipitation of NONO complexes in the liver nucleus and characterized bound RNAs and proteins. We found that NONO interacts with several RNA processing factors and that it primarily binds promoter-proximal introns of transcripts. This led us to hypothesize that NONO might be involved in the processing of its target RNAs. Furthermore, we found that the number of NONO-bound RNAs increases upon feeding and that about 40% of NONO target genes display circadian rhythmicity in their mRNA expression. We then showed that NONO regulates the rhythmicity and phase of these genes post-transcriptionally. To further explore the role of NONO in liver physiology we characterized the function of its bound RNAs. We found that a large fraction of circadian NONO-bound RNAs encodes proteins implicated in glucose uptake and macronutrient metabolism. The absence of NONO-mediated regulation of target RNAs profoundly impacts metabolic health. Indeed, NONO-deficient mice exhibit impaired glucose tolerance, which can be rescued by re-expression of NONO specifically in the liver. NONOdeficient mice also show reduced capacity to store glycogen and lipids in the liver. We also found that NONO-deficient mice burn more fat during the fasting (resting) phase and maintain a lean phenotype throughout their life. We therefore propose that NONO coordinates pre-mRNA processing of metabolic genes with the fasting/feeding cycle, thus allowing efficient energy utilization and storage. Disruption of the temporal coordination between metabolic demand and gene expression leads to the development of metabolic diseases, like obesity and diabetes. Our findings help to better understand how metabolic homeostasis is maintained in mammals, and identify novel therapeutic targets for treating diabetes and other associated metabolic dysfunctions.

Statistics

Downloads

20 downloads since deposited on 15 Feb 2018
20 downloads since 12 months
Detailed statistics

Additional indexing

Item Type:Dissertation
Referees:Brown Steven A, Arand Michael, Wolfrum Christian, Satchidananda Panda
Communities & Collections:04 Faculty of Medicine > Institute of Pharmacology and Toxicology
07 Faculty of Science > Institute of Pharmacology and Toxicology
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:27 October 2017
Deposited On:15 Feb 2018 09:10
Last Modified:19 Mar 2018 10:41
OA Status:Green

Download

Download PDF  'Role of the RNA-binding Protein NONO in Regulating Circadian Gene Expression and Metabolism'.
Preview
Content: Published Version
Filetype: PDF
Size: 7MB