scholarly journals miR-1 coordinately regulates lysosomal v-ATPase and biogenesis to impact proteotoxicity and muscle function during aging

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Isabelle Schiffer ◽  
Birgit Gerisch ◽  
Kazuto Kawamura ◽  
Raymond Laboy ◽  
Jennifer Hewitt ◽  
...  
Keyword(s):  
Nuclear Localization ◽  
Muscle Function ◽  
Muscle Development ◽  
Regulatory Subunit ◽  
Stress Growth ◽  
Atpase Subunits ◽  
Response To Stress ◽  
Pharyngeal Pumping ◽  
Direct Target ◽  
Lysosomal Acidification

Muscle function relies on the precise architecture of dynamic contractile elements, which must be fine-tuned to maintain motility throughout life. Muscle is also plastic, and remodeled in response to stress, growth, neural and metabolic inputs. The conserved muscle-enriched microRNA, miR-1, regulates distinct aspects of muscle development, but whether it plays a role during aging is unknown. Here we investigated Caenorhabditis elegans miR-1 in muscle function in response to proteostatic stress. mir-1 deletion improved mid-life muscle motility, pharyngeal pumping, and organismal longevity upon polyQ35 proteotoxic challenge. We identified multiple vacuolar ATPase subunits as subject to miR-1 control, and the regulatory subunit vha-13/ATP6V1A as a direct target downregulated via its 3′UTR to mediate miR-1 physiology. miR-1 further regulates nuclear localization of lysosomal biogenesis factor HLH-30/TFEB and lysosomal acidification. Our studies reveal that miR-1 coordinately regulates lysosomal v-ATPase and biogenesis to impact muscle function and health during aging.

scholarly journals miR-1 coordinately regulates lysosomal v-ATPase and biogenesis to affect muscle contractility upon proteotoxic challenge during ageing

2021 ◽  
Author(s):  
Isabelle Schiffer ◽  
Birgit Gerisch ◽  
Kazuto Kawamura ◽  
Raymond Laboy ◽  
Jennifer Hewitt ◽  
...  
Keyword(s):  
Muscle Function ◽  
Regulatory Subunit ◽  
C Elegans ◽  
Evolutionarily Conserved ◽  
Atpase Subunits ◽  
Muscle Biology ◽  
Response To Stress ◽  
Pharyngeal Pumping ◽  
Lysosomal Acidification

AbstractMuscle function relies on the precise architecture of dynamic contractile elements, which must be fine-tuned to maintain motility throughout life. Muscle is also highly plastic, and remodelled in response to stress, growth, neural and metabolic inputs. The evolutionarily conserved muscle-enriched microRNA, miR-1, regulates distinct aspects of muscle biology during development, but whether it plays a role during ageing is unknown. Here we investigated the role of C. elegans miR-1 in muscle function in response to proteostatic stress during adulthood. mir-1 deletion results in improved mid-life muscle motility, pharyngeal pumping, and organismal longevity under conditions of polyglutamine repeat proteotoxic challenge. We identified multiple vacuolar ATPase subunits as subject to miR-1 control, and the regulatory subunit vha-13/ATP6VIA as a direct target downregulated via its 3’UTR to mediate miR-1 physiology. miR-1 further regulates nuclear localization of lysosomal biogenesis factor HLH-30/TFEB and lysosomal acidification. In summary, our studies reveal that miR-1 coordinately regulates lysosomal v-ATPase and biogenesis to impact muscle function and health during ageing.

scholarly journals Integrated Regulation of PKA by Fast and Slow Neurotransmission in the Nucleus Accumbens Controls Plasticity and Stress Responses

2021 ◽  
Author(s):  
Rachel Thomas ◽  
Adan Hernandez ◽  
David R Benavides ◽  
Wei Li ◽  
Chunfeng Tan ◽  
...  
Keyword(s):  
Stress Responses ◽  
Regulatory Subunit ◽  
Signaling Mechanism ◽  
Acute And Chronic Stress ◽  
Dependent Protein ◽  
Response To Stress ◽  
Integrated Regulation ◽  
Midbrain Dopamine ◽  
Pka Regulatory Subunit ◽  
Neuropsychiatric Conditions

Cortical glutamate and midbrain dopamine neurotransmission converge to mediate striatum-dependent behaviors, while maladaptations in striatal circuitry contribute to mental disorders. Here we uncover a molecular mechanism by which glutamatergic and dopaminergic signaling integrate to regulate cAMP-dependent protein kinase (PKA) via phosphorylation of the PKA regulatory subunit, RIIβ. We find that glutamate-dependent reduction in Cdk5-dependent RIIβ phosphorylation alters the PKA holoenzyme auto-inhibitory state to increase PKA signaling in response to dopamine. Disruption of RIIβ phosphorylation by Cdk5, consequently, enhances cortico-ventral striatal synaptic plasticity. Acute and chronic stress in rats inversely modulates RIIβ phosphorylation and ventral striatal infusion of a small interfering peptide that selectively targets RIIβ regulation by Cdk5 improves behavioral response to stress. This new signaling mechanism integrating ventral striatal glutamate and dopamine neurotransmission is likely important to brain function, may contribute to neuropsychiatric conditions, and serves as a possible target for the development of novel therapeutics for stress-related disorders.

scholarly journals TORC1 signaling modulates Cdk8-dependent GAL gene expression in Saccharomyces cerevisiae

Genetics ◽  
2021 ◽  
Author(s):  
Riley Horvath ◽  
Nicole Hawe ◽  
Cindy Lam ◽  
Konstantin Mestnikov ◽  
Mariam Eji-Lasisi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Polymerase Ii ◽  
Nuclear Localization ◽  
Protein Phosphatase ◽  
Regulatory Subunit ◽  
Gal Genes ◽  
Dependent Site ◽  
Insight Into

Abstract Cdk8 of the RNA Polymerase II mediator kinase complex regulates gene expression by phosphorylating sequence-specific transcription factors. This function is conserved amongst eukaryotes, but the signals and mechanisms regulating Cdk8 activity and phosphorylation of its substrates are unknown. Full induction of the GAL genes in yeast requires phosphorylation of the transcriptional activator Gal4 by Cdk8. We used a screen to identify regulators of the Cdk8-dependent phosphorylation on Gal4, from which we identified multiple mutants with defects in TORC1 signaling. One mutant, designated gal four throttle 1 (gft1) was identified as a recessive allele of hom3, encoding aspartokinase, and mutations in hom3 caused effects typical of inhibition of TORC1, including rapamycin sensitivity and enhanced nuclear localization of the TORC1-responsive transcription factor Gat1. Mutations in hom3 also inhibit phosphorylation of Gal4 in vivo at the Cdk8-dependent site on Gal4, as did mutations of tor1, but these mutations did not affect activity of Cdk8 assayed in vitro. Disruption of cdc55, encoding a regulatory subunit of the TORC1-regulated protein phosphatase PP2A, suppressed the effect of hom3 and tor1 mutations on GAL expression, and also restored phosphorylation of Gal4 at the Cdk8-dependent site in vivo. These observations demonstrate that TORC1 signaling regulates GAL induction through the activity of PP2A/Cdc55, and suggest that Cdk8-dependent phosphorylation of Gal4 is opposed by PP2A/Cdc55 dephosphorylation. These results provide insight into how induction of transcription by a specific inducer can be modulated by global nutritional signals through regulation of Cdk8-dependent phosphorylation.

526 Late-Breaking: Poor Maternal Diet Alters the Muscle Proteome at Mid-gestation in Sheep

2021 ◽  
Vol 99 (Supplement_3) ◽  
pp. 149-150
Author(s):  
Sarah A Reed ◽  
Jeremy Balsbaugh ◽  
Xiaomeng Li ◽  
Timothy Moore ◽  
Amanda Jones ◽  
...  
Keyword(s):  
Fetal Development ◽  
Cellular Response ◽  
Muscle Development ◽  
Maternal Diet ◽  
Postnatal Growth ◽  
26S Proteasome ◽  
Mitogen Activated Protein Kinase ◽  
Mass Spectrometry Analysis ◽  
Initiation Factor ◽  
Regulatory Subunit

Abstract Maternal diet during gestation can negatively impact fetal development and postnatal growth and body composition. We hypothesized that poor maternal nutrition during gestation would alter protein expression in muscle at day 90 of gestation in sheep. Pregnant ewes (n = 24) were fed a control, restricted, or over-fed diet (100, 60, or 140% of NRC requirements for total digestible nutrients, respectively; n = 8 per treatment) starting at day 30 of gestation. At day 90 of gestation, fetal longissimus dorsi samples were collected (CON, RES, and OVER, respectively). Sarcoplasmic proteins were trypsin digested and subjected to multiplexed, label-based quantitative mass spectrometry analysis integrating Tandem Mass Tags. Differential expression of proteins was identified via two statistical techniques: ANOVA followed by Tukey’s HSD post-hoc tests, and regularized regression via elastic net. Significance was set at P < 0.05. Over-represented pathways containing differentially expressed proteins were identified by Reactome and included metabolism of proteins, immune system, cellular response to stress/external stimuli, developmental biology, and infectious disease. Within these pathways, 26S proteasome non-ATPase regulatory subunit (PSMD) 1, PSMD6, and PSMD14, components of the 26 S proteasome, were reduced 6 to 10% in OVER relative to CON offspring. Proteins involved in elongation and initiation, including eukaryotic initiation factor (EIF) 2 subunit 3, EIF 4 gamma 2, and EIF 3 subunit B were reduced 8 to 15% in OVER relative to CON offspring. Mitogen-activated protein kinase (MAPK) 1 and dual-specificity MAPK kinase (MEKK) 1 were increased by 12% in RES relative to OVER and CON offspring. Together, these data indicate that protein degradation and synthesis are altered in mid-gestation muscle development in OVER offspring, and that cell signaling related to cell proliferation is affected in RES offspring. These changes may contribute to the phenotypic and metabolic changes observed during fetal development and postnatal growth.

scholarly journals Disruption of Drosophila larval muscle structure and function by UNC45 knockdown

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Abiramy Karunendiran ◽  
Christine T. Nguyen ◽  
Virginijus Barzda ◽  
Bryan A. Stewart
Keyword(s):  
Muscle Function ◽  
Muscle Development ◽  
Protein Complexes ◽  
Second Harmonic ◽  
Pupal Stage ◽  
Microscopy Imaging ◽  
Synaptic Physiology ◽  
Late Embryogenesis ◽  
Body Wall Muscles ◽  
And Function

Abstract Background Proper muscle function is heavily dependent on highly ordered protein complexes. UNC45 is a USC (named since this region is shared by three proteins UNC45/CRO1/She4P) chaperone that is necessary for myosin incorporation into the thick filaments. UNC45 is expressed throughout the entire Drosophila life cycle and it has been shown to be important during late embryogenesis when initial muscle development occurs. However, the effects of UNC45 manipulation at later developmental times, after muscle development, have not yet been studied. Main results UNC45 was knocked down with RNAi in a manner that permitted survival to the pupal stage, allowing for characterization of sarcomere organization in the well-studied third instar larvae. Second harmonic generation (SHG) microscopy revealed changes in the striated pattern of body wall muscles as well as a reduction of signal intensity. This observation was confirmed with immunofluorescence and electron microscopy imaging, showing diminished UNC45 signal and disorganization of myosin and z-disk proteins. Concomitant alterations in both synaptic physiology and locomotor function were also found. Both nerve-stimulated response and spontaneous vesicle release were negatively affected, while larval movement was impaired. Conclusions This study highlights the dependency of normal sarcomere structure on UNC45 expression. We confirm the known role of UNC45 for myosin localization and further show the I-Z-I complex is also disrupted. This suggests a broad need for UNC45 to maintain sarcomere integrity. Newly discovered changes in synaptic physiology reveal the likely presence of a homeostatic response to partially maintain synaptic strength and muscle function.

scholarly journals Exercise attenuates juvenile irradiation-induced skeletal muscle decline by improving calcium handling and decreasing mitochondrial stress

2021 ◽  
Vol 154 (9) ◽  
Author(s):  
Thomas N. O’Connor ◽  
Jacob G. Kallenbach ◽  
Joe V. Chakkalakal ◽  
Robert T. Dirksen
Keyword(s):  
Skeletal Muscle ◽  
Childhood Cancer ◽  
Muscle Function ◽  
Muscle Development ◽  
Radiation Treatment ◽  
Wheel Running ◽  
Calcium Handling ◽  
Radiation Induced ◽  
Effects Of Radiation ◽  
And Function

Proper skeletal muscle development, maintenance, and function is necessary for movement. Decline in muscle function with age and disease is directly associated with a diminished quality of life. Radiation therapy is commonly used to treat certain forms of childhood cancer based on the cytotoxic effects of radiation on cancerous tissue. However, the adverse effects elicited by radiation are not always constrained to the diseased tissue and can accelerate muscle wasting and decline, which is particularly detrimental to juvenile cancer survivors. Exercise is effective at limiting muscle decline and improving muscle function in various diseases. Thus, we hypothesized 1 mo of voluntary endurance exercise following juvenile radiation treatment will reduce muscle damage and restore functional deficits that occur following radiation. Here, we show that following juvenile radiation, 1 mo of voluntary wheel running significantly improved muscle function in mice by promoting adaptations in intracellular calcium handling, improving mitochondrial turnover and reducing oxidative stress resulting from radiation-induced mitochondrial damage. These findings help guide caregivers in their approach to childhood cancer survivor recovery and have implications for other diseases where similar mechanisms of calcium handling and mitochondrial function are disrupted.

scholarly journals mRNA decapping is an evolutionarily conserved modulator of neuroendocrine signaling that controls development and ageing

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Fivos Borbolis ◽  
John Rallis ◽  
George Kanatouris ◽  
Nikolitsa Kokla ◽  
Antonis Karamalegkos ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nuclear Localization ◽  
Normal Development ◽  
Model Organisms ◽  
Mrna Decapping ◽  
Evolutionarily Conserved ◽  
Stress Survival ◽  
Response To Stress ◽  
Decapping Enzyme ◽  
Wing Morphogenesis

Eukaryotic 5’−3’ mRNA decay plays important roles during development and in response to stress, regulating gene expression post-transcriptionally. In Caenorhabditis elegans, deficiency of DCAP-1/DCP1, the essential co-factor of the major cytoplasmic mRNA decapping enzyme, impacts normal development, stress survival and ageing. Here, we show that overexpression of dcap-1 in neurons of worms is sufficient to increase lifespan through the function of the insulin/IGF-like signaling and its effector DAF-16/FOXO transcription factor. Neuronal DCAP-1 affects basal levels of INS-7, an ageing-related insulin-like peptide, which acts in the intestine to determine lifespan. Short-lived dcap-1 mutants exhibit a neurosecretion-dependent upregulation of intestinal ins-7 transcription, and diminished nuclear localization of DAF-16/FOXO. Moreover, neuronal overexpression of DCP1 in Drosophila melanogaster confers longevity in adults, while neuronal DCP1 deficiency shortens lifespan and affects wing morphogenesis, cell non-autonomously. Our genetic analysis in two model-organisms suggests a critical and conserved function of DCAP-1/DCP1 in developmental events and lifespan modulation.

scholarly journals The number of growing microtubules and nucleus-nucleus interactions uniquely regulate nuclear movement in Drosophila muscle

2020 ◽  
Author(s):  
Mary Ann Collins ◽  
L. Alexis Coon ◽  
Riya Thomas ◽  
Torrey R. Mandigo ◽  
Elizabeth Wynn ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Muscle Function ◽  
Cell Biology ◽  
Muscle Development ◽  
Eukaryotic Cell ◽  
Model System ◽  
Nuclear Movement ◽  
Precise Positioning ◽  
Live Embryo ◽  
Fundamental Process

ABSTRACTNuclear movement is a fundamental process of eukaryotic cell biology. Skeletal muscle presents an intriguing model to study nuclear movement because its development requires the precise positioning of multiple nuclei within a single cytoplasm. Furthermore, there is a high correlation between aberrant nuclear positioning and poor muscle function. Although many genes that regulate nuclear movement have been identified, the mechanisms by which these genes act is not known. Using Drosophila melanogaster muscle development as a model system, and a combination of live-embryo microscopy and laser ablation of nuclei, we have found that phenotypically similar mutants are based in different molecular disruptions. Specifically, ensconsin (Drosophila MAP7) regulates the number of growing microtubules that are used to move nuclei whereas bocksbeutel (Drosophila emerin) and klarsicht (Drosophila KASH-protein regulate interactions between nuclei.

scholarly journals Functional Characterization of Entamoeba histolytica Argonaute Proteins Reveals a Repetitive DR-Rich Motif Region That Controls Nuclear Localization

mSphere ◽  
2019 ◽  
Vol 4 (5) ◽  
Author(s):  
Hanbang Zhang ◽  
Vy Tran ◽  
Dipak Manna ◽  
Gretchen Ehrenkaufer ◽  
Upinder Singh
Keyword(s):  
Gene Silencing ◽  
Entamoeba Histolytica ◽  
Nuclear Localization ◽  
Functional Characterization ◽  
Protozoan Parasite ◽  
Rnai Pathway ◽  
Content Type ◽  
Localization Signal ◽  
Response To Stress ◽  
Paz Domain

ABSTRACT The RNA interference (RNAi) pathway regulates gene expression in many eukaryotic organisms. Argonaute (Ago) proteins, together with bound small RNAs (sRNAs), are key effectors that mediate gene silencing function. However, there is limited knowledge of Ago proteins and their functions in nonmodel systems. In the protozoan parasite Entamoeba histolytica, RNAi is a robust means for stable gene silencing mediated via large populations of antisense sRNAs. Here, we report functional characterization of three Ago proteins in E. histolytica (EhAgo2-1, EhAgo2-2, and EhAgo2-3). Our data show that each EhAgo protein has a distinct subcellular localization and binds 27-nucleotide (nt) sRNAs and that the localization of EhAgo proteins is altered in response to stress conditions. Via mutagenesis analyses, we demonstrated that the Ago PAZ (Piwi/Argonaute/Zwille) domain in all three EhAgos is essential for sRNA binding. With mutation of the PAZ domain in EhAgo2-2, there was no effect on the nuclear localization of the protein but a strong phenotype and a growth defect. We further show that EhAgo2-2 contains an unusual repetitive DR-rich (aspartic acid, arginine-rich) motif region which functions as a nuclear localization signal (NLS) and is both necessary and sufficient to mediate nuclear localization. Overall, our data delineate the localization and sRNA binding features of the three E. histolytica Ago proteins and demonstrate that the PAZ domain is necessary for sRNA binding. The repetitive DR-rich motif region in EhAgo2-2 has not previously been defined in other systems, which adds to the novel observations that can be made when studies of the RNAi pathway are extended to nonmodel systems. IMPORTANCE The protozoan parasite Entamoeba histolytica, which causes amebiasis and affects over 50 million people worldwide, contains an important RNAi pathway for gene silencing. Gene silencing via the RNAi pathway is mediated by the Argonaute (Ago) proteins. However, we lack knowledge on Ago function(s) in this nonmodel system. In this paper, we discovered that three E. histolytica Ago proteins (EhAgo2-1, EhAgo2-2, and EhAgo2-3) all bind 27-nt small RNAs and have distinct subcellular localizations, which change in response to stress conditions. The EhAgos bind small RNA populations via their PAZ domains. An unusual repetitive DR-rich motif region is identified in EhAgo2-2 that functions as a nuclear localization signal. Our results show for the first time an active nuclear transport process of the EhAgo2-2 RNA-induced silencing complex (RISC) in this parasite. These data add to the novel observations that can be made when studies of the RNAi pathway are extended to nonmodel systems.

scholarly journals High NaCl-induced inhibition of PTG contributes to activation of NFAT5 through attenuation of the negative effect of SHP-1

2013 ◽  
Vol 305 (3) ◽  
pp. F362-F369 ◽  
Author(s):  
Xiaoming Zhou ◽  
Hong Wang ◽  
Maurice B. Burg ◽  
Joan D. Ferraris
Keyword(s):  
Transcription Factor ◽  
Nuclear Localization ◽  
Transcriptional Activity ◽  
Protein Targeting ◽  
Inhibitory Effect ◽  
Catalytic Subunit ◽  
Regulatory Subunit ◽  
Protein Abundance ◽  
Negative Effect ◽  
Sirna Screening

Activation of the transcription factor NFAT5 by high NaCl involves changes in phosphorylation. By siRNA screening, we previously found that protein targeting to glycogen (PTG), a regulatory subunit of protein phosphatase1 (PP1), contributes to regulation of high NaCl-induced NFAT5 transcriptional activity. The present study addresses the mechanism involved. We find that high NaCl-induced inhibition of PTG elevates NFAT5 activity by increasing NFAT5 transactivating activity, protein abundance, and nuclear localization. PTG acts via a catalytic subunit PP1γ. PTG associates physically with PP1γ, and NaCl reduces both this association and remaining PTG-associated PP1γ activity. High NaCl-induced phosphorylation of p38, ERK, and SHP-1 contributes to activation of NFAT5. Knockdown of PTG does not affect phosphorylation of p38 or ERK. However, PTG and PP1γ bind to SHP-1, and knockdown of either PTG or PP1γ increases high NaCl-induced phosphorylation of SHP-1-S591, which inhibits SHP-1. Mutation of SHP-1-S591 to alanine, which cannot be phosphorylated, increases inhibition of NFAT5 by SHP-1. Thus high NaCl reduces the stimulatory effect of PTG and PP1γ on SHP-1, which in turn reduces the inhibitory effect of SHP-1 on NFAT5. Our findings add to the known functions of PTG, which was previously recognized only for its glycogenic activity.

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