The Snf1-related kinase, Hunk, is essential for mammary tumor metastasis.
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ABSTRACT:
SUBMITTER: Wertheim GB
PROVIDER: S-EPMC2734880 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:We previously identified a novel SNF1/AMPK-related protein kinase, Hunk, from a mammary tumor arising in an MMTV-neu transgenic mouse. The function of this kinase is unknown. Using targeted deletion in mice, we now demonstrate that Hunk is required for the metastasis of c-myc-induced mammary tumors, but is dispensable for normal development. Reconstitution experiments revealed that Hunk is sufficient to restore the metastatic potential of Hunk-deficient tumor cells, as well as defects in migration and invasion, and does so in a manner that requires its kinase activity. Consistent with a role for Hunk in the progression of human cancers, the human homologue of Hunk is overexpressed in aggressive subsets of carcinomas of the ovary, colon, and breast. In addition, a murine gene expression signature that distinguishes Hunk-wild type from Hunk-deficient mammary tumors predicts clinical outcome in women with breast cancer. Together, these findings establish a role for Hunk in metastasis and an in vivo function for this kinase. Hunk-deficient animals were crossed to mice harboring an MMTV-c-myc transgene (Leder et al., 1986). Hunk heterozygous, MMTV-c-myc mice were backcrossed to Hunk heterozygous animals. MMTV-c-myc female animals of each Hunk genotype were mated twice, then monitored twice weekly for mammary tumors. Mice possessing tumors with a maximum diameter of 20 mm were sacrificed and organs were examined at necropsy. Tumor nodules were identified by examination of organs through a Leica Wild MZ8 dissection microscope.
2009-12-31 | E-GEOD-14226 | biostudies-arrayexpress
Project description:We previously identified a novel SNF1/AMPK-related protein kinase, Hunk, from a mammary tumor arising in an MMTV-neu transgenic mouse. The function of this kinase is unknown. Using targeted deletion in mice, we now demonstrate that Hunk is required for the metastasis of c-myc-induced mammary tumors, but is dispensable for normal development. Reconstitution experiments revealed that Hunk is sufficient to restore the metastatic potential of Hunk-deficient tumor cells, as well as defects in migration and invasion, and does so in a manner that requires its kinase activity. Consistent with a role for Hunk in the progression of human cancers, the human homologue of Hunk is overexpressed in aggressive subsets of carcinomas of the ovary, colon, and breast. In addition, a murine gene expression signature that distinguishes Hunk-wild type from Hunk-deficient mammary tumors predicts clinical outcome in women with breast cancer. Together, these findings establish a role for Hunk in metastasis and an in vivo function for this kinase.
2009-12-31 | GSE14226 | GEO
Project description:Understanding the molecular pathways that contribute to the aggressive behavior of human cancers is a critical research priority. The SNF1/AMPK-related protein kinase Hunk is overexpressed in aggressive subsets of human breast, ovarian, and colon cancers. Analysis of Hunk(–/–) mice revealed that this kinase is required for metastasis of c-myc–induced mammary tumors but not c-myc–induced primary tumor formation. Similar to c-myc, amplification of the proto-oncogene HER2/neu occurs in 10%–30% of breast cancers and is associated with aggressive tumor behavior. By crossing Hunk(–/–) mice with transgenic mouse models for HER2/neu-induced mammary tumorigenesis, we report that Hunk is required for primary tumor formation induced by HER2/neu. Knockdown and reconstitution experiments in mouse and human breast cancer cell lines demonstrated that Hunk is required for maintenance of the tumorigenic phenotype in HER2/neu-transformed cells. This requirement is kinase dependent and resulted from the ability of Hunk to suppress apoptosis in association with downregulation of the tumor suppressor p27(kip1). Additionally, we find that Hunk is rapidly upregulated following HER2/neu activation in vivo and in vitro. These findings provide what we believe is the first evidence for a role for Hunk in primary tumorigenesis and cell survival and identify this kinase as an essential effector of the HER2/neu oncogenic pathway.
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Project description:SNF1-related kinase (SnRK1) in plants belongs to a conserved family that includes sucrose non-fermenting 1 kinase (SNF1) in yeast and AMP-activated protein kinase (AMPK) in animals. These kinases play important roles in the regulation of cellular energy homeostasis and in response to stresses that deplete ATP, they inhibit energy consuming anabolic pathways and promote catabolism. Energy stress is sensed by increased AMP:ATP ratios and in plants, 5'-AMP inhibits inactivation of phosphorylated SnRK1 by phosphatase. In previous studies, we showed that geminivirus pathogenicity proteins interact with both SnRK1 and adenosine kinase (ADK), which phosphorylates adenosine to generate 5'-AMP. This suggested a relationship between SnRK1 and ADK, which we investigate in the studies described here. We demonstrate that SnRK1 and ADK physically associate in the cytoplasm, and that SnRK1 stimulates ADK in vitro by an unknown, non-enzymatic mechanism. Further, altering SnRK1 or ADK activity in transgenic plants altered the activity of the other kinase, providing evidence for in vivo linkage but also revealing that in vivo regulation of these activities is complex. This study establishes the existence of SnRK1-ADK complexes that may play important roles in energy homeostasis and cellular responses to biotic and abiotic stress.
| S-EPMC3907550 | biostudies-literature
Project description:Epidermal growth factor receptor (EGFR) is commonly over-expressed in metastatic breast cancer yet metastatic breast cancer is generally resistant to anti-EGFR therapies, and the mechanism for resistance to EGFR inhibitors in this setting is not fully understood. Hormonally up-regulated neu-associated kinase (HUNK) kinase is up-regulated in aggressive breast cancers and is thought to play a role in breast cancer metastasis. However, no studies have been conducted to examine a relationship between EGFR and HUNK in breast cancer metastasis. We performed a kinase substrate screen and identified that EGFR is phosphorylated by HUNK. Our studies show that HUNK phosphorylates EGFR at T654, enhancing receptor stability and downstream signaling. We found that increased phosphorylation of T654 EGFR correlates with increased epithelial to mesenchymal, migration and invasion, and metastasis. In addition, we found that HUNK expression correlates with overall survival and distant metastasis free survival. This study shows that HUNK directly phosphorylates EGFR at T654 to promote metastasis and is the first study to show that the phosphorylation of this site in EGFR regulates metastasis.
| S-EPMC6989402 | biostudies-literature
Project description:Endocytosis is regulated in response to changing environmental conditions to adjust plasma membrane (PM) protein composition for optimal cell growth. Protein networks involved in cargo capture and sorting, membrane sculpting and deformation, and vesicle scission have been well-characterized, but less is known about the networks that sense extracellular cues and relay signals to trigger endocytosis of specific cargo. Hal4 and Hal5 are yeast Snf1-related kinases that were previously reported to regulate nutrient transporter stability by an unknown mechanism. Here we demonstrate that loss of Hal4 and Hal5 activates endocytosis of many different kinds of PM proteins, including Art1-mediated and Art1-independent endocytic events. Acute inhibition of Hal5 in the absence of Hal4 triggers rapid endocytosis, suggesting that Hal kinases function in a nutrient-sensing relay upstream of the endocytic response. Interestingly, Hal5 localizes to the PM, but shifts away from the cell surface in response to stimulation with specific nutrients. We propose that Hal5 functions as a nutrient-responsive regulator of PM protein stability, antagonizing endocytosis and promoting stability of endocytic cargos at the PM in nutrient-limiting conditions.
| S-EPMC7176151 | biostudies-literature
Project description:Members of the conserved SNF1/AMP-activated protein kinase (AMPK) family regulate cellular responses to environmental and nutritional stress in eukaryotes. Yeast SNF1 and animal AMPKs form a complex with regulatory SNF4/AMPKgamma and SIP1/SIP2/GAL83/AMPKbeta subunits. The beta-subunits function as target selective adaptors that anchor the catalytic kinase and regulator SNF4/gamma-subunits to their kinase association (KIS) and association with the SNF1 complex (ASC) domains. Here we demonstrate that plant SNF1-related protein kinases (SnRKs) interact with an adaptor-regulator protein, AKINbetagamma, in which an N-terminal KIS domain characteristic of beta-subunits is fused with a C-terminal region related to the SNF4/AMPKgamma proteins. AKINbetagamma is constitutively expressed in plants, suppresses the yeast delta snf4 mutation, and shows glucose-regulated interaction with the Arabidopsis SnRK, AKIN11. Our results suggest that evolution of AKINbetagamma reflects a unique function of SNF1-related protein kinases in plant glucose and stress signalling.
| S-EPMC1083798 | biostudies-literature
Project description:Ischaemic heart disease limits oxygen and metabolic substrate availability to the heart, resulting in tissue death. Here, we demonstrate that the AMP-activated protein kinase (AMPK)-related protein Snf1-related kinase (SNRK) decreases cardiac metabolic substrate usage and mitochondrial uncoupling, and protects against ischaemia/reperfusion. Hearts from transgenic mice overexpressing SNRK have decreased glucose and palmitate metabolism and oxygen consumption, but maintained power and function. They also exhibit decreased uncoupling protein 3 (UCP3) and mitochondrial uncoupling. Conversely, Snrk knockout mouse hearts have increased glucose and palmitate oxidation and UCP3. SNRK knockdown in cardiac cells decreases mitochondrial efficiency, which is abolished with UCP3 knockdown. We show that Tribbles homologue 3 (Trib3) binds to SNRK, and downregulates UCP3 through PPAR?. Finally, SNRK is increased in cardiomyopathy patients, and SNRK reduces infarct size after ischaemia/reperfusion. SNRK also decreases cardiac cell death in a UCP3-dependent manner. Our results suggest that SNRK improves cardiac mitochondrial efficiency and ischaemic protection.
| S-EPMC5286102 | biostudies-literature
Project description:c-Jun N-terminal kinase 2 (JNK2) isoforms are transcribed from the jnk2 gene and are highly homologous with jnk1 and jnk3 transcriptional products. JNK proteins mediate cell proliferation, stress response, and migration when activated by a variety of stimuli, including receptor tyrosine kinases (RTKs), but their ability to influence tumor metastasis is ill defined. To evaluate JNK2 in this manner, we used the highly metastatic 4T1.2 mammary tumor cells. Short hairpin RNA expression directed toward JNK2 (shJNK2) decreases tumor cell invasion. In vivo, shJNK2 expression slows tumor growth and inhibits lung metastasis. Subsequent analysis of tumors showed that shJNK2 tumors express lower GRB2-associated binding protein 2 (GAB2). In vitro, knockdown of JNK2 or GAB2 inhibits Akt activation by hepatocyte growth factor (HGF), insulin, and heregulin-1, while phosphorylation of ERK is constitutive and Src dependent. Knockdown of GAB2 phenocopies knockdown of JNK2 in vivo by reducing tumor growth and metastasis, supporting that JNK2 mediates tumor progression by regulating GAB2. The influence of jnk2 in the host or microenvironment was also evaluated using syngeneic jnk2-/- and jnk2+/+ mice. Jnk2-/- mice experience longer survival and less bone and lung metastasis compared to jnk2+/+ mice after intracardiac injection of 4T1.2 cells. GAB2 has previously been shown to mediate osteoclast differentiation, and osteoclasts are critical mediators of tumor-related osteolysis. Thus, studies focusing on the role of JNK2 on osteoclast differentiation were undertaken. ShJNK2 expression impairs osteoclast differentiation, independently of GAB2. Further, shJNK2 4T1.2 cells express less RANKL, a stimulant of osteoclast differentiation. Together, our data support that JNK2 conveys Src/phosphotidylinositol 3-kinase (PI3K) signals important for tumor growth and metastasis by enhancing GAB2 expression. In osteoclast progenitor cells, JNK2 promotes differentiation, which may contribute to the progression of bone metastasis. These studies identify JNK2 as a tumor and host target to inhibit breast cancer growth and metastasis.
| S-EPMC3111003 | biostudies-literature
Project description:The AMP-activated protein kinase (AMPK) is a major stress sensor of mammalian cells. AMPK's homolog in the yeast Saccharomyces cerevisiae, the SNF1 protein kinase, is a central regulator of carbon metabolism that inhibits the Snf3/Rgt2-Rgt1 glucose sensing pathway and activates genes involved in respiration. We present evidence that glucose induces modification of the Snf1 catalytic subunt of SNF1 with the small ubiquitin-like modifier protein SUMO, catalyzed by the SUMO (E3) ligase Mms21. Our results suggest that SUMOylation of Snf1 inhibits its function in two ways: by interaction of SUMO attached to lysine 549 with a SUMO-interacting sequence motif located near the active site of Snf1, and by targeting Snf1 for destruction via the Slx5-Slx8 (SUMO-directed) ubiquitin ligase. These findings reveal another way SNF1 function is regulated in response to carbon source.
| S-EPMC3808588 | biostudies-literature