Sleep Need, the Key Regulator of Sleep Homeostasis, Is Indicated and Controlled by Phosphorylation of Threonine 221 in Salt Inducible Kinase 3

Sleep need drives sleep and plays a key role in homeostatic regulation of sleep. So far sleep need can only be inferred by animal behaviors and indicated by electroencephalography (EEG). Here we report that threonine 221 (T221) of the salt inducible kinase 3 (SIK3) was important for the catalytic activity and stability of SIK3. T221 phosphorylation in the mouse brain indicates sleep need: more sleep resulting in less phosphorylation and less sleep more phosphorylation during daily sleep/wake cycle and after sleep deprivation (SD). Sleep need was reduced in SIK3 loss of function (LOF) mutants and by T221 mutation to alanine (T221A). Sleep rebound after SD was also decreased in SIK3 LOF and T221A mutant mice. Other kinases such as SIK1 and SIK2 or other sites in SIK3 do not fulfil criteria to be both an indicator and a controller of sleep need. Our results reveal SIK3 T221 phosphorylation as the first and only chemical modification which indicates and controls sleep need.

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Seasonal variation in sleep homeostasis in migratory geese: a rebound of NREM sleep following sleep deprivation in summer but not in winter

SLEEP ◽

10.1093/sleep/zsaa244 ◽

2020 ◽

Cited By ~ 1

Author(s):

Sjoerd J van Hasselt ◽

Gert-Jan Mekenkamp ◽

Jan Komdeur ◽

Giancarlo Allocca ◽

Alexei L Vyssotski ◽

...

Keyword(s):

Sleep Deprivation ◽

Physiological State ◽

Spectral Composition ◽

Nrem Sleep ◽

Sleep Time ◽

Lunar Cycle ◽

Homeostatic Regulation ◽

Data Loggers ◽

Sleep Homeostasis ◽

Barnacle Geese

Abstract Sleep is a behavioral and physiological state that is thought to serve important functions. Many animals go through phases in the annual cycle where sleep time might be limited, for example, during the migration and breeding phases. This leads to the question whether there are seasonal changes in sleep homeostasis. Using electroencephalogram (EEG) data loggers, we measured sleep in summer and winter in 13 barnacle geese (Branta leucopsis) under semi-natural conditions. During both seasons, we examined the homeostatic regulation of sleep by depriving the birds of sleep for 4 and 8 h after sunset. In winter, barnacle geese showed a clear diurnal rhythm in sleep and wakefulness. In summer, this rhythm was less pronounced, with sleep being spread out over the 24-h cycle. On average, the geese slept 1.5 h less per day in summer compared with winter. In both seasons, the amount of NREM sleep was additionally affected by the lunar cycle, with 2 h NREM sleep less during full moon compared to new moon. During summer, the geese responded to 4 and 8 h of sleep deprivation with a compensatory increase in NREM sleep time. In winter, this homeostatic response was absent. Overall, sleep deprivation only resulted in minor changes in the spectral composition of the sleep EEG. In conclusion, barnacle geese display season-dependent homeostatic regulation of sleep. These results demonstrate that sleep homeostasis is not a rigid phenomenon and suggest that some species may tolerate sleep loss under certain conditions or during certain periods of the year.

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Sleep after arousal from hibernation is not homeostatically regulated

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1999.276.2.r522 ◽

1999 ◽

Vol 276 (2) ◽

pp. R522-R529 ◽

Cited By ~ 10

Author(s):

Jennie E. Larkin ◽

H. Craig Heller

Keyword(s):

Sleep Deprivation ◽

Brain Function ◽

Extended Period ◽

Nrem Sleep ◽

Wave Activity ◽

Ground Squirrels ◽

Recovery Sleep ◽

Sleep Homeostasis ◽

Homeostatic Response ◽

Periodic Arousals

Electroencephalographic slow-wave activity (SWA) in non-rapid eye movement (NREM) sleep is directly related to prior sleep/wake history, with high levels of SWA following extended periods of wake. Therefore, SWA has been thought to reflect the level of accumulated sleep need. The discovery that euthermic intervals between hibernation bouts are spent primarily in sleep and that this sleep is characterized by high and monotonically declining SWA has led to speculation that sleep homeostasis may play a fundamental role in the regulation of the timing of bouts of hibernation and periodic arousals to euthermia. It was proposed that because the SWA profile seen after arousal from hibernation is strikingly similar to what is seen in nonhibernating mammals after extended periods of wakefulness, that hibernating mammals may arouse from hibernation with significant accumulated sleep need. This sleep need may accumulate during hibernation because the low brain temperatures during hibernation may not be compatible with sleep restorative processes. In the present study, golden-mantled ground squirrels were sleep deprived during the first 4 h of interbout euthermia by injection of caffeine (20 mg/kg ip). We predicted that if the SWA peaks after bouts of hibernation reflected a homeostatic response to an accumulated sleep need, sleep deprivation should simply have displaced and possibly augmented the SWA to subsequent recovery sleep. Instead we found that after caffeine-induced sleep deprivation of animals just aroused from hibernation, the anticipated high SWA typical of recovery sleep did not occur. Similar results were found in a study that induced sleep deprivation by gentle handling (19). These findings indicate that the SWA peak immediately after hibernation does not represent homeostatic regulation of NREM sleep, as it normally does after prolonged wakefulness during euthermia, but instead may reflect some other neurological process in the recovery of brain function from an extended period at low temperature.

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Impaired Response to Sleep Deprivation in Heterozygous Disc1 Mutant Mice

The World Journal of Biological Psychiatry ◽

10.1080/15622975.2021.1907724 ◽

2021 ◽

pp. 1-30

Author(s):

Chih-Yu Tsao ◽

Li-Heng Tuan ◽

Lukas Jyuhn-Hsiarn Lee ◽

Chih-Min Liu ◽

Hai-Gwo Hwu ◽

...

Keyword(s):

Sleep Deprivation ◽

Mutant Mice

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Aicardi-Goutières syndrome-associated mutation at ADAR1 gene locus activates innate immune response in mouse brain

Journal of Neuroinflammation ◽

10.1186/s12974-021-02217-9 ◽

2021 ◽

Vol 18 (1) ◽

Author(s):

Xinfeng Guo ◽

Clayton A. Wiley ◽

Richard A. Steinman ◽

Yi Sheng ◽

Beihong Ji ◽

...

Keyword(s):

Rna Sequencing ◽

Rna Editing ◽

Mouse Brain ◽

Mutant Mouse ◽

Cell Types ◽

Mutant Mice ◽

Blue Staining ◽

Sequencing Analysis ◽

Elevated Type ◽

The Brain

Abstract Background Aicardi-Goutières syndrome (AGS) is a severe infant or juvenile-onset autoimmune disease characterized by inflammatory encephalopathy with an elevated type 1 interferon-stimulated gene (ISG) expression signature in the brain. Mutations in seven different protein-coding genes, all linked to DNA/RNA metabolism or sensing, have been identified in AGS patients, but none of them has been demonstrated to activate the IFN pathway in the brain of an animal. The molecular mechanism of inflammatory encephalopathy in AGS has not been well defined. Adenosine Deaminase Acting on RNA 1 (ADAR1) is one of the AGS-associated genes. It carries out A-to-I RNA editing that converts adenosine to inosine at double-stranded RNA regions. Whether an AGS-associated mutation in ADAR1 activates the IFN pathway and causes autoimmune pathogenesis in the brain is yet to be determined. Methods Mutations in the ADAR1 gene found in AGS patients were introduced into the mouse genome via CRISPR/Cas9 technology. Molecular activities of the specific p.K999N mutation were investigated by measuring the RNA editing levels in brain mRNA substrates of ADAR1 through RNA sequencing analysis. IFN pathway activation in the brain was assessed by measuring ISG expression at the mRNA and protein level through real-time RT-PCR and Luminex assays, respectively. The locations in the brain and neural cell types that express ISGs were determined by RNA in situ hybridization (ISH). Potential AGS-related brain morphologic changes were assessed with immunohistological analysis. Von Kossa and Luxol Fast Blue staining was performed on brain tissue to assess calcification and myelin, respectively. Results Mice bearing the ADAR1 p.K999N were viable though smaller than wild type sibs. RNA sequencing analysis of neuron-specific RNA substrates revealed altered RNA editing activities of the mutant ADAR1 protein. Mutant mice exhibited dramatically elevated levels of multiple ISGs within the brain. RNA ISH of brain sections showed selective activation of ISG expression in neurons and microglia in a patchy pattern. ISG-15 mRNA was upregulated in ADAR1 mutant brain neurons whereas CXCL10 mRNA was elevated in adjacent astroglia. No calcification or gliosis was detected in the mutant brain. Conclusions We demonstrated that an AGS-associated mutation in ADAR1, specifically the p.K999N mutation, activates the IFN pathway in the mouse brain. The ADAR1 p.K999N mutant mouse replicates aspects of the brain interferonopathy of AGS. Neurons and microglia express different ISGs. Basal ganglia calcification and leukodystrophy seen in AGS patients were not observed in K999N mutant mice, indicating that development of the full clinical phenotype may need an additional stimulus besides AGS mutations. This mutant mouse presents a robust tool for the investigation of AGS and neuroinflammatory diseases including the modeling of potential “second hits” that enable severe phenotypes of clinically variable diseases.

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Genetic analysis of Pycr1 and Pycr2 in mice

Genetics ◽

10.1093/genetics/iyab048 ◽

2021 ◽

Author(s):

Morgane G Stum ◽

Abigail L D Tadenev ◽

Kevin L Seburn ◽

Kathy E Miers ◽

Pak P Poon ◽

...

Keyword(s):

Neuromuscular Disorders ◽

Subcutaneous Fat ◽

Mutant Mice ◽

Loss Of Function ◽

Proline Biosynthesis ◽

Cell Counts ◽

Chemically Induced ◽

White Blood Cell Counts ◽

Redundant Functions ◽

Altered Lipid

Abstract The final step in proline biosynthesis is catalyzed by three pyrroline-5-carboxylate reductases, PYCR1, PYCR2, and PYCR3, which convert pyrroline-5-carboxylate (P5C) to proline. Mutations in human PYCR1 and ALDH18A1 (P5C Synthetase) cause Cutis Laxa (CL), whereas mutations in PYCR2 cause hypomyelinating leukodystrophy 10 (HLD10). Here, we investigated the genetics of Pycr1 and Pycr2 in mice. A null allele of Pycr1 did not show integument or CL-related phenotypes. We also studied a novel chemically-induced mutation in Pycr2. Mice with recessive loss-of-function mutations in Pycr2 showed phenotypes consistent with neurological and neuromuscular disorders, including weight loss, kyphosis, and hind-limb clasping. The peripheral nervous system was largely unaffected, with only mild axonal atrophy in peripheral nerves. A severe loss of subcutaneous fat in Pycr2 mutant mice is reminiscent of a CL-like phenotype, but primary features such as elastin abnormalities were not observed. Aged Pycr2 mutant mice had reduced white blood cell counts and altered lipid metabolism, suggesting a generalized metabolic disorder. PYCR1 and -2 have similar enzymatic and cellular activities, and consistent with previous studies, both were localized in the mitochondria in fibroblasts. Both PYCR1 and -2 were able to complement the loss of Pro3, the yeast enzyme that converts P5C to proline, confirming their activity as P5C reductases. In mice, Pycr1; Pycr2 double mutants were sub-viable and unhealthy compared to either single mutant, indicating the genes are largely functionally redundant. Proline levels were not reduced, and precursors were not increased in serum from Pycr2 mutant mice or in lysates from skin fibroblast cultures, but placing Pycr2 mutant mice on a proline-free diet worsened the phenotype. Thus, Pycr1 and -2 have redundant functions in proline biosynthesis, and their loss makes proline a semi-essential amino acid. These findings have implications for understanding the genetics of CL and HLD10, and for modeling these disorders in mice.

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GUCY2D Gene Loss-of-Function Mutations Responsible for Leber Congenital Amaurosis 1

10.21203/rs.2.11649/v1 ◽

2019 ◽

Author(s):

Feng Xue ◽

Tianying Wei ◽

Junhui Sun ◽

Yuqin Luo ◽

Yanan Huo ◽

...

Keyword(s):

Catalytic Activity ◽

Guanylate Cyclase ◽

Cyclic Guanosine Monophosphate ◽

Guanosine Monophosphate ◽

Loss Of Function ◽

Wild Type ◽

Leber Congenital Amaurosis ◽

Cellular Location ◽

Next Generation Sequencing Ngs ◽

Generation Sequencing

Abstract Background: Leber congenital amaurosis (LCA) is a group of severe congenital neurodegenerative diseases. Variants in guanylate cyclase 2D (GUCY2D), which encoded guanylate cyclase protein (ROS-GC1) associate with LCA1, accounting for 6–21% of all LCA cases. Methods: In this study, one family with LCA1 was recruited from China. A combination of next-generation sequencing (NGS) and Sanger sequencing was used for disease-causing mutations screening. Additionally, immunohistochemistry and HPLC-coupled tandem mass-spectrometry (HPLC-MS/MS) were used to confirm the cellular location and catalytic activity of ROS-GC1 mutants, respectively. Results: We found three novel mutations (c.139_139delC, c.835G>A and c.2783G>A) in GUCY2D gene. The results showed that mutation c.139_139delC results in a truncated protein and destroys the structure of ROS-GC1 protein. Mutations c.835G>A and c.2783G>A exert no effects on cellular location, whereas significantly reduce the catalytic activity of ROS-GC1. Conclusions: Our findings highlight the clinical range of LCA. Moreover we used HPLC-MS/MS to analyze the concentration of 3', 5'-cyclic guanosine monophosphate (cGMP), suggesting that HPLC-MS/MS can be an effective alternative method to evaluate the catalytic activity of wild type (wt) and mutant ROS-GC1.

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Human xylosyltransferase I and N-terminal truncated forms: functional characterization of the core enzyme

Biochemical Journal ◽

10.1042/bj20051606 ◽

2006 ◽

Vol 394 (1) ◽

pp. 163-171 ◽

Cited By ~ 11

Author(s):

Sandra Müller ◽

Jennifer Disse ◽

Manuela Schöttler ◽

Sylvia Schön ◽

Christian Prante ◽

...

Keyword(s):

Amino Acids ◽

Catalytic Activity ◽

Binding Capacity ◽

Functional Characterization ◽

Terminal Region ◽

Binding Motif ◽

Heparin Binding ◽

Loss Of Function ◽

The Impact

Human XT-I (xylosyltransferase I; EC 2.4.2.26) initiates the biosynthesis of the glycosaminoglycan linkage region and is a diagnostic marker of an enhanced proteoglycan biosynthesis. In the present study, we have investigated mutant enzymes of human XT-I and assessed the impact of the N-terminal region on the enzymatic activity. Soluble mutant enzymes of human XT-I with deletions at the N-terminal domain were expressed in insect cells and analysed for catalytic activity. As many as 260 amino acids could be truncated at the N-terminal region of the enzyme without affecting its catalytic activity. However, truncation of 266, 272 and 273 amino acids resulted in a 70, 90 and >98% loss in catalytic activity. Interestingly, deletion of the single 12 amino acid motif G261KEAISALSRAK272 leads to a loss-of-function XT-I mutant. This is in agreement with our findings analysing the importance of the Cys residues where we have shown that C276A mutation resulted in a nearly inactive XT-I enzyme. Moreover, we investigated the location of the heparin-binding site of human XT-I using the truncated mutants. Heparin binding was observed to be slightly altered in mutants lacking 289 or 568 amino acids, but deletion of the potential heparin-binding motif P721KKVFKI727 did not lead to a loss of heparin binding capacity. The effect of heparin or UDP on the XT-I activity of all mutants was not significantly different from that of the wild-type. Our study demonstrates that over 80% of the nucleotide sequence of the XT-I-cDNA is necessary for expressing a recombinant enzyme with full catalytic activity.

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Homeostatic regulation through GABA and acetylcholine muscarinic receptors of motor trigeminal neurons following sleep deprivation

Brain Structure and Function ◽

10.1007/s00429-017-1392-4 ◽

2017 ◽

Vol 222 (7) ◽

pp. 3163-3178 ◽

Cited By ~ 4

Author(s):

Hanieh Toossi ◽

Esther Del Cid-Pellitero ◽

Barbara E. Jones

Keyword(s):

Sleep Deprivation ◽

Muscarinic Receptors ◽

Homeostatic Regulation ◽

Trigeminal Neurons

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The European starling (Sturnus vulgaris) shows signs of NREM sleep homeostasis but has very little REM sleep and no REM sleep homeostasis

SLEEP ◽

10.1093/sleep/zsz311 ◽

2019 ◽

Vol 43 (6) ◽

Cited By ~ 4

Author(s):

Sjoerd J van Hasselt ◽

Maria Rusche ◽

Alexei L Vyssotski ◽

Simon Verhulst ◽

Niels C Rattenborg ◽

...

Keyword(s):

Sleep Deprivation ◽

Eye Movement ◽

Rem Sleep ◽

Spectral Power ◽

Nrem Sleep ◽

Sleep Time ◽

European Starling ◽

Dark Phase ◽

Rapid Eye Movement ◽

Sleep Homeostasis

Abstract Most of our knowledge about the regulation and function of sleep is based on studies in a restricted number of mammalian species, particularly nocturnal rodents. Hence, there is still much to learn from comparative studies in other species. Birds are interesting because they appear to share key aspects of sleep with mammals, including the presence of two different forms of sleep, i.e. non-rapid eye movement (NREM) and rapid eye movement (REM) sleep. We examined sleep architecture and sleep homeostasis in the European starling, using miniature dataloggers for electroencephalogram (EEG) recordings. Under controlled laboratory conditions with a 12:12 h light–dark cycle, the birds displayed a pronounced daily rhythm in sleep and wakefulness with most sleep occurring during the dark phase. Sleep mainly consisted of NREM sleep. In fact, the amount of REM sleep added up to only 1~2% of total sleep time. Animals were subjected to 4 or 8 h sleep deprivation to assess sleep homeostatic responses. Sleep deprivation induced changes in subsequent NREM sleep EEG spectral qualities for several hours, with increased spectral power from 1.17 Hz up to at least 25 Hz. In contrast, power below 1.17 Hz was decreased after sleep deprivation. Sleep deprivation also resulted in a small compensatory increase in NREM sleep time the next day. Changes in EEG spectral power and sleep time were largely similar after 4 and 8 h sleep deprivation. REM sleep was not noticeably compensated after sleep deprivation. In conclusion, starlings display signs of NREM sleep homeostasis but the results do not support the notion of important REM sleep functions.

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