Transcriptome of the GSH-Depleted lens reveals changes in detoxification and EMT signaling genes, transport systems, and lipid homeostasis

Jeremy A. Whitson, Xiang Zhang, Mario Medvedovic, Jenny Chen, Zongbo Wei, Vincent M. Monnier, Xingjun Fan

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

PURPOSE. To understand the effects of glutathione (GSH)-deficiency on genetic processes that regulate lens homeostasis and prevent cataractogenesis. METHODS. The transcriptome of lens epithelia and fiber cells was obtained from C57BL/6 LEGSKO (lens GSH-synthesis knockout) and buthionine sulfoximine (BSO)-treated LEGSKO mice and compared to C57BL/6 wild-type mice using RNA-Seq. Transcriptomic data were confirmed by qPCR and Western blot/ELISA on a subset of genes. RESULTS. RNA-Seq results were in excellent agreement with qPCR (correlation coefficients 0.87-0.94 and P < 5E-6 for a subset of 36 mRNAs). Of 24,415 transcripts mapped to the mouse genome, 441 genes showed significantly modulated expression. Pathway analysis indicated major changes in epithelial-mesenchymal transition (EMT) signaling, visual cycle, small molecule biochemistry, and lipid metabolism. GSH-deficient lenses showed upregulation of detoxification genes, including Aldh1a1, Aldh3a1 (aldehyde dehydrogenases), Mt1, Mt2 (metallothioneins), Ces1g (carboxylesterase), and Slc14a1 (urea transporter UT-B). Genes in canonical EMT pathways, including Wnt10a, showed upregulation in lens epithelia samples. Severely GSH-deficient lens epithelia showed downregulation of vision-related genes (including crystallins). The BSO-treated LEGSKO lens epithelia transcriptome has significant correlation (r = 0.63, P < 0.005) to that of lens epithelia undergoing EMT. Protein expression data correlated with transcriptomic data and confirmed EMT signaling activation. CONCLUSIONS. These results show that GSH-deficiency in the lens leads to expression of detoxifying genes and activation of EMT signaling, in addition to changes in transport systems and lipid homeostasis. These data provide insight into the adaptation and consequences of GSH-deficiency in the lens and suggest that GSH plays an important role in lenticular EMT pathology.

Original languageEnglish (US)
Pages (from-to)2666-2684
Number of pages19
JournalInvestigative Ophthalmology and Visual Science
Volume58
Issue number5
DOIs
StatePublished - May 1 2017

Fingerprint

Epithelial-Mesenchymal Transition
Transcriptome
Lenses
Homeostasis
Lipids
Genes
Epithelium
Buthionine Sulfoximine
Up-Regulation
Genetic Phenomena
RNA
Carboxylesterase
Aldehyde Dehydrogenase
Crystallins
Metallothionein
Lipid Metabolism
Knockout Mice
Biochemistry
Transcriptional Activation
Glutathione

Keywords

  • Cataract
  • Epithelial mesenchymal transition
  • Gene expression
  • Glutathione
  • LEGSKO mouse
  • Oxidative stress
  • RNA-seq

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

Cite this

Transcriptome of the GSH-Depleted lens reveals changes in detoxification and EMT signaling genes, transport systems, and lipid homeostasis. / Whitson, Jeremy A.; Zhang, Xiang; Medvedovic, Mario; Chen, Jenny; Wei, Zongbo; Monnier, Vincent M.; Fan, Xingjun.

In: Investigative Ophthalmology and Visual Science, Vol. 58, No. 5, 01.05.2017, p. 2666-2684.

Research output: Contribution to journalArticle

Whitson, Jeremy A. ; Zhang, Xiang ; Medvedovic, Mario ; Chen, Jenny ; Wei, Zongbo ; Monnier, Vincent M. ; Fan, Xingjun. / Transcriptome of the GSH-Depleted lens reveals changes in detoxification and EMT signaling genes, transport systems, and lipid homeostasis. In: Investigative Ophthalmology and Visual Science. 2017 ; Vol. 58, No. 5. pp. 2666-2684.
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abstract = "PURPOSE. To understand the effects of glutathione (GSH)-deficiency on genetic processes that regulate lens homeostasis and prevent cataractogenesis. METHODS. The transcriptome of lens epithelia and fiber cells was obtained from C57BL/6 LEGSKO (lens GSH-synthesis knockout) and buthionine sulfoximine (BSO)-treated LEGSKO mice and compared to C57BL/6 wild-type mice using RNA-Seq. Transcriptomic data were confirmed by qPCR and Western blot/ELISA on a subset of genes. RESULTS. RNA-Seq results were in excellent agreement with qPCR (correlation coefficients 0.87-0.94 and P < 5E-6 for a subset of 36 mRNAs). Of 24,415 transcripts mapped to the mouse genome, 441 genes showed significantly modulated expression. Pathway analysis indicated major changes in epithelial-mesenchymal transition (EMT) signaling, visual cycle, small molecule biochemistry, and lipid metabolism. GSH-deficient lenses showed upregulation of detoxification genes, including Aldh1a1, Aldh3a1 (aldehyde dehydrogenases), Mt1, Mt2 (metallothioneins), Ces1g (carboxylesterase), and Slc14a1 (urea transporter UT-B). Genes in canonical EMT pathways, including Wnt10a, showed upregulation in lens epithelia samples. Severely GSH-deficient lens epithelia showed downregulation of vision-related genes (including crystallins). The BSO-treated LEGSKO lens epithelia transcriptome has significant correlation (r = 0.63, P < 0.005) to that of lens epithelia undergoing EMT. Protein expression data correlated with transcriptomic data and confirmed EMT signaling activation. CONCLUSIONS. These results show that GSH-deficiency in the lens leads to expression of detoxifying genes and activation of EMT signaling, in addition to changes in transport systems and lipid homeostasis. These data provide insight into the adaptation and consequences of GSH-deficiency in the lens and suggest that GSH plays an important role in lenticular EMT pathology.",
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T1 - Transcriptome of the GSH-Depleted lens reveals changes in detoxification and EMT signaling genes, transport systems, and lipid homeostasis

AU - Whitson, Jeremy A.

AU - Zhang, Xiang

AU - Medvedovic, Mario

AU - Chen, Jenny

AU - Wei, Zongbo

AU - Monnier, Vincent M.

AU - Fan, Xingjun

PY - 2017/5/1

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N2 - PURPOSE. To understand the effects of glutathione (GSH)-deficiency on genetic processes that regulate lens homeostasis and prevent cataractogenesis. METHODS. The transcriptome of lens epithelia and fiber cells was obtained from C57BL/6 LEGSKO (lens GSH-synthesis knockout) and buthionine sulfoximine (BSO)-treated LEGSKO mice and compared to C57BL/6 wild-type mice using RNA-Seq. Transcriptomic data were confirmed by qPCR and Western blot/ELISA on a subset of genes. RESULTS. RNA-Seq results were in excellent agreement with qPCR (correlation coefficients 0.87-0.94 and P < 5E-6 for a subset of 36 mRNAs). Of 24,415 transcripts mapped to the mouse genome, 441 genes showed significantly modulated expression. Pathway analysis indicated major changes in epithelial-mesenchymal transition (EMT) signaling, visual cycle, small molecule biochemistry, and lipid metabolism. GSH-deficient lenses showed upregulation of detoxification genes, including Aldh1a1, Aldh3a1 (aldehyde dehydrogenases), Mt1, Mt2 (metallothioneins), Ces1g (carboxylesterase), and Slc14a1 (urea transporter UT-B). Genes in canonical EMT pathways, including Wnt10a, showed upregulation in lens epithelia samples. Severely GSH-deficient lens epithelia showed downregulation of vision-related genes (including crystallins). The BSO-treated LEGSKO lens epithelia transcriptome has significant correlation (r = 0.63, P < 0.005) to that of lens epithelia undergoing EMT. Protein expression data correlated with transcriptomic data and confirmed EMT signaling activation. CONCLUSIONS. These results show that GSH-deficiency in the lens leads to expression of detoxifying genes and activation of EMT signaling, in addition to changes in transport systems and lipid homeostasis. These data provide insight into the adaptation and consequences of GSH-deficiency in the lens and suggest that GSH plays an important role in lenticular EMT pathology.

AB - PURPOSE. To understand the effects of glutathione (GSH)-deficiency on genetic processes that regulate lens homeostasis and prevent cataractogenesis. METHODS. The transcriptome of lens epithelia and fiber cells was obtained from C57BL/6 LEGSKO (lens GSH-synthesis knockout) and buthionine sulfoximine (BSO)-treated LEGSKO mice and compared to C57BL/6 wild-type mice using RNA-Seq. Transcriptomic data were confirmed by qPCR and Western blot/ELISA on a subset of genes. RESULTS. RNA-Seq results were in excellent agreement with qPCR (correlation coefficients 0.87-0.94 and P < 5E-6 for a subset of 36 mRNAs). Of 24,415 transcripts mapped to the mouse genome, 441 genes showed significantly modulated expression. Pathway analysis indicated major changes in epithelial-mesenchymal transition (EMT) signaling, visual cycle, small molecule biochemistry, and lipid metabolism. GSH-deficient lenses showed upregulation of detoxification genes, including Aldh1a1, Aldh3a1 (aldehyde dehydrogenases), Mt1, Mt2 (metallothioneins), Ces1g (carboxylesterase), and Slc14a1 (urea transporter UT-B). Genes in canonical EMT pathways, including Wnt10a, showed upregulation in lens epithelia samples. Severely GSH-deficient lens epithelia showed downregulation of vision-related genes (including crystallins). The BSO-treated LEGSKO lens epithelia transcriptome has significant correlation (r = 0.63, P < 0.005) to that of lens epithelia undergoing EMT. Protein expression data correlated with transcriptomic data and confirmed EMT signaling activation. CONCLUSIONS. These results show that GSH-deficiency in the lens leads to expression of detoxifying genes and activation of EMT signaling, in addition to changes in transport systems and lipid homeostasis. These data provide insight into the adaptation and consequences of GSH-deficiency in the lens and suggest that GSH plays an important role in lenticular EMT pathology.

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KW - Epithelial mesenchymal transition

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KW - LEGSKO mouse

KW - Oxidative stress

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