Introduction to the Symposium: The Origins and Evolution of Metabolic Pathways in Animals

Protein kinase D (PKD), a family of serine/threonine kinases, can be activated by a multitude of stimuli in a protein kinase C-dependent or -independent manner. PKD is involved in signal transduction pathways controlling cell proliferation, apoptosis, motility, and protein trafficking. Despite its versatile functions, few genuine in vivo substrates for PKD have been identified. In this study we demonstrate that the transcription factor cAMPresponse element-binding protein (CREB) is a direct substrate for PKD. PKD1 and CREB interact in cells, and activated PKD1 provokes CREB phosphorylation at Ser-133 both in vitro and in vivo. A constitutive active mutant of PKD1 stimulates GAL4-CREB-mediated transcription in a Ser-133-dependent manner, activates CRE-responsive promoters, and increases the expression of CREB target genes. PKD1 also enhances transcription mediated by two other members of the CREB family, ATF-1 and CREM. Our results describe a novel mechanism for PKD-induced signaling through activation of the transcription factor CREB and suggest that stimulus-induced phosphorylation of CREB, reported to be mediated by protein kinase C, may involve downstream activated PKD.The mammalian PKD 3 family of serine/threonine kinases includes the isoforms PKD1 (mouse PKD and human PKC), PKD2, and PKD3 (also named PKC). PKD was originally considered to be a member of the PKC family (1, 2) but is now classified in the calcium/calmodulin-dependent kinase group based on sequence similarities in the kinase domain (3). The PKDs share a similar architecture consisting of a C-terminal catalytic domain, an N-terminal regulatory domain that encompasses two cysteine-rich regions (C1a and C1b), and a pleckstrin homology (PH) domain (4 -7). A comparison of the amino acid sequences of PKD1-3 reveals that the highest homology lies in the catalytic domain, followed by C1a, C1b, and PH domain, suggesting that isoform-specific functions may be due to the regulatory part of the kinase (8).PKD1 can be activated by several mechanisms. The most studied mechanism involves a sequential activation of phospholipase C that results in the generation of the second messengers inositol 1,4,5-triphosphate and diacylglycerol (DAG) and subsequent activation of the classical (␣, ␤I, ␤II, and ␥) and novel (␦, ⑀, , and ) PKC isoforms. Binding of DAG to the C1b domain of PKD1 directs its translocation to the plasma membrane where activated novel PKC phosphorylates PKD1 at Ser-744/748 in the activation loop, causing activation of the enzyme. The activated enzyme can be imported via its C1b motif into the nucleus, where it transiently accumulates before being exported to the cytosol through a CRM1-dependent nuclear export pathway that requires the PH domain of PKD (4 -7). Mutations and deletions in the regulatory domain induce activation of PKD to various extents, and the entire regulatory domain has an inhibitory effect on the kinase activity (9). Indeed, PKD can also be activated through caspase-mediated cleavage of the regulatory domain (10). G␤␥ subunits ...

Section: Methodsmentioning

confidence: 99%

Protein Kinase D Induces Transcription through Direct Phosphorylation of the cAMP-response Element-binding Protein

Johannessen

Delghandi

Rykx

et al. 2007

“…No data are known on the involvement of phosphatases in the regulation of PRL signaling. To assess the possible role of endogenous tyrosine phosphatases in STAT5 activation, we treated mammary epithelial cells with sodium pervanadate (PV), a known protein-tyrosine phosphatase inhibitor (51). In these studies we employed the peroxy derivative of sodium vanadate, which penetrates intact cells more readily.…”

Section: Mgf Activation By Prl Is Affected By the Protein-tyrosine Kimentioning

confidence: 99%

Activation of STAT Factors by Prolactin, Interferon-γ, Growth Hormones, and a Tyrosine Phosphatase Inhibitor in Rabbit Primary Mammary Epithelial Cells

Schindler

Larose

et al. 1995

In numerous studies on mammary epithelial cell lines multiple factors, added to the medium or contained in the serum, were required for casein gene expression. It has been shown in these systems that the mammary gland factor (MGF) is implicated in the activation of the ␤-casein gene promoter. In the present study, we determined the relationship between known agents that affect casein gene expression and MGF activity using the properties of rabbit primary mammary epithelial cells to respond to PRL alone, when cultured in chemically defined medium. We demonstrate that MGF is rapidly activated by PRL alone or by human growth hormone, a natural ligand of many PRL receptors (PRL-Rs), in the cytoplasm and accumulated in the nucleus. The MGF activation by PRL occurred in the absence of endogenous extracellular matrix, a condition where casein synthesis is known to be markedly reduced. Different inhibitors of protein-tyrosine kinases, which have been shown to reduce casein mRNA synthesis, but not of protein kinase C, decrease the MGF activity. A tyrosine phosphatase inhibitor, sodium pervanadate, induced two GAS-binding complexes related to MGF and STAT1. Our data show that MGF is a latent cytoplasmic factor rapidly activated in mammary epithelial cells, by a mechanism involving a tyrosine kinase and a tyrosine phosphatase.

“…Protein-tyrosine phosphatases (PTPs) also play an important role in the attenuation of signals generated by protein-tyrosine kinases involved in mitogenesis. The extent and duration of protein-tyrosine kinase-mediated tyrosine phosphorylation can be greatly enhanced by treatment with the PTP inhibitor, sodium orthovanadate, or its peroxy derivatives (21,22). It has also been shown that overexpression of dual specificity MAPK-phospha-tase 1 (MKP-1) inhibits ERK2 activity and relieves the inhibitory effects of mitogens on muscle-specific gene expression (23).…”

mentioning

confidence: 99%

12-O-Tetradecanoylphorbol-13-acetate (TPA)-induced c-Jun N-terminal Kinase (JNK) Phosphatase Renders Immortalized or Transformed Epithelial Cells Refractory to TPA-inducible JNK Activity

Zhou¹,

Lin²,

Gu³

et al. 2000

Members of the mitogen-activated protein kinase (MAPK) 1 superfamily are proline-directed serine/threonine protein kinases that play pivotal roles in transducing various extracellular signals to the nucleus. They consist of three major subfamilies: MAPK/extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK)/stress-activated protein kinase (SAPK), and p38/Mpk2. MAPK/ERK is activated mainly by growth factors and phorbol esters and is associated with cellular proliferation and differentiation (1-3). JNK/SAPK and p38/ Mpk2 are activated by various extracellular stimuli, including growth factors, phorbol esters, UV irradiation and ionizing radiation, heat shock, hyperosmolarity, and cytotoxic drugs (3-6). Activation of these protein kinases leads to various responses, including gene expression, cell proliferation, differentiation, cell cycle arrest, apoptosis, early development, etc., depending on the cell type (7-11).MAPKs phosphorylate diverse target proteins in the membrane or cytosol as well as a number of nuclear transcription factors, indicating their critical role in orchestrating many short and long term changes in cell function (11,12). This has been confirmed recently using specific chemical inhibitors or by expressing mutant versions of different MAPKs or their upstream activators. These studies show that ERKs play a pivotal role in mediating neuronal differentiation in PC12 cells, as well as growth factor-stimulated proliferation and oncogenic transformation in fibroblasts (13,14). Similar approaches have yielded results supporting the view that JNK/SAPK and p38/ Mpk2 are critical in processes mediating platelet aggregation and secretion, in generation of inflammatory cytokines, and in pathways leading to apoptotic death in a number of cell types (15-18).Full activation of MAPKs requires phosphorylation of critical tyrosine and threonine residues. Several upstream dual specificity kinases catalyzing this modification have now been identified (1, 2, 9). Once activated, MAPKs phosphorylate and regulate several cellular proteins, including other protein kinases, cytoskeletal elements, stathmin, phospholipase A 2 , and transcription factors, notably Myc, Jun,19,20). The broad range of substrates indicates a pivotal role for MAPKs in cellular signal transduction, suggesting that the extent and duration of their activation play a key role in controlling cell function.A precise balance of the activities of protein kinases and protein phosphatases (PPs) plays a major role in receptormediated signaling pathways and cell cycle control. Several components of the ERK1/ERK2 pathway are subjected to regulation by PPs such as PP1 and PP2A. Protein-tyrosine phosphatases (PTPs) also play an important role in the attenuation of signals generated by protein-tyrosine kinases involved in mitogenesis. The extent and duration of protein-tyrosine kinase-mediated tyrosine phosphorylation can be greatly enhanced by treatment with the PTP inhibitor, sodium orthovanadate, or its peroxy derivatives (21, 22). It h...