Modulating serine palmitoyl transferase (SPT) expression and activity unveils a crucial role in lipid-induced insulin resistance in rat skeletal muscle cells

Maria L. Watson, Matthew Coghlan, Harinder S. Hundal (Lead / Corresponding author)

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    61 Citations (Scopus)

    Abstract

    Saturated fatty acids, such as palmitate, promote accumulation of ceramide, which impairs activation and signalling of PKB (protein kinase B; also known as Akt) to important end points such as glucose transport. SPT (serine palmitoyl transferase) is a key enzyme regulating ceramide synthesis from palmitate and represents a potential molecular target in curbing lipid-induced insulin resistance. In the present Study we explore the effects of palmitate upon insulin action in L6 muscle cells in which SPT expression/activity has been decreased by shRNA (small-hairpin RNA) or Sustained incubation with myriocin, an SPT inhibitor. Incubation of L6 myotubes with palmitate (for 16 h) increases intramyocellular ceramide and reduces insulin-stimulated PKB activation and glucose uptake. PKB inhibition was not associated with impaired IRS (insulin receptor substrate) signalling and was ameliorated by short-term treatment with myriocin. Silencing SPT expression (similar to 90%) by shRNA or chronic cell incubation with myriocin (for 7 days) markedly suppressed SPT activity and palmitate-driven ceramide synthesis; however, challenging these muscle cells with palmitate still inhibited the hormonal activation of PKB. This inhibition was associated with reduced IRS1/p85-PI3K (phosphoinositide 3-kinase) coupling that arises from diverting palmitate towards greater DAG (diacylglycerol) synthesis, which elevates IRS1 serine phosphorylation via activation of DAG-sensitive PKCs (protein kinase Cs). Treatment of SPT-shRNA cells or those treated chronically with myriocin with PKC inhibitors antagonized palmitate-induced loss in insulin signalling. The findings of the present study indicate that SPT plays a crucial role in desensitizing muscle cells to insulin in response to incubation with palmitate. While short-term inhibition of SPT ameliorates palmitate/ceramide-induced insulin resistance, Sustained loss/reduction in SPT expression/activity promotes greater partitioning of palmitate towards DAG synthesis, which impacts negatively upon IRS1-directed insulin signalling.

    Original languageEnglish
    Pages (from-to)791-801
    Number of pages11
    JournalBiochemical Journal
    Volume417
    Issue number1
    DOIs
    Publication statusPublished - 1 Feb 2009

    Keywords

    • ceramide
    • diacylglycerol
    • insulin receptor substrate 1 (IRS1)
    • myriocin
    • palmitate
    • protein kinase B (PKB)
    • protein kinase C (PKC)
    • PROTEIN-KINASE-C
    • PLECKSTRIN HOMOLOGY DOMAIN
    • FATTY-ACIDS
    • RECEPTOR SUBSTRATE-1
    • DEPENDENT MECHANISM
    • CERAMIDE SYNTHESIS
    • 3T3-L1 ADIPOCYTES
    • KAPPA-B
    • ACTIVATION
    • INHIBITION

    Cite this

    @article{235e3ecab97c482289387fa0a85c759b,
    title = "Modulating serine palmitoyl transferase (SPT) expression and activity unveils a crucial role in lipid-induced insulin resistance in rat skeletal muscle cells",
    abstract = "Saturated fatty acids, such as palmitate, promote accumulation of ceramide, which impairs activation and signalling of PKB (protein kinase B; also known as Akt) to important end points such as glucose transport. SPT (serine palmitoyl transferase) is a key enzyme regulating ceramide synthesis from palmitate and represents a potential molecular target in curbing lipid-induced insulin resistance. In the present Study we explore the effects of palmitate upon insulin action in L6 muscle cells in which SPT expression/activity has been decreased by shRNA (small-hairpin RNA) or Sustained incubation with myriocin, an SPT inhibitor. Incubation of L6 myotubes with palmitate (for 16 h) increases intramyocellular ceramide and reduces insulin-stimulated PKB activation and glucose uptake. PKB inhibition was not associated with impaired IRS (insulin receptor substrate) signalling and was ameliorated by short-term treatment with myriocin. Silencing SPT expression (similar to 90{\%}) by shRNA or chronic cell incubation with myriocin (for 7 days) markedly suppressed SPT activity and palmitate-driven ceramide synthesis; however, challenging these muscle cells with palmitate still inhibited the hormonal activation of PKB. This inhibition was associated with reduced IRS1/p85-PI3K (phosphoinositide 3-kinase) coupling that arises from diverting palmitate towards greater DAG (diacylglycerol) synthesis, which elevates IRS1 serine phosphorylation via activation of DAG-sensitive PKCs (protein kinase Cs). Treatment of SPT-shRNA cells or those treated chronically with myriocin with PKC inhibitors antagonized palmitate-induced loss in insulin signalling. The findings of the present study indicate that SPT plays a crucial role in desensitizing muscle cells to insulin in response to incubation with palmitate. While short-term inhibition of SPT ameliorates palmitate/ceramide-induced insulin resistance, Sustained loss/reduction in SPT expression/activity promotes greater partitioning of palmitate towards DAG synthesis, which impacts negatively upon IRS1-directed insulin signalling.",
    keywords = "ceramide, diacylglycerol, insulin receptor substrate 1 (IRS1), myriocin, palmitate, protein kinase B (PKB), protein kinase C (PKC), PROTEIN-KINASE-C, PLECKSTRIN HOMOLOGY DOMAIN, FATTY-ACIDS, RECEPTOR SUBSTRATE-1, DEPENDENT MECHANISM, CERAMIDE SYNTHESIS, 3T3-L1 ADIPOCYTES, KAPPA-B, ACTIVATION, INHIBITION",
    author = "Watson, {Maria L.} and Matthew Coghlan and Hundal, {Harinder S.}",
    year = "2009",
    month = "2",
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    doi = "10.1042/BJ20081149",
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    volume = "417",
    pages = "791--801",
    journal = "Biochemical Journal",
    issn = "0264-6021",
    publisher = "Portland Press",
    number = "1",

    }

    TY - JOUR

    T1 - Modulating serine palmitoyl transferase (SPT) expression and activity unveils a crucial role in lipid-induced insulin resistance in rat skeletal muscle cells

    AU - Watson, Maria L.

    AU - Coghlan, Matthew

    AU - Hundal, Harinder S.

    PY - 2009/2/1

    Y1 - 2009/2/1

    N2 - Saturated fatty acids, such as palmitate, promote accumulation of ceramide, which impairs activation and signalling of PKB (protein kinase B; also known as Akt) to important end points such as glucose transport. SPT (serine palmitoyl transferase) is a key enzyme regulating ceramide synthesis from palmitate and represents a potential molecular target in curbing lipid-induced insulin resistance. In the present Study we explore the effects of palmitate upon insulin action in L6 muscle cells in which SPT expression/activity has been decreased by shRNA (small-hairpin RNA) or Sustained incubation with myriocin, an SPT inhibitor. Incubation of L6 myotubes with palmitate (for 16 h) increases intramyocellular ceramide and reduces insulin-stimulated PKB activation and glucose uptake. PKB inhibition was not associated with impaired IRS (insulin receptor substrate) signalling and was ameliorated by short-term treatment with myriocin. Silencing SPT expression (similar to 90%) by shRNA or chronic cell incubation with myriocin (for 7 days) markedly suppressed SPT activity and palmitate-driven ceramide synthesis; however, challenging these muscle cells with palmitate still inhibited the hormonal activation of PKB. This inhibition was associated with reduced IRS1/p85-PI3K (phosphoinositide 3-kinase) coupling that arises from diverting palmitate towards greater DAG (diacylglycerol) synthesis, which elevates IRS1 serine phosphorylation via activation of DAG-sensitive PKCs (protein kinase Cs). Treatment of SPT-shRNA cells or those treated chronically with myriocin with PKC inhibitors antagonized palmitate-induced loss in insulin signalling. The findings of the present study indicate that SPT plays a crucial role in desensitizing muscle cells to insulin in response to incubation with palmitate. While short-term inhibition of SPT ameliorates palmitate/ceramide-induced insulin resistance, Sustained loss/reduction in SPT expression/activity promotes greater partitioning of palmitate towards DAG synthesis, which impacts negatively upon IRS1-directed insulin signalling.

    AB - Saturated fatty acids, such as palmitate, promote accumulation of ceramide, which impairs activation and signalling of PKB (protein kinase B; also known as Akt) to important end points such as glucose transport. SPT (serine palmitoyl transferase) is a key enzyme regulating ceramide synthesis from palmitate and represents a potential molecular target in curbing lipid-induced insulin resistance. In the present Study we explore the effects of palmitate upon insulin action in L6 muscle cells in which SPT expression/activity has been decreased by shRNA (small-hairpin RNA) or Sustained incubation with myriocin, an SPT inhibitor. Incubation of L6 myotubes with palmitate (for 16 h) increases intramyocellular ceramide and reduces insulin-stimulated PKB activation and glucose uptake. PKB inhibition was not associated with impaired IRS (insulin receptor substrate) signalling and was ameliorated by short-term treatment with myriocin. Silencing SPT expression (similar to 90%) by shRNA or chronic cell incubation with myriocin (for 7 days) markedly suppressed SPT activity and palmitate-driven ceramide synthesis; however, challenging these muscle cells with palmitate still inhibited the hormonal activation of PKB. This inhibition was associated with reduced IRS1/p85-PI3K (phosphoinositide 3-kinase) coupling that arises from diverting palmitate towards greater DAG (diacylglycerol) synthesis, which elevates IRS1 serine phosphorylation via activation of DAG-sensitive PKCs (protein kinase Cs). Treatment of SPT-shRNA cells or those treated chronically with myriocin with PKC inhibitors antagonized palmitate-induced loss in insulin signalling. The findings of the present study indicate that SPT plays a crucial role in desensitizing muscle cells to insulin in response to incubation with palmitate. While short-term inhibition of SPT ameliorates palmitate/ceramide-induced insulin resistance, Sustained loss/reduction in SPT expression/activity promotes greater partitioning of palmitate towards DAG synthesis, which impacts negatively upon IRS1-directed insulin signalling.

    KW - ceramide

    KW - diacylglycerol

    KW - insulin receptor substrate 1 (IRS1)

    KW - myriocin

    KW - palmitate

    KW - protein kinase B (PKB)

    KW - protein kinase C (PKC)

    KW - PROTEIN-KINASE-C

    KW - PLECKSTRIN HOMOLOGY DOMAIN

    KW - FATTY-ACIDS

    KW - RECEPTOR SUBSTRATE-1

    KW - DEPENDENT MECHANISM

    KW - CERAMIDE SYNTHESIS

    KW - 3T3-L1 ADIPOCYTES

    KW - KAPPA-B

    KW - ACTIVATION

    KW - INHIBITION

    UR - http://www.scopus.com/inward/record.url?scp=59849098608&partnerID=8YFLogxK

    U2 - 10.1042/BJ20081149

    DO - 10.1042/BJ20081149

    M3 - Article

    C2 - 18922131

    VL - 417

    SP - 791

    EP - 801

    JO - Biochemical Journal

    JF - Biochemical Journal

    SN - 0264-6021

    IS - 1

    ER -