Amy K. Rines

Snf1-related kinase inhibits colon cancer cell proliferation through calcyclin-binding protein-dependent reduction of β-catenin

Sucrose nonfermenting 1 (Snf1)‐related kinase (SNRK) is a serine/threonine kinase with sequence similarity to AMP‐activated protein kinase (AMPK); however, its function is not well characterized. We conducted a gene array to determine which genes are regulated by SNRK. The array demonstrated that SNRK overexpression increased the levels of genes involved in cell proliferation, including calcyclin‐binding protein (CacyBP), a member of the ubiquitin ligase complex that targets nonphosphorylated β‐catenin for degradation. We confirmed that SNRK increased CacyBP mRNA and protein, and decreased β‐catenin protein in HCT116 and RKO colon cancer cells. Furthermore, SNRK inhibited colon cancer cell proliferation, and CacyBP down‐regulation reversed the SNRK‐mediated decrease in proliferation and β‐catenin. SNRK overexpression also decreased β‐catenin nuclear localization and target gene transcription, and β‐catenin down‐regulation reversed the effects of SNRK knockdown on proliferation. SNRK transcript levels were reduced in human colon tumors compared to normal tissue by 35.82%, and stable knockdown of SNRK increased colon cancer cell tumorigenicity. Our results demonstrate that SNRK is down‐regulated in colon cancer and inhibits colon cancer cell proliferation through CacyBP up‐regulation and β‐catenin degradation, resulting in reduced proliferation signaling. These findings reveal a novel function for SNRK in the regulation of colon cancer cell proliferation and β‐catenin signaling.—Rines, A. K., Burke, M. A., Fernandez, R. P., Volpert, O. V., Ardehali, H. Snf1‐related kinase inhibits colon cancer cell proliferation through calcyclin binding protein‐dependent reduction of β‐catenin. FASEB J. 26, 4685–4695 (2012). www.fasebj.org

A new pROM king for the mitoK(ATP) dance: ROMK takes the lead.

Ischemic heart disease remains the leading cause of death in the developed world, and mechanisms to reduce cardiac ischemic damage are being actively investigated. A potent protective mechanism for the heart is ischemic preconditioning (IPC), a phenomenon in which brief ischemic episodes protect the heart from tissue damage and cell death resulting from subsequent periods of ischemia and reperfusion.1 The precise molecular mechanisms at play during IPC are not known, but the opening of a mitochondrial ATP-sensitive potassium channel, mitoKATP, is believed to be necessary for IPC-induced activation of several prosurvival pathways and processes.2 Article, see p 446 Although the discovery of a putative mitoKATP occurred more than 20 years ago,3 progress on identifying its molecular composition has been limited. The mitoKATP was determined to be both functionally and molecularly distinct from sarcolemmal KATP channels, which have a relatively minimal role in IPC protection and are insensitive to several drugs that affect the mitoKATP.4, 5 Initial studies using immunoreactivity identified the inward rectifying K+-channel subunit Kir6.1 as localizing to mitochondria,6,7 and thus Kir6.1 became an attractive candidate as a possible subunit of the mitoKATP. However, further research by mass spectrometry into the specificity of the Kir6.1 antibody revealed that the antibody does not recognize Kir6.1, and that Kir6.1 is not isolated from more thorough proteomic screens of mitochondria.8 Another investigation led to purification of an inward-rectifying K+-channel component of …

Phosphorylation of contractile proteins in response to α- and β-adrenergic stimulation in neonatal cardiomyocytes

alpha- and beta-Adrenergic receptor agonists induce an inotropic response in the adult heart by promoting the phosphorylation of several regulatory proteins, including myosin-binding protein C (MyBP-C), cardiac troponin I (cTnI), and phospholamban (PLB). However, the adrenergic-induced phosphorylation of these proteins has not been characterized in the developing heart. Accordingly, we evaluated MyBP-C, cTnI, and PLB phosphorylation in cultured neonatal rat cardiomyocytes (NRCMs) after alpha- and beta-receptor activation with phenylephrine and isoproterenol. alpha-Receptor stimulation increased, whereas beta-receptor activation reduced MyBP-C phosphorylation. Isoelectric-focusing experiments indicated that the amount of monophosphorylated MyBP-C was sensitive to alpha-adrenergic activation, but diphosphorylated and triphosphorylated MyBP-C levels were largely unaffected. The phosphorylation of cTnI and PLB was consistent with the mechanism observed in adult hearts: alpha- and beta-Receptor stimulation phosphorylated both proteins. For cTnI, the greatest difference associated with beta-receptor activation was observed in the diphosphorylated state, whereas alpha-receptor activation was associated with a marked increase in the tetraphosphorylated protein and absence of the unphosphorylated state. Despite these apparent changes in cTnI and PLB phosphorylation, beta-receptor activation failed to alter calcium transients in NRCMs. Collectively, these findings suggest that, unlike cTnI and PLB, MyBP-C and inotropy are not coupled to beta-adrenergic stimulation in NRCMs. Therefore, cTnI and PLB probably play a more central role in modulating contractile function in NRCMs in response to catecholamines than does MyBP-C, and MyBP-C may have a structural role in stabilizing thick filament assembly rather than influencing cross-bridge formation in developing hearts.

Corrigendum: Snf1-related kinase improves cardiac mitochondrial efficiency and decreases mitochondrial uncoupling

This corrects the article DOI: 10.1038/ncomms14095.