Brain-selective Kinase 2 (BRSK2) Phosphorylation on PCTAIRE1 Negatively Regulates Glucose-stimulated Insulin Secretion in Pancreatic -Cells

State Key Laboratory of Genetic Engineering, School of Life Science, Institutes of Biomedical Sciences, Fudan University, Shanghai, 200433, China.
Journal of Biological Chemistry (Impact Factor: 4.57). 07/2012; 287(36):30368-75. DOI: 10.1074/jbc.M112.375618
Source: PubMed


Brain-selective kinase 2 (BRSK2) has been shown to play an essential role in neuronal polarization. In the present study, we show that BRSK2 is also abundantly expressed in pancreatic islets and MIN6 β-cell line. Yeast two-hybrid screening, GST fusion protein pull-down, and co-immunoprecipitation assays reveal that BRSK2 interacts with CDK-related protein kinase PCTAIRE1, a kinase involved in neurite outgrowth and neurotransmitter release. In MIN6 cells, BRSK2 co-localizes with PCTAIRE1 in the cytoplasm and phosphorylates one of its serine residues, Ser-12. Phosphorylation of PCTAIRE1 by BRSK2 reduces glucose-stimulated insulin secretion (GSIS) in MIN6 cells. Conversely, knockdown of BRSK2 by siRNA increases serum insulin levels in mice. Our results reveal a novel function of BRSK2 in the regulation of GSIS in β-cells via a PCTAIRE1-dependent mechanism and suggest that BRSK2 is an attractive target for developing novel diabetic drugs.

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    • "24 h post-transfection, cells were either left untreated or were treated with Tm, Tg or normal medium for 24 h. Cells were then fixed with 4 % (w/v) paraformaldehyde, probed with rabbit BRSK2 polyclonal antibodies [either produced in our laboratory (Chen et al. 2012) or a VCP monoclonal antibody (BD Biosciences), a fluorescein-conjugated goat anti-rabbit secondary antibody (Alexa 488) or a goat anti-mouse secondary antibody (Alexa 546) (Invitrogen)]. Nuclei were visualized by incubating cells with 0.1 mg DAPI/ml (Sigma) in the dark for 10 min at room temperature. "
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    ABSTRACT: Endoplasmic reticulum-associated protein degradation (ERAD) removes improperly-folded proteins from the ER membrane into the cytosol where they undergo proteasomal degradation. Valosin-containing protein (VCP)/p97 mediates in the extraction of ERAD substrates from the ER. BRSK2 (also known as SAD-A), a serine/threonine kinase of the AMP-activated protein kinase family affected VCP/p97 activity in ERAD. In addition, BRSK2 interacted with VCP/p97 via three of the four functional domains of VCP/p97. Immunofluorescence demonstrated that BRSK2 and VCP/p97 were co-localized and also that knockdown of endogenous BRSK2 induced increased levels of CD3δ, a substrate in ERAD for VCP/p97. Thus, BRSK2 might affect the activity of VCP/p97 in ERAD.
    Biotechnology Letters 08/2013; 35(12). DOI:10.1007/s10529-013-1295-2 · 1.59 Impact Factor
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    • "Mammalian CDK5 plays a large number of roles in neural development (Su and Tsai, 2011), and it will be of interest to determine whether some CDK5 functions may be mediated by SAD regulation and whether other neurally expressed CDKs (e.g., PCTAIRE1) also contribute to SAD inhibition. An added complexity is that SAD-A has been reported capable of phosphorylating PCTAIRE1 (Chen et al., 2012). Our studies leave open the identity of the SAD ALT kinase important for sensory axon branching. "
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    ABSTRACT: Studies of brain-specific kinase 2 (BRSK2), an AMP-activated protein kinase (AMPK)-related kinase, and its homologs suggest that they are multifunctional regulators of cell-cycle progression. BRSK2, which contains a ubiquitin-associated (UBA) domain, is polyubiquitinated in cells. However, the regulatory mechanisms and exact biological function of BRSK2 remain unclear. Herein, we show that BRSK2 co-localizes with the centrosomes during mitosis. We also demonstrate that BRSK2 protein levels fluctuate during the cell cycle, peaking during mitosis and declining in G1 phase. Furthermore, Cdh1, rather than Cdc20, promotes the degradation of BRSK2 in vivo. Consistent with this finding, knock-down of endogenous Cdh1 blocks BRSK2 degradation during the G1 phase. The conserved KEN box of BRSK2 is required for anaphase-promoting complex/cyclosome-Cdh1 (APC/C(Cdh1))-dependent degradation. Additionally, overexpression of either BRSK2(WT) or BRSK2(ΔKEN) increases the percentage of cells in G2/M. Thus, our results provide the first evidence that BRSK2 regulates cell-cycle progression controlled by APC/C(Cdh1) through the ubiquitin-proteasome pathway.
    PLoS ONE 09/2012; 7(9):e45932. DOI:10.1371/journal.pone.0045932 · 3.23 Impact Factor
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