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Assembly-induced folding regulates interleukin 12 biogenesis and secretion.

ABSTRACT: Members of the IL-12 family perform essential functions in immunoregulation by connecting innate and adaptive immunity and are emerging therapeutic targets. They are unique among other interleukins in forming heterodimers that arise from extensive subunit sharing within the family, leading to the production of at least four functionally distinct heterodimers from only five subunits. This raises important questions about how the assembly of IL-12 family members is regulated and controlled in the cell. Here, using cell-biological approaches, we have dissected basic principles that underlie the biogenesis of the founding member of the family, IL-12. Within the native IL-12 heterodimer, composed of IL-12? and IL-12?, IL-12? possesses three intramolecular and one intermolecular disulfide bridges. We show that, in isolation, IL-12? fails to form its native structure but, instead, misfolds, forming incorrect disulfide bonds. Co-expression of its ? subunit inhibits misfolding and thus allows secretion of biologically active heterodimeric IL-12. On the basis of these findings, we identified the disulfide bonds in IL-12? that are critical for assembly-induced secretion and biological activity of IL-12 versus misfolding and degradation of IL-12?. Surprisingly, two of the three disulfide bridges in IL-12? are dispensable for IL-12 secretion, stability, and biological activity. Extending our findings, we show that misfolding also occurs for IL-23?, another IL-12 family protein. Our results indicate that assembly-induced folding is key in IL-12 family biogenesis and secretion. The identification of essential disulfide bonds that underlie this process lays the basis for a simplified yet functional IL-12 cytokine.

SUBMITTER: Reitberger S

PROVIDER: S-EPMC5427282 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

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Assembly-induced folding regulates interleukin 12 biogenesis and secretion.

Reitberger Susanne S Haimerl Pascal P Aschenbrenner Isabel I Esser-von Bieren Julia J Feige Matthias J MJ

The Journal of biological chemistry 20170321 19

Members of the IL-12 family perform essential functions in immunoregulation by connecting innate and adaptive immunity and are emerging therapeutic targets. They are unique among other interleukins in forming heterodimers that arise from extensive subunit sharing within the family, leading to the production of at least four functionally distinct heterodimers from only five subunits. This raises important questions about how the assembly of IL-12 family members is regulated and controlled in the ...[more]

PMID: 28325840

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Project description:ObjectivePhysical exercise training is associated with increased glucose uptake in skeletal muscle and improved glycemic control. HDAC5, a class IIa histone deacetylase, has been shown to regulate transcription of the insulin-responsive glucose transporter GLUT4 in cultured muscle cells. In this study, we analyzed the contribution of HDAC5 to the transcriptional network in muscle and the beneficial effect of muscle contraction and regular exercise on glucose metabolism.MethodsHDAC5 knockout mice (KO) and wild-type (WT) littermates were trained for 8 weeks on treadmills, metabolically phenotyped, and compared to sedentary controls. Hdac5-deficient skeletal muscle and cultured Hdac5-knockdown (KD) C2C12 myotubes were utilized for studies of gene expression and glucose metabolism. Chromatin immunoprecipitation (ChIP) studies were conducted to analyze Il6 promoter activity using H3K9ac and HDAC5 antibodies.ResultsGlobal transcriptome analysis of Hdac5 KO gastrocnemius muscle demonstrated activation of the IL-6 signaling pathway. Accordingly, knockdown of Hdac5 in C2C12 myotubes led to higher expression and secretion of IL-6 with enhanced insulin-stimulated activation of AKT that was reversed by Il6 knockdown. Moreover, Hdac5-deficient myotubes exhibited enhanced glucose uptake, glycogen synthesis, and elevated expression levels of the glucose transporter GLUT4. Transcription of Il6 was further enhanced by electrical pulse stimulation in Hdac5-deficient C2C12 myotubes. ChIP identified a ∼1 kb fragment of the Il6 promoter that interacts with HDAC5 and demonstrated increased activation-associated histone marker AcH3K9 in Hdac5-deficient muscle cells. Exercise intervention of HDAC5 KO mice resulted in improved systemic glucose tolerance as compared to WT controls.ConclusionsWe identified HDAC5 as a negative epigenetic regulator of IL-6 synthesis and release in skeletal muscle. HDAC5 may exert beneficial effects through two different mechanisms, transcriptional control of genes required for glucose disposal and utilization, and HDAC5-dependent IL-6 signaling cross-talk to improve glucose uptake in muscle in response to exercise.

Dataset Information

Assembly-induced folding regulates interleukin 12 biogenesis and secretion.

Publications

Assembly-induced folding regulates interleukin 12 biogenesis and secretion.

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