Dataset Information

Silencing of eIF5B Changes Proteostasis by Providing Negative Feedback on MAPK Signaling

ABSTRACT: Protein translational machinery is an important component of proteostasis network (PN) that maintains cellular proteostasis and regulates aging and other cellular processes. Ample evidence indicates that inhibition of translation initiation factor activities enhances stress resistance and extends life span in model organisms. Eukaryotic translation initiation factor 5B (eIF5B) is responsible for joining of pre-40S subunits with 60S ribosomal subunits to give a 80S-like complex in protein translational initiation and its silencing may disrupt proteostasis and trigger cellular processes associated with stress responses. In the present work, eIF5B was genetically manipulated in 293T cells. The physiological aspects of eIF5B-knockdown cells (eIF5B-KN) were characterized showing that they grew slower, had a lower level of intracellular reactive oxygen species (ROS), an increased resistance to oxidative stress and an enhanced autophagy. Proteomic analysis showed that silencing of eIF5B resulted in up-regulation of 88 proteins and down-regulation of 130 proteins in eIF5B-KN compared to control cells, which involved in diverse cellular processes including metabolism, RNA processing, and protein metabolism, and DNA synthesis. The autonomous downregulation of the MAPK singnaling pathway was identified that led to the prolonged S-phase cell-cycle arrest and contributed to the slow growth of eIF5B-KN cells. Glutamine transporters were found to be downregulated which enhanced formation of autophagy. Furthermore, eIF5B knockdown compromised the integrity of 28S rRNA and 8.5.8S rRNA that can be rescued via restoring the eIF5B expression level. Taken together, these results demonstrated that eIF5B silencing providesd a negative feedback to down regulate the MAPK signaling pathway which reconstitutesd the proteostasis, resulting in a decrease in cell growth and an enhanced resistance to oxidative stress. Our data provide a useful resource to further biological exploration into functions of protein synthesis in regulation of proteostasis and aging and suggest that eIF5B plays a role in aging process.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Kidney

SUBMITTER: Xu Jiang

LAB HEAD: Haiteng Deng

PROVIDER: PXD003111 | Pride | 2016-06-13

REPOSITORIES: Pride

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Publications

Proteomics data in support of the quantification of the changes of bovine milk proteins during mammary gland involution.

Boggs Irina I Hine Brad B Smolenksi Grant G Hettinga Kasper K Zhang Lina L Wheeler Thomas T TT

Data in brief 20160513

Here we provide data from three proteomics techniques; two-dimensional electrophoresis (2-DE) followed by identification of selected spots using PSD MALDI-TOF MS/MS, one-dimensional gel electrophoresis followed by LC-MS/MS analysis of gel slices (GeLC) and dimethyl isotopic labelling of tryptic peptides followed by Orbitrap MS/MS (DML), to quantify the changes in the repertoire of bovine milk proteins that occurs after drying off. We analysed skim milk and whey sampled at day 0 and either day 3 ...[more]

PMID: 27274532

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Project description:Protein translational machinery is an important component of proteostasis network (PN) that maintains cellular proteostasis and regulates aging and other cellular processes. Ample evidence indicates that inhibition of translation initiation factor activities enhances stress resistance and extends life span in model organisms. Eukaryotic translation initiation factor 5B (eIF5B) is responsible for joining of pre-40S subunits with 60S ribosomal subunits to give a 80S-like complex in protein translational initiation and its silencing may disrupt proteostasis and trigger cellular processes associated with stress responses. In the present work, eIF5B was genetically manipulated in 293T cells. The physiological aspects of eIF5B-knockdown cells (eIF5B-KN) were characterized showing that they grew slower, had a lower level of intracellular reactive oxygen species (ROS), an increased resistance to oxidative stress and an enhanced autophagy. Proteomic analysis showed that silencing of eIF5B resulted in up-regulation of 88 proteins and down-regulation of 130 proteins in eIF5B-KN compared to control cells, which involved in diverse cellular processes including metabolism, RNA processing, and protein metabolism, and DNA synthesis. The autonomous downregulation of the MAPK singnaling pathway was identified that led to the prolonged S-phase cell-cycle arrest and contributed to the slow growth of eIF5B-KN cells. Glutamine transporters were found to be downregulated which enhanced formation of autophagy. Furthermore, eIF5B knockdown compromised the integrity of 28S rRNA and 8.5.8S rRNA that can be rescued via restoring the eIF5B expression level. Taken together, these results demonstrated that eIF5B silencing providesd a negative feedback to down regulate the MAPK signaling pathway which reconstitutesd the proteostasis, resulting in a decrease in cell growth and an enhanced resistance to oxidative stress. Our data provide a useful resource to further biological exploration into functions of protein synthesis in regulation of proteostasis and aging and suggest that eIF5B plays a role in aging process.

Project description:Protein function is controlled by the cellular proteostasis network. Proteostasis is energetically costly and those costs must be balanced with the energy needs of other physiological functions. Hypertonic stress causes widespread protein damage in C. elegans. Suppression and management of protein damage is essential for optimal survival under hypertonic conditions. ASH chemosensory neurons allow C. elegans to detect and avoid strongly hypertonic environments. We demonstrate that gene mutations that disrupt ASH mediated hypertonic avoidance behavior or genetic ablation of ASH neurons enhance survival during hypertonic stress. Enhanced survival is not due to altered systemic volume homeostasis or organic osmolyte accumulation. Instead, loss of ASH neuron function reduces protein damage in non-neuronal cells. Improved proteostasis capacity is due in part to upregulation of genes that play important roles in managing protein damage. We propose that inhibitory signaling from ASH neurons normally suppresses expression of genes required for non-neuronal cell proteostasis. Because all cells have the capacity to sense and respond to stressors, inhibitory neuronal signaling may be important for minimizing activation of cellular stress resistance and proteostasis pathways during short duration and less extreme stressors or stressors that can be avoided by behavioral changes. Neuronal regulation of systemic proteostasis allows the nervous system to monitor environmental variables and more effectively partition finite energy resources between different organismal processes. Our studies add to a growing body of work demonstrating that intercellular communication between neuronal and non-neuronal cells plays a critical role in integrating cellular stress resistance with other organismal physiological demands and associated energy costs. mRNA expression profiling of synchronized L4 stage wild-type N2 Bristol and VC1262 osm-9(ok1677) C. elegans strains under control (51mM NaCl) and hypertonic stress (200mM NaCl).

Project description:A detailed knowledge of the mechanisms underlying brain aging is fundamental to understand its functional decline and the baseline upon which brain pathologies superimpose. Endogenous protective mechanisms must contribute to the adaptability and plasticity still present in the healthy aged brain. Apolipoprotein D (ApoD) is one of the few genes with a consistent and evolutionarily conserved up-regulation in the aged brain. ApoD protecting roles upon stress or injury are well known, but a study of the effects of ApoD expression in the normal aging process is still missing. Using an ApoD-knockout mouse we analyze the effects of ApoD on factors contributing to the functional maintenance of the aged brain. We focused our cellular and molecular analyses in cortex and hippocampus at an age representing the onset of senescence where mortality risks are below 25%, avoiding bias towards long-lived animals. Lack of ApoD causes a prematurely aged brain without altering lifespan. Age-dependent hyperkinesia and memory deficits are accompanied by differential molecular effects in cortex and hippocampus. Transcriptome analyses reveal distinct effects of ApoD loss on the molecular age-dependent patterns of cortex and hippocampus, with different cell-type contributions to age-regulated gene expression. Markers of glial reactivity, proteostasis, and oxidative and inflammatory damage reveal early signs of aging and enhanced brain deterioration in the ApoD-knockout brain. The lack of ApoD results in an age-enhanced significant reduction in neuronal calcium-dependent functionality markers and signs of early reduction of neuronal numbers in the cortex, thus impinging upon parameters clearly differentiating neurodegenerative conditions from healthy brain aging. Our data support the hypothesis that the physiological increased brain expression of ApoD represents a homeostatic anti-aging mechanism. The brain cortex and hippocampus of young and aged mice of wild-type and ApoD-KO genotypes were used for RNA extraction and hybridization on Affymetrix microarrays. We aim at identifying distinct effects of ApoD loss on the molecular age-dependent patterns of cortex and hippocampus.

Dataset Information

Silencing of eIF5B Changes Proteostasis by Providing Negative Feedback on MAPK Signaling

Publications

Proteomics data in support of the quantification of the changes of bovine milk proteins during mammary gland involution.

Similar Datasets

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