Protein kinase C (PKC)-induced PKC degradation: a model for down-regulation

140

128

waf1/cip1 and p27 kip1 . Reversal of these cell cycle regulatory effects was coincident with activator-induced down-regulation of PKC ␣, ␦, and ⑀. Differential down-regulation of individual PKC isozymes revealed that PKC ␣ in particular is sufficient to mediate cell cycle arrest by PKC agonists in this system. Taken together, the data implicate PKC ␣ in negative regulation of intestinal epithelial cell growth both in vitro and in situ via pathways which involve modulation of Cip/Kip family cyclin-dependent kinase inhibitors and the retinoblastoma growth suppressor protein.

Section: P21supporting

confidence: 91%

Protein Kinase C Isozyme-mediated Cell Cycle Arrest Involves Induction of p21 and p27 and Hypophosphorylation of the Retinoblastoma Protein in Intestinal Epithelial Cells

Frey

Saxon

Zhao

et al. 1997

140

128

“…Members of the PKC family undergo down-regulation following activation (33). In IEC-18 cells, PMA treatment results in progressive down-regulation of phorbol ester-responsive PKC isozymes, albeit with different kinetics (Fig.…”

Section: Loss Of Pkc/pkc␣ Signaling Results In Recovery Of the Translmentioning

confidence: 99%

Protein Kinase C α Signaling Inhibits Cyclin D1 Translation in Intestinal Epithelial Cells

Hizli¹,

Black²,

Pysz³

et al. 2006

Cyclin D1 is a key regulator of cell proliferation, acting as a mitogen sensor and linking extracellular signaling to the cell cycle machinery. Strict control of cyclin D1 levels is critical for maintenance of tissue homeostasis. We have reported previously that protein kinase C ␣ (PKC␣) Tight control of cell proliferation is essential for maintenance of normal homeostasis in self-renewing tissues such as the intestinal epithelium. Previous studies have identified protein kinase C (PKC) 4 signaling as an important negative regulator of cell growth/cell cycle progression in intestinal epithelial cells (1-3). Several members of the PKC family are predominantly expressed/activated in non-proliferating and terminally differentiated intestinal cells (4 -6), pointing to a major function in regulation of post-mitotic events in this tissue. Consistent with this role, we have demonstrated that PKC␣ signaling triggers a program of cell cycle withdrawal in non-transformed intestinal crypt cells, paralleling the membrane translocation/activation of this enzyme precisely at the point of growth arrest within intestinal crypts in situ (3,4,7,8).,Although the extracellular signals that trigger growth arrest in the intestine in situ remain poorly characterized, a possible physiological activator of PKC␣ in this system is transforming growth factor ␤. This potent growth inhibitory factor is known to promote G 0 /G 1 arrest in intestinal epithelial cells (9 -11) and to activate PKC␣ signaling in other systems (12).One of the earliest events following PKC␣ activation in intestinal epithelial cells is down-regulation of cyclin D1 (3,7,8), indicating that this molecule is an important target of PKC␣ control. Cyclin D1 is a key regulator of cell proliferation, acting as a mitogen sensor and linking extracellular signaling to the cell cycle machinery (13). Progression through early G 1 involves the activity of holoenzymes consisting of cyclin D (D1, D2, or D3 depending on the cell type) in association with cdk4 or cdk6. Cyclin D-dependent kinases promote cell cycle progression by initiating phosphorylation/inactivation of the retinoblastoma growth suppressor protein in mid G 1 , a process that is completed later in G 1 by cyclin E/cdk2. Cyclin D/cdk complexes also have an important non-catalytic function that involves sequestration of cdk-inhibitory proteins of the Cip/Kip family, thus relieving repression of cyclin E-and cyclin A-cdk2 activity.Precise regulation of cyclin D1 accumulation is of critical importance. Increased expression of the molecule shortens the G 1 interval in many cell types and can reduce/overcome dependence on physiological growth stimuli (14). Decreased levels of the protein, on the other hand, lengthen G 1 phase and reduce proliferation (14, 15). Thus, cyclin D1 expression is subject to strict control at multiple levels, including transcription, message stability and nucleocytoplasmic transport, protein synthesis, and protein turnover (14, 16 -21). Notably, aberrant overexpression of cyclin D1 is a key componen...

“…However, the effect of nPKC isoform inhibitors to slow the kinetics of PKC␣ and PKC␦ downregulation (i.e. to regulate PKC␣ in trans) suggests a more general role for nPKC kinase activity in the events that govern PKC trafficking/down-regulation (20,21).…”

Section: Kinases That Mediate Npkc Isoform Phosphorylation-mentioning

confidence: 99%

Cross-regulation of Novel Protein Kinase C (PKC) Isoform Function in Cardiomyocytes

Rybin¹,

Sabri²,

Short³

et al. 2003

103

Recent studies identify conventional protein kinase C (PKC) isoform phosphorylations at conserved residues in the activation loop and C terminus as maturational events that influence enzyme activity and targeting but are not dynamically regulated by second messengers. In contrast, this study identifies phorbol 12-myristoyl 13-acetate (PMA)-and norepinephrine-induced phosphorylations of PKC⑀ (at the C-terminal hydrophobic motif) and PKC␦ (at the activation loop) as events that accompany endogenous novel PKC (nPKC) isoform activation in neonatal rat cardiomyocytes. Agonist-induced nPKC phosphorylations are prevented (and the kinetics of PMAdependent PKC down-regulation are slowed) by pharmacologic inhibitors of nPKC kinase activity. PKC␦ is recovered from PMA-treated cultures with increased in vitro lipid-independent kinase activity (and altered substrate specificity); the PMA-dependent increase in PKC␦ kinase activity is attenuated when PKC␦ activation loop phosphorylation is prevented. To distinguish roles of individual nPKC isoforms in nPKC phosphorylations, wild-type (WT) and dominant negative (DN) PKC␦ and PKC⑀ mutants were introduced into cardiomyocyte cultures using adenovirus-mediated gene transfer. WT-PKC␦ and WT-PKC⑀ are highly phosphorylated at activation loop and hydrophobic motif sites, even in the absence of allosteric activators. DN-PKC␦ is phosphorylated at the activation loop but not the hydrophobic motif; DN-PKC⑀ is phosphorylated at the hydrophobic motif but not the activation loop. Collectively, these results identify a role for PKC⑀ in nPKC activation loop phosphorylations and PKC␦ in nPKC hydrophobic motif phosphorylations. Agonist-induced nPKC isoform phosphorylations that accompany activation/translocation of the enzyme contribute to the regulation of PKC␦ kinase activity, may influence nPKC isoform trafficking/down-regulation, and introduce functionally important cross-talk for nPKC signaling pathways in cardiomyocytes. Protein kinase C (PKC)1 constitutes a family of serine/threonine kinases that play key roles in signal transduction by receptors coupled to phosphoinositide hydrolysis (1). The basic primary structure of all PKC isozymes consists of a single polypeptide chain with N-terminal regulatory and C-terminal catalytic domains. PKC isoforms are subdivided into three distinct subfamilies based upon structural differences in their N-terminal regulatory domains. Conventional PKC isoforms (cPKCs) contain an autoinhibitory pseudosubstrate domain followed by two membrane-targeting modules, the C1 domain that binds diacylglycerol and phosphatidylserine (PS) and the C2 domain that binds anionic lipids in a calcium-dependent manner. Novel PKC (nPKC) isoforms lack the C2 domain and are maximally activated by diacylglycerol/phorbol ester in the absence of calcium; atypical PKCs are regulated by lipids but are not activated by calcium and diacylglycerol. Current models of PKC isoform activation focus largely on the allosteric changes that are induced by cofactor binding to N-terminal membrane-t...