Browsing by Author "Young, Sara K."

Now showing 1 - 7 of 7
  • Thumbnail Image
    Item
    Bmi1 promotes erythroid development through regulating ribosome biogenesis
    (Wiley, 2015-03) Gao, Rui; Chen, Sisi; Kobayashi, Michihiro; Yu, Hao; Zhang, Yingchi; Wan, Yang; Young, Sara K.; Soltis, Anthony; Yu, Ming; Vemula, Sasidhar; Fraenkel, Ernest; Cantor, Alan; Antipin, Yevgeniy; Xu, Yang; Yoder, Mervin C.; Wek, Ronald C.; Ellis, Steven R.; Kapur, Reuben; Zhu, Xiaofan; Liu, Yan; Department of Pediatrics, Indiana University School of Medicine
    While Polycomb group protein Bmi1 is important for stem cell maintenance, its role in lineage commitment is largely unknown. We have identified Bmi1 as a novel regulator of erythroid development. Bmi1 is highly expressed in mouse erythroid progenitor cells and its deficiency impairs erythroid differentiation. BMI1 is also important for human erythroid development. Furthermore, we discovered that loss of Bmi1 in erythroid progenitor cells results in decreased transcription of multiple ribosomal protein genes and impaired ribosome biogenesis. Bmi1 deficiency stabilizes p53 protein, leading to upregulation of p21 expression and subsequent G0/G1 cell cycle arrest. Genetic inhibition of p53 activity rescues the erythroid defects seen in the Bmi1 null mice, demonstrating that a p53-dependent mechanism underlies the pathophysiology of the anemia. Mechanistically, Bmi1 is associated with multiple ribosomal protein genes and may positively regulate their expression in erythroid progenitor cells. Thus, Bmi1 promotes erythroid development, at least in part through regulating ribosome biogenesis. Ribosomopathies are human disorders of ribosome dysfunction, including Diamond-Blackfan anemia (DBA) and 5q- syndrome, in which genetic abnormalities cause impaired ribosome biogenesis, resulting in specific clinical phenotypes. We observed that BMI1 expression in human hematopoietic stem and progenitor cells from patients with DBA is correlated with the expression of some ribosomal protein genes, suggesting that BMI1 deficiency may play a pathological role in DBA and other ribosomopathies.
  • Thumbnail Image
    Item
    CHOP links endoplasmic reticulum stress to NF-κB activation in the pathogenesis of nonalcoholic steatohepatitis
    (2015) Willy, Jeffrey A.; Young, Sara K.; Stevens, James L.; Masuoka, Howard C.; Wek, Ronald C.; Department of Biochemistry and Molecular Biology, IU School of Medicine
    Free fatty acid induction of inflammation and cell death is an important feature of nonalcoholic steatohepatitis (NASH) and has been associated with disruption of the endoplasmic reticulum and activation of the Unfolded Protein Response (UPR). Following chronic UPR activation, the transcription factor CHOP (GADD153/DDIT3) triggers cell death; however, the mechanisms linking the UPR or CHOP to hepatoceullular injury and inflammation in the pathogenesis of NASH are not well understood. Using HepG2 and primary human hepatocytes, we found that CHOP induces cell death and inflammatory responses following saturated free fatty acid exposure by activating NF-κB through a pathway involving IRAK2 expression, resulting in secretion of cytokines IL-8 and TNFα directly from hepatocytes. TNFα facilitates hepatocyte death upon exposure to saturated free fatty acids and secretion of both IL-8 and TNFα contribute to inflammation. Interestingly, CHOP/NF-κB signaling is not conserved in primary rodent hepatocytes. Our studies suggest that CHOP plays a vital role in the pathophysiology of NASH through induction of secreted factors that trigger inflammation and hepatocellular death via a signaling pathway specific to human hepatocytes.
  • Thumbnail Image
    Item
    Function of inhibitor of Bruton's tyrosine kinase isoform α (IBTKα) in nonalcoholic steatohepatitis links autophagy and the unfolded protein response
    (American Society for Biochemistry and Molecular Biology, 2017-08-25) Willy, Jeffrey A.; Young, Sara K.; Mosley, Amber L.; Gawrieh, Samer; Stevens, James L.; Masuoka, Howard C.; Wek, Ronald C.; Biochemistry and Molecular Biology, School of Medicine
    Nonalcoholic fatty liver disease (steatosis) is the most prevalent liver disease in the Western world. One of the advanced pathologies is nonalcoholic steatohepatitis (NASH), which is associated with induction of the unfolded protein response (UPR) and disruption of autophagic flux. However, the mechanisms by which these processes contribute to the pathogenesis of human diseases are unclear. Herein, we identify the α isoform of the inhibitor of Bruton's tyrosine kinase (IBTKα) as a member of the UPR, whose expression is preferentially translated during endoplasmic reticulum (ER) stress. We found that IBTKα is located in the ER and associates with proteins LC3b, SEC16A, and SEC31A and plays a previously unrecognized role in phagophore initiation from ER exit sites. Depletion of IBTKα helps prevent accumulation of autophagosome intermediates stemming from exposure to saturated free fatty acids and rescues hepatocytes from death. Of note, induction of IBTKα and the UPR, along with inhibition of autophagic flux, was associated with progression from steatosis to NASH in liver biopsies. These results indicate a function for IBTKα in NASH that links autophagy with activation of the UPR.
  • Thumbnail Image
    Item
    Nuclear Matrix Protein 4 Is a Novel Regulator of Ribosome Biogenesis and Controls the Unfolded Protein Response via Repression of Gadd34 Expression
    (American Society for Biochemistry and Molecular Biology, 2016-06-24) Young, Sara K.; Shao, Yu; Bidwell, Joseph P.; Wek, Ronald C.; Biochemistry and Molecular Biology, School of Medicine
    The unfolded protein response (UPR) maintains protein homeostasis by governing the processing capacity of the endoplasmic reticulum (ER) to manage ER client loads; however, key regulators within the UPR remain to be identified. Activation of the UPR sensor PERK (EIFAK3/PEK) results in the phosphorylation of the α subunit of eIF2 (eIF2α-P), which represses translation initiation and reduces influx of newly synthesized proteins into the overloaded ER. As part of this adaptive response, eIF2α-P also induces a feedback mechanism through enhanced transcriptional and translational expression of Gadd34 (Ppp1r15A),which targets type 1 protein phosphatase for dephosphorylation of eIF2α-P to restore protein synthesis. Here we describe a novel mechanism by which Gadd34 expression is regulated through the activity of the zinc finger transcription factor NMP4 (ZNF384, CIZ). NMP4 functions to suppress bone anabolism, and we suggest that this occurs due to decreased protein synthesis of factors involved in bone formation through NMP4-mediated dampening of Gadd34 and c-Myc expression. Loss of Nmp4 resulted in an increase in c-Myc and Gadd34 expression that facilitated enhanced ribosome biogenesis and global protein synthesis. Importantly, protein synthesis was sustained during pharmacological induction of the UPR through a mechanism suggested to involve GADD34-mediated dephosphorylation of eIF2α-P. Sustained protein synthesis sensitized cells to pharmacological induction of the UPR, and the observed decrease in cell viability was restored upon inhibition of GADD34 activity. We conclude that NMP4 is a key regulator of ribosome biogenesis and the UPR, which together play a central role in determining cell viability during endoplasmic reticulum stress.
  • Thumbnail Image
    Item
    Ribosome Elongation Stall Directs Gene-specific Translation in the Integrated Stress Response
    (American Society for Biochemistry and Molecular Biology, 2016-03-18) Young, Sara K.; Palam, Lakshmi Reddy; Wu, Cheng; Sachs, Matthew S.; Wek, Ronald C.; Department of Biochemistry & Molecular Biology, IU School of Medicine
    Upon exposure to environmental stress, phosphorylation of the α subunit of eIF2 (eIF2α-P) represses global protein synthesis, coincident with preferential translation of gene transcripts that mitigate stress damage or alternatively trigger apoptosis. Because there are multiple mammalian eIF2 kinases, each responding to different stress arrangements, this translational control scheme is referred to as the integrated stress response (ISR). Included among the preferentially translated mRNAs induced by eIF2α-P is that encoding the transcription factor CHOP (DDIT3/GADD153). Enhanced levels of CHOP promote cell death when ISR signaling is insufficient to restore cell homeostasis. Preferential translation of CHOP mRNA occurs by a mechanism involving ribosome bypass of an inhibitory upstream ORF (uORF) situated in the 5'-leader of the CHOP mRNA. In this study, we used biochemical and genetic approaches to define the inhibitory features of the CHOP uORF and the biological consequences of loss of the CHOP uORF on CHOP expression during stress. We discovered that specific sequences within the CHOP uORF serve to stall elongating ribosomes and prevent ribosome reinitiation at the downstream CHOP coding sequence. As a consequence, deletion of the CHOP uORF substantially increases the levels and modifies the pattern of induction of CHOP expression in the ISR. Enhanced CHOP expression leads to increased expression of key CHOP target genes, culminating in increased cell death in response to stress.
  • Thumbnail Image
    Item
    Ribosome Reinitiation Directs Gene-specific Translation and Regulates the Integrated Stress Response
    (American Society for Biochemistry and Molecular Biology, 2015-11-20) Young, Sara K.; Willy, Jeffrey A.; Wu, Cheng; Sachs, Matthew S.; Wek, Ronald C.; Department of Biochemistry & Molecular Biology, IU School of Medicine
    Young, S. K., Willy, J. A., Wu, C., Sachs, M. S., & Wek, R. C. (2015). Ribosome Reinitiation Directs Gene-specific Translation and Regulates the Integrated Stress Response. The Journal of Biological Chemistry, 290(47), 28257–28271. http://doi.org/10.1074/jbc.M115.693184
  • Thumbnail Image
    Item
    Translation Regulation of the Glutamyl-prolyl-tRNA Synthetase Gene EPRS through Bypass of Upstream Open Reading Frames with Noncanonical Initiation Codons
    (American Society for Biochemistry and Molecular Biology, 2016-05-13) Young, Sara K.; Baird, Thomas D.; Wek, Ronald C.; Department of Biochemistry and Molecular Biology, School of Medicine
    In the integrated stress response, phosphorylation of eIF2α (eIF2α-P) reduces protein synthesis while concomitantly promoting preferential translation of specific transcripts associated with stress adaptation. Translation of the glutamyl-prolyl-tRNA synthetase gene EPRS is enhanced in response to eIF2α-P. To identify the underlying mechanism of translation control, we employed biochemical approaches to determine the regulatory features by which upstream ORFs (uORFs) direct downstream translation control and expression of the EPRS coding region. Our findings reveal that translation of two inhibitory uORFs encoded by noncanonical CUG and UUG initiation codons in the EPRS mRNA 5'-leader serve to dampen levels of translation initiation at the EPRS coding region. By a mechanism suggested to involve increased translation initiation stringency during stress-induced eIF2α-P, we observed facilitated ribosome bypass of these uORFs, allowing for increased translation of the EPRS coding region. Importantly, EPRS protein expression is enhanced through this preferential translation mechanism in response to multiple known activators of eIF2α-P and likely serves to facilitate stress adaptation in response to a variety of cellular stresses. The rules presented here for the regulated ribosome bypass of noncanonical initiation codons in the EPRS 5'-leader add complexity into the nature of uORF-mediated translation control mechanisms during eIF2α-P and additionally illustrate the roles that previously unexamined uORFs with noncanonical initiation codons can play in modulating gene expression.